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Unformatted text preview: Psychology 335 Behavioral Neuroendocrinology Fall 2008 George Wade 545-0772 gwade@cns.umass.edu 2 - Behavioral Neuroendocrinology CONTENTS Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Structure and function of the Nervous System . . . . . . . . . . . . . Endocrine Physiology . . . . . . . . . . . . . . . . . . . . . . . . . . Sexual Differentiation . . . . . . . . . . . . . . . . . . . . . . . . . . 5 21 45 85 Male Copulatory Behavior . . . . . . . . . . . . . . . . . . . . . . . 123 Female Copulatory Behavior . . . . . . . . . . . . . . . . . . . . . . 151 Parental Behavior . . . . . . . . . . . . . . . . . . . . . . . . . . . . 193 Affiliation and Aggression . . . . . . . . . . . . . . . . . . . . . . . . 219 Hunger and Energy Balance . . . . . . . . . . . . . . . . . . . . . . 255 Nutrition and Fertility . . . . . . . . . . . . . . . . . . . . . . . . . . 307 Biological Rhythms and Seasonal Reproduction . . . . . . . . . . . . 341 Stress . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 365 Thirst . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 389 List of Acronyms . . . . . . . . . . . . . . . . . . . . . . . . . . . . 409 Behavioral Neuroendocrinology - 3 4 - Behavioral Neuroendocrinology Introduction - 1 Introduction Behavioral Neuroendocrinology - 5 Introduction - 2 6 - Behavioral Neuroendocrinology Introduction - 3 Psychology 335 – Behavioral Neuroendocrinology Who I am: George Wade 525 Tobin Hall gwade@cns.umass.edu (Don’t use SPARK.) 545-0772 Office hours: Pretty much any time. Behavioral Neuroendocrinology - 7 Introduction - 4 How the course works Grading: • Exams are short answer – a few words to a paragraph – no multiple choice. • Exam questions will come only from class lectures, unless I tell you otherwise. • There will be an evening review session before each exam. • This course will not accept research participation credits. 8 - Behavioral Neuroendocrinology Introduction - 5 Academic honesty: All students are expected to adhere scrupulously to the University policy concerning academic honesty. For more information on the University’s academic honesty policy, check this web site: www.umass.edu/dean_students/code_conduct/acad_honest.htm Attendance: You are expected to attend class. If you miss a class for some reason, you are responsible for getting the material. To encourage you to attend class, I will discuss some material in each lecture that will not be posted online. It will be included on the exams for extra credit. Reading materials: Randy S. Nelson. An Introduction to Behavioral Endocrinology, third edition, Sinauer, 2005. (Second edition is OK, too. Page numbers for readings in syllabus and posted online.) Other readings available for download on SPARK. Behavioral Neuroendocrinology - 9 Introduction - 6 Useful advice: There will be an optional course pack for this class for sale at the Textbook Annex. It contains printouts of all of the slides. It is also available for download on SPARK, so you do not have to purchase it. The course pack is not (not, not, not) an alternative to coming to class. Much of what I say in class is not included in the course pack. I will also post the slides for each lecture as PowerPoint files online at SPARK. 10 - Behavioral Neuroendocrinology Introduction - 7 Useful advice: Don’t get behind. Go over the material after class and make sure you understand what I said. You don’t have to commit it to memory at that time. But if you understand the material first, studying for the exam will be much easier. Don’t wait until the last minute to figure out how something works. Useful advice: Ask questions in class: I welcome questions in class. If you don’t understand something, chances are a bunch of others don’t get it either, and you’ll be doing them a favor by asking. Come see me for help: If you find that don’t understand something after class, come see me for help. After class is a good time, but it can be kind of hectic. The best way to get help is to call or e-mail me to set up a time to talk. That way, you can be sure I’ll be there. My schedule is pretty flexible. Behavioral Neuroendocrinology - 11 Introduction - 8 So what’s behavioral neuroendocrinology, anyway? Actually, it’s just a pretentious name for what’s used to be called ‘hormones and behavior.’ There are two major communications systems in the body. • Neuroscientists study the workings of the nervous system. • Endocrinologists study the workings of the various glands in the body. • Neuroendocrinologists study how these two systems interact with one another. 12 - Behavioral Neuroendocrinology Introduction - 9 Neural and endocrine systems have both similarities and differences. Similarities: 1. Both use chemicals to communicate among cells. • The nervous system uses chemicals called neurotransmitters. • The endocrine system uses chemicals called hormones. • Some chemicals can act as either hormones or neurotransmitters, depending on what kind of cell releases them. 2. Both neurotransmitters and hormones act via receptors on or in the target cells. Neural and endocrine systems have both similarities and differences. Differences: 1. Hormones and neurotransmitters act over very different distances. • Neurotransmitters act over very short distances – synapses. • Hormones are secreted directly into the bloodstream and are carried to distant targets. Behavioral Neuroendocrinology - 13 Introduction - 10 Neural and endocrine systems have both similarities and differences. Differences: 2. Hormones and neurotransmitters act with different latencies and durations. • Neurotransmitters typically act instantaneously and for a short duration. • Hormones typically act more slowly, because it takes time to travel through the bloodstream. Hormone action may be prolonged – until it is cleared from the bloodstream. Neural and endocrine systems have both similarities and differences. Differences: 3. Differences in voluntary (conscious) control. • We have voluntary control over many, but not all, neural signals. • On the other hand, we generally can’t consciously control hormone secretion. 14 - Behavioral Neuroendocrinology Introduction - 11 Interactions between hormones and behavior are reciprocal – each affects the other. Hormones behavior – examples??? female sexual behavior. male sexual behavior. food intake. arousal and alertness. parental behavior. • Estrogens (ovaries) • Androgens (testes) • Insulin (pancreas) • Epinephrine (adrenal) • Oxytocin (posterior pituitary) Hormones almost always act directly on the brain in order to alter behavior, but there can be some exceptions (insulin, for example). Interactions between hormones and behavior are reciprocal – each affects the other. Behavior hormones – examples??? reflex ovulation. epinephrine. insulin secretion. oxytocin and milk letdown. oxytocin and milk letdown. • Mating stimulation • Fear or surprise • Smell of food • Nipple stimulation • Sound of a crying baby • Sporting events testosterone secretion. Behavioral Neuroendocrinology - 15 Introduction - 12 Sometimes sequences of hormone interactions can be quite complex. The mating of shrews is an example: anestrous smells activates brain, pituitary and ovaries behavior ’s hormones act on brain mates with ’s penis stimulates ’s vagina ovulates Endocrine manipulations are quite common in our society and have a long history – thousands of years. Castration of male animals: Why do it? • Easy to do • Positive effects on behavior • May improve physical properties • We don’t need a whole bunch of males anyway – females are bottleneck for reproduction 16 - Behavioral Neuroendocrinology Introduction - 13 Everyday example. “Ha, ha, ha, Biff. Guess what? After we go to the drugstore and the post office, I’m going to the vet’s to get tutored.” Actually, there’s a long history of castrating men – eunuchs • Punishment • “Testosterone-sensitive” jobs – guarding the harem • Castrati – male sopranos – until late 1700’s • Over a million eunuchs in contemporary India - Hijras Guarding the Sultan’s harem Carlo Broschi Farinelli (1705-1782) Contemporary Indian Hijras (HIJ-ruhs) Behavioral Neuroendocrinology - 17 Introduction - 14 First experimental endocrinology done by A.A. Berthold in 1840’s Caponization of roosters appearance red comb, colorful plumage pale comb, bland plumage behavior crows, struts, fights, mates nada + red comb, colorful plumage crows, struts, fights, mates This means that the presence of testes is responsible for these male-like characteristics. (Hormonal or neural?) Topics we’ll cover: 1. How the nervous system works. 2. The basics of endocrine function. 3. Sexual differentiation. 4. Male copulatory behavior. 5. Female copulatory behavior. 6. Parental behavior. 7. Social behaviors – affiliation and aggression. 8. Food intake and energy balance. 9. Interactions between energy balance and reproduction. 10. Rhythms and seasonal breeding. 11. Stress. 18 - Behavioral Neuroendocrinology Introduction - 15 Some of the topics we won’t cover: 1. 2. 3. 4. 5. 6. Hormones and mood. Neuroimmunology. Hormones and cognitive function. Hormones and sensory function. Hormones and thirst – maybe. Un-mammals. Behavioral Neuroendocrinology - 19 Introduction - 16 20 - Behavioral Neuroendocrinology Structure and Function of the Nervous System - 1 Structure and Function of the Nervous System Behavioral Neuroendocrinology - 21 Structure and Function of the Nervous System - 2 22 - Behavioral Neuroendocrinology Structure and Function of the Nervous System - 3 Everything you ever wanted to know about the nervous system in two easy lessons. Structure of an idealized nerve cell (neuron) (glial cells) bifurcation Behavioral Neuroendocrinology - 23 Structure and Function of the Nervous System - 4 The contacts between neurons are called synapses The “innards” of the soma: Cytoplasm and organelles. (ATP) 24 - Behavioral Neuroendocrinology Structure and Function of the Nervous System - 5 Four different types of synapses 1. Synapse on smooth dendrite – axodendritic synapse 2. Synapse on dendritic spine – axodendritic synapse 3. Synapse on soma (cell body) – axosomatic synapse 4. Synapse on terminal button from another cell – axoaxonic synapse Structure of a synapse Neurotransmitter being released into synaptic cleft Behavioral Neuroendocrinology - 25 Structure and Function of the Nervous System - 6 Synaptic vesicle presynaptic neuron postsynaptic neuron 26 - Behavioral Neuroendocrinology Structure and Function of the Nervous System - 7 Electrical currents in neuronal membranes are caused by the movement of electrically charged atoms (ions) in and out of the cell – through ion channels. Na+ K+ Some neurotransmitters bind to receptor sites on the outside end of ion channels and cause them to open or close. Behavioral Neuroendocrinology - 27 Structure and Function of the Nervous System - 8 Sequential opening and closing of ion channels change the electrical potential across the neuronal cell membrane and generate an action potential. The axon hillock integrates excitatory postsynaptic potentials (EPSP’s) and inhibitory postsynaptic potentials (IPSP’s) and “decides” whether or not an action potential will be generated. Synapses closer to axon hillock have more impact than those farther away. Activity of inhibitory synapses produces IPSPs (blue) in postsynaptic neuron axon hillock Activity of excitatory synapses produces EPSPs (red) in postsynaptic neuron Axon hillock reaches threshold of excitation; action potential is triggered in axon IPSPs counteract EPSPs; action potential is not triggered in axon 28 - Behavioral Neuroendocrinology Structure and Function of the Nervous System - 9 size of action potential doesn’t diminish Action potentials moving down the axon are ‘all-or-nothing.’ There aren’t big ones or little ones – they’re all the same size. Information about the intensity of a stimulus is coded by the frequency of the action potentials. Behavioral Neuroendocrinology - 29 Structure and Function of the Nervous System - 10 Synaptic transmission: summary Three ways to turn off synaptic transmission: 1. Destruction of the neurotransmitter in the synaptic cleft. 2. Reuptake of the neurotransmitter by the presynaptic neuron. 3. Autoreceptors on the presynaptic membrane inhibit further neurotransmitter release. 30 - Behavioral Neuroendocrinology Structure and Function of the Nervous System - 11 Enzymes are biological catalysts. They are proteins (usually), and they determine which chemical reactions take place in the body. Agonists and antagonists Agonists are chemicals that mimic the effects of a neurotransmitter or hormone. Antagonists are chemicals that block the effects of a neurotransmitter or hormone by blocking the binding site. Agonists and antagonists can act either directly or indirectly. occupies binding site without having any biological effect binds to a different site but prevents agonist from having its effect at other site Behavioral Neuroendocrinology - 31 Structure and Function of the Nervous System - 12 Gross structure of the nervous system. Where’s where. 32 - Behavioral Neuroendocrinology Structure and Function of the Nervous System - 13 Transverse (frontal) Horizontal Saggital Behavioral Neuroendocrinology - 33 Structure and Function of the Nervous System - 14 Subdivisions of the brain. Top view: Sagittal view: Notice that the brain is hollow – filled with cerebrospinal fluid. 34 - Behavioral Neuroendocrinology Structure and Function of the Nervous System - 15 The brain is hollow; it contains fluid-filled ventricles Cerebrospinal fluid circulates through the cerebral ventricles – front to back. Flow of CSF: front to back Behavioral Neuroendocrinology - 35 Structure and Function of the Nervous System - 16 (2) The hypothalamus and pituitary gland(s). 36 - Behavioral Neuroendocrinology Structure and Function of the Nervous System - 17 Cranial nerves: The autonomic nervous system controls the functioning of your innards It consists of two opposing parts: • Sympathetic nervous system • Parasympathetic nervous system The autonomic nervous system controls the function of some important glands. Behavioral Neuroendocrinology - 37 Structure and Function of the Nervous System - 18 sympathetic parasympathetic vagus n. autonomic nerves also carry sensory information to the brain Some techniques for studying how the brain works. 38 - Behavioral Neuroendocrinology Structure and Function of the Nervous System - 19 Stereotaxic instrument Landmarks used to find specific sites in the brain. Behavioral Neuroendocrinology - 39 Structure and Function of the Nervous System - 20 A stereotaxic atlas is a map of the brain. 1.1 mm anterior to bregma dorsomedial hypothalamus 0 6 4 2 0 2 distance from midline (mm) 4 6 -2 depth below bregma (mm) -4 From Bregma: -7.5 mm down ±1.0 mm lateral 1.1 mm anterior -6 -8 -10 40 - Behavioral Neuroendocrinology Structure and Function of the Nervous System - 21 Stereotaxic instrument for the human brain. Electrodes (fine metal wires) can be implanted in the brain for use in electrical stimulation or recording or making lesions. 1. Electrical recording 2. Electrical stimulation 3. Lesioning Behavioral Neuroendocrinology - 41 Structure and Function of the Nervous System - 22 Electrodes are fastened to the skull using jeweler’s screws and dental cement. Bregma Cannulas (fine hypodermic tubes) can be implanted in the brain in order to infuse substances directly into the brain. 42 - Behavioral Neuroendocrinology Structure and Function of the Nervous System - 23 Tracers can be used to plot the connections of a particular area of the brain. AAA BBB VMH retrograde tracer (taken up by terminal buttons) CCC DDD anterograde tracer (taken up by soma) Tract tracing. Anterograde tracing: soma → terminals Retrograde tracing: terminals → soma PHA-L is an anterograde tracer. Behavioral Neuroendocrinology - 43 Structure and Function of the Nervous System - 24 44 - Behavioral Neuroendocrinology Endocrine Physiology - 1 Endocrine Physiology Behavioral Neuroendocrinology - 45 Endocrine Physiology - 2 46 - Behavioral Neuroendocrinology Endocrine Physiology - 3 A brief tour of your endocrine glands and how they work. Types of chemical signaling: Intracrine: chemical signal within a cell Autocrine: secreted chemical acts on same cell - autoreceptors Paracrine: secreted chemical acts on neighbors – like synaptic transmission Endocrine: secreted chemical carried by blood Exocrine: chemical secreted into ducts Paracrine (neurotransmitters) Endocrine (hormones) Behavioral Neuroendocrinology - 47 Endocrine Physiology - 4 There are three kinds of hormones: Amines Steroids Peptides There are three principal kinds of hormones: Amines are derivatives of single amino acids. Examples include epinephrine, and melatonin. Sources include adrenal medulla and pineal. each bend is a carbon (C) atom amino groups epinephrine from the adrenal medulla derived from tyrosine melatonin from the pineal gland derived from tryptophan 48 - Behavioral Neuroendocrinology Endocrine Physiology - 5 There are three principal kinds of hormones: Steroids are all derivatives of cholesterol. Examples include progesterone, testosterone, and cortisol. Sources are ovaries, testes, and adrenal cortex. C A B D basic steroid structure (4 rings, 17 carbons) cholesterol There are three principal kinds of hormones: Peptide and protein hormones are chains of amino acids. Examples include insulin and luteinizing hormone. Sources include the pancreas and the pituitary gland. ● Peptides are short chains, ≈ 50 amino acids or fewer ● Proteins are longer chains of amino acids Behavioral Neuroendocrinology - 49 Endocrine Physiology - 6 Proteins and peptides are made up of chains of amino acids side chain amino acid Formation of a peptide bond amino acid - H2O amino acid dipeptide 50 - Behavioral Neuroendocrinology Endocrine Physiology - 7 H2N HOOC Protein structure can be enormously complicated. This diversity allows proteins to have so many functions. Different ways of representing the 3-D structure of proteins. Behavioral Neuroendocrinology - 51 Endocrine Physiology - 8 Your Glands: Top to Bottom (mostly) The hypothalamus and pituitary gland(s) are the two most important endocrine structures in the body (from a neuroendocrinologist’s point of view) 52 - Behavioral Neuroendocrinology Endocrine Physiology - 9 Actually, the pituitary gland is two separate glands right next to each other – the anterior pituitary and the posterior pituitary. Control of the anterior pituitary hypophyseal portal vein (releasing hormones) (tropic hormones) Portal vein: a blood vessel that joins two capillary beds Behavioral Neuroendocrinology - 53 Endocrine Physiology - 10 Control of the posterior pituitary The posterior pituitary is actually part of the brain; the anterior pituitary is not. The anterior pituitary gland controls the thyroid, the adrenal cortex, the ovaries, and the testes. cortex 54 - Behavioral Neuroendocrinology Endocrine Physiology - 11 (posterior) Other glands are controlled by direct innervation. (the medulla) Behavioral Neuroendocrinology - 55 Endocrine Physiology - 12 In the human brain, the pineal gland is buried deep under the cerebral cortex. In lower vertebrates like lizards the pineal is a third eye and responds to directly to light and dark. In mammals it in not directly sensitive to light. Instead, it gets information about light and dark via neural projections from the eyes. The pineal plays an important role in seasonal reproduction by secreting the amine, melatonin. melatonin 56 - Behavioral Neuroendocrinology Endocrine Physiology - 13 The pathway from photoreceptors in the eye to the pineal gland is circuitous in mammals. NE sympathetic nerve (superior cervical ganglion) (posterior) Behavioral Neuroendocrinology - 57 Endocrine Physiology - 14 The posterior pituitary releases two peptide hormones, oxytocin and vasopressin. Oxytocin causes milk letdown during nursing and causes uterine contractions during labor and orgasms. Vasopressin, also known as antidiuretic hormone, plays an important role in maintaining water balance. Both peptides affect social and reproductive behaviors by acting as neurotransmitters. Each one has just 9 amino acids. oxytocin vasopressin 58 - Behavioral Neuroendocrinology Endocrine Physiology - 15 The thyroid has multiple effects, particularly on metabolic rate. Thyroid hormone is extremely important for normal neural development - cretinism thyroid hormone (a.k.a. T4, thyroxine) http://www.nytimes.com/2006/12/16/health/16iodine.html Behavioral Neuroendocrinology - 59 Endocrine Physiology - 16 Thyroid hormone is actually a prohormone. thyroid hormone (a.k.a. T4, thyroxine) deiodination triiodothyronine (a.k.a. T3) (the more active form) Secretion of thyroid hormone is controlled by a negative feedback system involving the hypothalamus, anterior pituitary gland and the thyroid. 60 - Behavioral Neuroendocrinology Endocrine Physiology - 17 Negative feedback in endocrine systems acts much like the thermostat and furnace in your house negative feedback (–) Thermostat air ducts wiring Bedroom Heat Living room Bathroom Kitchen (+) Furnace Negative feedback in endocrine systems acts much like the thermostat and furnace in your house negative Hypothalamus feedback (–) portal vein (+) bloodstream releasing hormone Anterior pituitary bloodstream Nose Hormone Muscles Brain Liver (+) Gland tropic hormone Behavioral Neuroendocrinology - 61 Endocrine Physiology - 18 Releasing hormones: are produced in the hypothalamus are secreted into the hypophyseal portal system stimulate tropic hormone secretion by the anterior pituitary Negative feedback in the HPT axis: Hypothalamus (-) thyrotropin-releasing hormone (TRH) triiodothyronine – T3 (-) Anterior pituitary thyroid-stimulating hormone (TSH, or thyrotropin) Thyroid 62 - Behavioral Neuroendocrinology Endocrine Physiology - 19 What if the thyroid can’t produce T3 or T4? Hypothalamus (-) thyrotropin-releasing hormone (TRH) triiodothyronine – T3 (-) Anterior pituitary thyroid-stimulating hormone (TSH, or thyrotropin) Thyroid Goiter due to thyroid insufficiency Behavioral Neuroendocrinology - 63 Endocrine Physiology - 20 Parathyroid hormone controls calcium metabolism 64 - Behavioral Neuroendocrinology Endocrine Physiology - 21 The pancreas Behavioral Neuroendocrinology - 65 Endocrine Physiology - 22 The pancreas produces two hormones: insulin and glucagon Insulin (lowers blood glucose) Glucagon (raises blood glucose) Insulin is produced as a prohormone (proinsulin) and then converted to insulin enzymatically (C-peptide) Proinsulin Insulin 66 - Behavioral Neuroendocrinology Endocrine Physiology - 23 Sympathetic Parasympathetic The pancreas is innervated by both sympathetic and parasympathetic nerves. Parasympathetic nerves stimulate insulin secretion. Sympathetic nerves inhibit insulin secretion. (+) (-) Pancreas Type 1 diabetes is caused by the inability to produce insulin – an autoimmune disease. Type 2 diabetes is caused by target tissue insensitivity to insulin. Behavioral Neuroendocrinology - 67 Endocrine Physiology - 24 (s) The adrenal glands sit right on top of the kidneys kidney (renal) bladder 68 - Behavioral Neuroendocrinology Endocrine Physiology - 25 The adrenals are actually two glands in one: the cortex and the medulla (cortex = bark) (medulla = marrow) Adrenal cortex: Controlled by the pituitary gland (ACTH) Secretes steroid hormones (relatives of sex hormones) cortisol (glucocorticoid) Adrenal medulla: Controlled by direct sympathetic innervation Secretes epinephrine (adrenalin, a catecholamine) epinephrine (catecholamine) Behavioral Neuroendocrinology - 69 Endocrine Physiology - 26 Interrelationships among steroid hormones cholesterol progestins (progesterone) androgens (testosterone) aromatase glucocorticoids (estradiol) (cortisol) estrogens Negative feedback in the HPA axis: Hypothalamus (-) Cortisol or corticosterone (glucocorticoids) Corticotropin-releasing hormone (CRH) (-) Anterior pituitary Adrenocorticotropic hormone (ACTH) Adrenal cortex 70 - Behavioral Neuroendocrinology Endocrine Physiology - 27 Sympathetic Secretion of epinephrine by the adrenal medulla is stimulated by sympathetic nerves. There is no parasympathetic innervation of the adrenal medulla adrenal medulla (inside) The secretory (hormone-producing) cells in the adrenal medulla are just (very) simplified neurons. Behavioral Neuroendocrinology - 71 Endocrine Physiology - 28 The ovaries produce two kinds of steroid hormones: estrogens and progestins (an estrogen) estradiol progesterone (a progestin) 72 - Behavioral Neuroendocrinology Endocrine Physiology - 29 Negative feedback in the HPG ( ) axis; Hypothalamus (-) Estradiol, progesterone Gonadotropin-releasing hormone (GnRH) (-) Anterior pituitary Luteinizing hormone (LH) Ovary The testes produce androgens. testosterone (an androgen) The adrenal medulla is also a source of androgens – in men and women. Behavioral Neuroendocrinology - 73 Endocrine Physiology - 30 Negative feedback in the HPG ( ) axis; Hypothalamus (-) Testosterone, dihydrotestosterone Gonadotropin-releasing hormone (GnRH) (-) Anterior pituitary Luteinizing hormone (LH) Testis Testosterone as a prohormone (converted to active form in target tissue) 5α-reduction (5α-reductase) n t io iza ) at tase om a ar rom testosterone dihydrotestosterone (genitalia, hair) (Propecia, Avodart – 5α-reductase inhibitors) (a aromatic (benzene) ring estradiol (brain) 74 - Behavioral Neuroendocrinology Endocrine Physiology - 31 Hormone secretion from other places – places that don’t fit the traditional notions of glands Adipose (fat) tissue produces the hormone, leptin, which tells the brain about the availability of calories The walls of the stomach produce ghrelin, which is probably a hunger signal. The upper part of the small intestine (duodenum) produces cholecystokinin, which signals the brain to end a meal. The pancreas (ghrelin) (CCK) Behavioral Neuroendocrinology - 75 Endocrine Physiology - 32 How hormone receptors work. There are two types of receptors: Membrane receptors – amines and peptides (on outer surface of cell) Nuclear receptors – steroids and T3 (inside cell) 76 - Behavioral Neuroendocrinology Endocrine Physiology - 33 There are three types of receptors located in membranes: 1. Ion-channel-linked receptors 2. Receptor kinases 3. G protein-linked receptors Ion-channel-linked receptors (e.g., neurotransmitters) Ligand Ions Ligand: the molecule that is bound by the receptor. (can be an agonist or antagonist; natural or synthetic) Behavioral Neuroendocrinology - 77 Endocrine Physiology - 34 Receptor kinases Ligand active catalytic domain (active enzyme) inactive catalytic domain (inactive enzyme) Kinases are enzymes that catalyze the transfer of a highenergy phosphate group from ATP to another molecule. protein (inactive) kinase phosphorylated protein (active) adenosine { ADP ATP 3 phosphates (adenosine triphosphate) (adenosine diphosphate) 78 - Behavioral Neuroendocrinology Endocrine Physiology - 35 G protein-linked receptors (most peptide & protein hormones) ligand G protein activated G protein enzyme (or ion channel) activated enzyme (or ion channel) second messengers (cAMP) 1. Ligand binds to receptor, causing G protein to bind to receptor inside the cell. 2. Activated G protein interacts with another trans-membrane protein. 3. Activated protein alters cell function. G protein-linked receptors: up close and personal NH2 ligand-binding site COOH G protein binding sites Behavioral Neuroendocrinology - 79 Endocrine Physiology - 36 Steroids and thyroid hormones actually pass through the cell membrane and bind to intracellular receptors. Thyroid hormones and steroids are relatively small molecules and have similar mechanisms of action. triiodothyronine (T3) estradiol (an estrogen) 80 - Behavioral Neuroendocrinology Endocrine Physiology - 37 Genetic transcription and translation: DNA transcription messenger RNA translation protein Mechanism of steroid hormone action 1. Steroid passively diffuses across cell membrane. 2. The steroid is bound by receptor in cytoplasm. 3. There is a change in the conformation (shape) of the steroid receptor. 4. The steroid-receptor complex moves into the cell nucleus. 5. The steroid-receptor complex associates with specific DNA sequences and initiates genetic transcription. (The receptor is a transcription factor.) 6. Synthesis of messenger RNA (mRNA) changes. 7. The mRNA moves into the cytoplasm where it is translated into protein. 8. The protein(s) are processed into their final form. Behavioral Neuroendocrinology - 81 Endocrine Physiology - 38 Antiestrogens: Induction of genetic transcription (RNA synthesis) by estrogen receptors requires the assistance of coactivators Antiestrogens like tamoxifen bind to estrogen receptors, enter the cell nucleus, but do not bind to the coactivators. Genetic transcription is not affected. 82 - Behavioral Neuroendocrinology Endocrine Physiology - 39 Behavioral Neuroendocrinology - 83 Endocrine Physiology - 40 84 - Behavioral Neuroendocrinology Sexual Differentiation - 1 Sexual Differentiation Behavioral Neuroendocrinology - 85 Sexual Differentiation - 2 86 - Behavioral Neuroendocrinology Sexual Differentiation - 3 Sex determination and differentiation Sexual dimorphisms and how they get there. Behavioral Neuroendocrinology - 87 Sexual Differentiation - 4 Sexual dimorphisms Dimorphism: the existence of two different forms (as of color or size) of a species especially in the same population <sexual dimorphism> - robins Behaviors can be sexually dimorphic, too - aggression Dimorphisms are not necessarily absolute. There is often overlap between males and females What are some sexual dimorphisms? 88 - Behavioral Neuroendocrinology Sexual Differentiation - 5 Just talking about sexual dimorphism can influence (or create?) it. Men versus women: Math performance Math Test Read Text Four different kinds of text: 1. No Gender Difference (ND) Condition: A meta-analysis across multiple countries revealed that males and females performed equally well on math tests. 2. Standard Stereotype Threat (S) Condition: The role of the female body in the arts was discussed with relation to women’s identity. 3. Genetic (G) Condition: Males perform 5 percentile points better on math tests than women because of some genes that are found on the Y chromosome. 4. Experiential (E) Condition: Males perform 5 percentile points better on math tests than women because teachers biased their expectations during early school formative years. Math Test Behavioral Neuroendocrinology - 89 Sexual Differentiation - 6 Dar-Nimrod, I. and Heine, S.J. Exposure to scientific theories affects women’s math performance. Science, 314: 435, 2006. Sex and gender: Are they different? If so, how? 90 - Behavioral Neuroendocrinology Sexual Differentiation - 7 Why have two sexes? Having two sexes really complicates things and takes up a tremendous amount of time and energy. Isn’t parthenogenesis good enough for us? (parthenogenesis: eggs develop without fertilization seen in some types of fish, amphibians, and reptiles) How do we get to be males and females? Behavioral Neuroendocrinology - 91 Sexual Differentiation - 8 Mammalian sex determination XY SRY gene testis determination factor bipotential gonad (medulla) testis XX No SRY gene bipotential gonad (cortex) bipotential gonad (cortex) ovary ovary (knockout) testis determination factor XY-SRY No SRY gene XX+SRY (transgenic) bipotential gonad (medulla) testis Undifferentiated internal plumbing immature gonad note: two sets of pipes, one and one 92 - Behavioral Neuroendocrinology Sexual Differentiation - 9 The fetal testis produces two substances which determine whether the internal plumbing will be male or female (or neither or both). Mullerian inhibitory hormone (a peptide) acts on the female plumbing (Mullerian ducts) and causes it to atrophy. Defeminization Testosterone (a steroid) acts on the male plumbing (Wolffian ducts) and makes them grow. Masculinization Internal plumbing Mullerian inhibitory hormone (inhibits growth of female plumbing) no MIH means female plumbing develops testosterone (stimulates growth of male plumbing) no testosterone means no male plumbing If the testes produce testosterone, but not MIH, the fetus will develop both and internal plumbing If the testes produce MIH, but not testosterone, the fetus will develop neither nor internal plumbing Behavioral Neuroendocrinology - 93 Sexual Differentiation - 10 Differentiation of the external plumbing ovary nada testis testosterone 5α-DHT note: one set of tissues, either or , but not both Summary: differentiation of the and genitalia In the absence of testes, the internal and external genitalia develop in a normal female pattern. Nothing else is necessary. The default is female. In males, Mullerian inhibitory hormone defeminizes the internal genitalia and make the Mullerian ducts regress. Testosterone masculinizes the internal genitalia by stimulating growth of the Wolffian ducts. In males, testosterone (via 5α-DHT) also masculinizes the external genitalia. In the absence of 5α-DHT, the default is female external genitalia. 94 - Behavioral Neuroendocrinology Sexual Differentiation - 11 Hermaphrodite: animal with both male and female reproductive systems. Pseudohermaphrodite: animal with ovaries or testes and ambiguous external genitalia. Jaimie Lee Curtis is not a hermaphrodite!!! The Tfm mutation: A disorder of morphological sexual differentiation – in humans and other species 1. Born anatomically normal girls and raised as females. 2. Normal anatomical changes at puberty but fail to menstruate. 3. Upon seeking medical help, it turns out that they are genetic males (XY) and have testes, not ovaries. 4. Therapeutic treatment is usually to remove the testes and treat with exogenous ovarian hormones. 5. But they are infertile, of course. Behavioral Neuroendocrinology - 95 Sexual Differentiation - 12 The Tfm mutation: So how does this happen? 1. They have one X and one Y chromosome – normal male makeup. 2. The Y chromosome is normal – SRY gene is fine, and they develop normal testes. 3. Their testes produce Mullerian inhibitory hormone and testosterone – Mullerian (female) ducts degenerate. 4. But there is a defective gene on the X chromosome – the gene for the androgen receptor. 5. Their tissues cannot respond to the androgens secreted by their testes – Wolffian ducts degenerate, external genitalia remain female. 6. At puberty their testes produce very high levels of testosterone – no negative feedback. 7. Enough of this testosterone is aromatized (converted to estradiol) so that female secondary sexual characteristics develop normally – except for pubic/axillary hair. Sexual differentiation in birds and reptiles – “unmammals.” 96 - Behavioral Neuroendocrinology Sexual Differentiation - 13 Birds aren’t mammals. estrogens female sexual development ovary WZ tissues testis ZZ tissues male sexual development In the absence of estrogens, the default is to become a male. Sex determination by incubation temperature in reptiles. High temp = High temp = lizards & alligators many turtles snapping turtles & crocodiles Behavioral Neuroendocrinology - 97 Sexual Differentiation - 14 Sex hormones can override the effects of incubation temperature on sexual differentiation. ‘ ‘ ’ temperature + testosterone = ’ temperature + estradiol = Thus, temperature controls sexual differentiation by determining whether the gonads produce testosterone or estradiol. Temperature may direct sexual differentiation in reptiles by controlling the activity of the enzymes catalyzing the synthesis of androgens and estrogens. Hypothetical example: testosterone aromatase estradiol testosterone 5α-reductase 5α-DHT Mostly female Mostly male 98 - Behavioral Neuroendocrinology Sexual Differentiation - 15 Sexual differentiation of reproductive behaviors in mammals mounting lordosis There is a sex difference in adult behavioral responsiveness to gonadal hormones* testosterone propionate (TP)* mounting lordosis estradiol+progesterone* estradiol+progesterone* no lordosis no mounting TP* * activating effects of gonadal hormones Behavioral Neuroendocrinology - 99 Sexual Differentiation - 16 Organization and activation of the neural circuits controlling copulatory behavior What accounts for this sex difference in activational effects of sex hormones? Do hormones produced early in life determine adult sensitivity to hormones, as they do in the reproductive organs? That is, do they organize the neural circuitry? To test this hypothesis, Phoenix, et al. (1959) looked at the effects of treating pregnant guinea pigs with testosterone. Effects of prenatal exposure to testosterone on adult sexual behavior in guinea pigs: moms newborn adult E+ P TP lordosis no mounting pregnant + oil vehicle E+ P TP no lordosis mounting E+ P TP no lordosis mounting pregnant + testosterone E+ P TP no lordosis mounting 100 - Behavioral Neuroendocrinology Sexual Differentiation - 17 Prenatal exposure to exogenous testosterone masculinizes and defeminizes the neural circuits controlling copulatory behavior in female guinea pigs. It has no effect on males – Why? Organizing and activating effects of steroids. These actions of testosterone early in development are called organizing effects – they cause permanent changes in the nervous system. In adulthood, ovarian and testicular hormones act on the differentiated brain to have activating effects – these effects are transient and last only as long as the hormone is present. Behavioral Neuroendocrinology - 101 Sexual Differentiation - 18 So what happens to males if you deprive them of androgens early in development? You can’t do this experiment in guinea pigs, because they’re already sexually differentiated at birth. (~60-day gestation period) To study the effects of androgen withdrawal on sexual differentiation, people use rodents whose pups are immature at birth, i.e., mice, rats, and hamsters. (18-21-day gestation period) 102 - Behavioral Neuroendocrinology Sexual Differentiation - 19 Effects of neonatal gonadectomy on adult copulatory behavior in rats (neonate = newborn) neonate adult E+ P TP ovariectomy at birth lordosis no mounting Ovaries are not necessary for normal female development. E+ P TP lordosis no mounting castration at birth In the absence of testes, starting at birth, genetic males are not masculinized or defeminized. Effects of neonatal treatment with testosterone on adult copulatory behavior in rats neonate adult ovariectomy + TP E+ P TP no lordosis mounting Neonatal testosterone treatment masculinizes and defeminizes female rats. E+ P TP no lordosis mounting castration + TP Testosterone treatment prevents the effects of neonatal castration. Behavioral Neuroendocrinology - 103 Sexual Differentiation - 20 Summary: organizing actions of androgens on copulatory behaviors 1. The default condition is female. In the absence of gonadal secretions, the mammalian brain develops in a normal female pattern. 2. The presence of testosterone during proscribed periods of development masculinizes and defeminizes the brain, regardless of genetic sex. 3. Treatment with testosterone has no effect on sexual differentiation when given outside this ‘critical period.’ 4. Testosterone exposure during early development permanently changes (organizes) the neural circuits controlling sex behaviors. 5. Treatment with ovarian or testicular hormones in adulthood has only transient activational effects on behaviors. One or two neural circuits for rodent copulatory behaviors? Q: Are the neural circuits controlling copulatory behavior like the internal reproductive structures with two precursors, one for male copulatory behavior and one for female copulatory behavior? Or are they like the external genitalia with only one precursor, making male and female copulatory behaviors mutually exclusive? A: In a number of situations you can see masculinization without defeminization. Thus, male and female copulatory behavior are controlled by different circuits which can function independently. Examples include normal hamsters and rats exposed to moderate levels of testosterone postnatally. 104 - Behavioral Neuroendocrinology Sexual Differentiation - 21 The aromatization hypothesis Many of the organizational effects of testosterone on the nervous system are actually mediated by aromatized metabolites (i.e., estrogens). testes Testosterone in bloodstream T ar om cell nucleus Evidence for the aromatization hypothesis: Treatment with estradiol mimics many of the organizing effects of testosterone. ata se E2 E2+ER cytoplasm Treatment with antiestrogens can block some of the organizing effects of testosterone. Treatment with 5α-dihydrotestosterone does not masculinize or defeminize the nervous system, because 5α-DHT cannot be aromatized. If it is aromatized metabolites of androgens that are really masculinizing and defeminizing developing rat brains, how come rat fetuses aren’t masculinized and defeminized by their mothers’ estrogens? After all, estrogens readily cross the placenta from mom to fetus. Behavioral Neuroendocrinology - 105 Sexual Differentiation - 22 It’s α-feto-protein (αFP) to the rescue! Perinatal rodents have high levels of α-feto-protein in circulation. This protein sequesters (ties up) circulating estrogens, but it does not bind testosterone. fetal testes testosterone T ar om cell nucleus ata se mom’s ovaries E2 E2+ER estradiol αFP αFP+E2 (inactivated) The process of sexual differentiation can be modified by the environment – even in rodents. 106 - Behavioral Neuroendocrinology Sexual Differentiation - 23 Uterine position effect: a female fetus lying between two of her brothers can be slightly masculinized/defeminized compared with a sister who is in between two females ovary 0M female 2M female vagina The mother rat’s licking of her male pups’ anogenital region is necessary for normal male development. Behavioral Neuroendocrinology - 107 Sexual Differentiation - 24 Sex differences in the brain and behaviors. Q: Are there sex differences in brain anatomy that you can actually see? A: That depends on what you mean by ‘see.’ Examples in mammals: • Dendritic shaft versus spine synapses. • Sexually dimorphic nucleus of the preoptic area (SDN). • Spinal nucleus of the bulbocavernosus muscle (SNB). preoptic area 108 - Behavioral Neuroendocrinology Sexual Differentiation - 25 The sexually dimorphic nucleus of the preoptic area (SDN) SDN-POA SDN-POA SDN-POA + TP Q: So what does this thing do? A: Damned if I know. Spinal nucleus of the bulbocavernosus muscle (SNB) Contraction of the BC and LA causes erections in rats. In humans, the BC aids in the expulsion of urine ( and constriction of the vaginal opening ( ). ) Colon Behavioral Neuroendocrinology - 109 Sexual Differentiation - 26 Development of the SNB depends on the presence of androgens during early development. Both males and females start out with plenty of SNB neurons. Androgens prevent programmed cell death and ‘rescue’ the SNB neurons in males (or in females treated with exogenous hormone). Development of the SNB system in Tfm males is completely feminine Number of SNB Motoneurons 200 160 120 80 40 0 (Tfm = no androgen receptor) Wild Type Male Tfm Male After Breedlove + Arnold (1981) 110 - Behavioral Neuroendocrinology Sexual Differentiation - 27 Could the failure of females to show male-like behaviors simply be due to the fact that they lack male external genitalia? “You can’t be a carpenter without the tools.” -- Frank Beach Sex differences in noncopulatory behaviors “So! Planning on roaming the neighborhood with some of your buddies today?” Behavioral Neuroendocrinology - 111 Sexual Differentiation - 28 Urination postures in dogs : leg lift : squat Early (prenatal) androgen exposure increases leg-lifting in adult Neonatal does not prevent leg lifting in adults – differentiation is prenatal Organization without activation – happens without any adult hormone treatments Rough-and-tumble play in juvenile Rhesus monkeys > > +T > (prenatal) Does not depend on masculinization of the genitalia = Organization without activation 112 - Behavioral Neuroendocrinology Sexual Differentiation - 29 Sexual differentiation of human behaviors Humans are rotten experimental animals, so we have to rely on so-called ‘experiments of nature’ to gain information about the role of hormones in sexual differentiation. One example is seen in the condition called compensatory adrenal hypertrophy (CAH). Normal adrenocortical function cholesterol progestins (progesterone) androgens (testosterone) glucocorticoids (cortisol) Behavioral Neuroendocrinology - 113 Sexual Differentiation - 30 Compensatory adrenal hypertrophy (CAH) cholesterol progestins (progesterone) androgens (testosterone) In CAH the adrenal cortex is unable to convert progestins to glucocorticoids, with the result that substantial amounts of androgens are secreted instead. No negative feedback in CAH Normal: Hypothalamus CAH: Hypothalamus (-) CRH (-) CRH cortisol (-) Anterior pituitary cortisol (-) Anterior pituitary ACTH ACTH Adrenal cortex Adrenal cortex androgens androgens 114 - Behavioral Neuroendocrinology Sexual Differentiation - 31 CAH girls: 1. More rough-and-tumble play 2. Thought of as ‘tomboys’ 3. Prefer male playmates 4. Prefer ‘male typical’ toys 5. Reduced interest in infant care 6. Reduced interest in appearance, makeup, jewelry 7. Less fantasizing about marriage and motherhood 8. Slightly higher incidence of lesbianism Interpretive problems? CAH boys? They’re perfectly normal. Why?? Behavioral Neuroendocrinology - 115 Sexual Differentiation - 32 Guevedoces of the Dominican Republic 1. Congenital deficit in type two 5α-reductase 2. Born with female external genitalia 3. Reared as girls 4. Testes, not ovaries, become active at puberty What happens? What does it mean? Interpretive problems? ‘Guevedoces’ literally means penis at 12 years. More info at: http://www.usrf.org/news/010308-guevedoces.html Read “John and Joan” in the “For discussion” folder on SPARK for next time. 116 - Behavioral Neuroendocrinology Sexual Differentiation - 33 The famous case of Joan/John. One of two twin boys had his penis irreparably mutilated in a botched circumcision. He was taken to Johns Hopkins where they advised the parents to rear him as a girl. His penis and testes were removed surgically, and reconstructive surgeons created female external genitalia. From seventeen months of age he was reared as a girl, and his twin brother was reared as a boy. What happened? What does it mean? Sexual orientation and sexual preference. Vera looked around the room. Not another chicken anywhere. And then it struck her – this was a hay bar. Behavioral Neuroendocrinology - 117 Sexual Differentiation - 34 Biological differences between heterosexuals and homosexuals? Why does it matter? 1. Differences in hormone levels? Nope 2. Inherited differences? 3. Neuroanatomical differences? Nope Yes, but . . . Interstitial nuclei of the anterior hypothalamus (INAH) > heterosexual (straight) >> (gay) gay Interpretive problems? Cause or effect? What’s it do? 118 - Behavioral Neuroendocrinology Sexual Differentiation - 35 Sex differences in finger length and sexual orientation. There is a sex difference in the relative lengths of the index (2D) and ring (4D) fingers in people. The 2D:4D ratio is greater in women than in men – the two fingers are more nearly equal in women than in men. This sex difference is thought to be determined by testosterone exposure in utero. More testosterone = smaller 2D:4D ratio. 4D 2D The 2D:4D ratio is greater in women than in men, and the difference is more striking in the right hand. Behavioral Neuroendocrinology - 119 Sexual Differentiation - 36 Lesbians have lower 2D:4D ratios than heterosexual women. But there is no difference between homosexual and heterosexual men. NS * What does this mean???? If 2D:4D ratio reflects the degree of prenatal androgen exposure – and it almost certainly does – then lesbians may have been exposed to higher levels of testosterone prior to birth than heterosexual women were. 120 - Behavioral Neuroendocrinology Sexual Differentiation - 37 What causes heterosexuality? What causes homosexuality? The bottom line is that we don’t know what shapes the direction of sexual attraction in either sex. One can make a case for either socialization or biological factors as playing a role. But there aren’t clear, unambiguous data in either direction. When listening to arguments over nature versus nurture on this issue, keep your bullshit detectors carefully tuned. Behavioral Neuroendocrinology - 121 Sexual Differentiation - 38 122 - Behavioral Neuroendocrinology Male Copulatory Behavior - 1 Male Copulatory Behavior Behavioral Neuroendocrinology - 123 Male Copulatory Behavior - 2 124 - Behavioral Neuroendocrinology Male Copulatory Behavior - 3 Neuroendocrinology of male copulatory behavior. Rusty makes his move What do males and females want from each other, i.e., what are their reproductive strategies? Is the ideal sex partner the ideal mate? (Only an issue for species which actually have ‘mates.’) Behavioral Neuroendocrinology - 125 Male Copulatory Behavior - 4 History of the evidence that something from the testes stimulates male sex drive. 1. Changes in testes and behavior at puberty (antiquity). 2. Seasonal changes in testis size and libido (antiquity). 3. Effects of castration on libido (antiquity). 4. Testis transplants (A. A. Berthold ~1849) 5. Treatment with testicular extracts (C. Brown-Sequard ~1889). 6. Treatment with purified steroids (C. Stone et al. 1930’s). The ‘hydraulic theory’ of male sexual motivation, ~1880. 1. Between copulatory episodes, fluid (semen) builds up in the seminal vesicles. 2. This information is conveyed to the brain via sensory nerves innervating the seminal vesicles. 3. This pressure can be very uncomfortable, and if unrelieved, can be excruciating and permanently damage the male reproductive system. Well, this is just a bunch of bullshit! It’s just a line guys use to convince women to have sex with them. Don’t fall for it. 126 - Behavioral Neuroendocrinology Male Copulatory Behavior - 5 Motivated behaviors (e.g., eating, drinking, copulation, parental behaviors) are typically divided into two phases, appetitive and consummatory. Food intake: Appetitive phase: finding food. Consummatory phase: eating it. Mating behavior: Appetitive phase: finding and courting a mate. Consummatory phase: copulation. Male rat mating behavior has three components. Mounting: male approaches female from behind and places his forepaws on her hindquarters Intromission: male mounts female, inserts penis in vagina, thrusts rapidly, dismounts quickly Ejaculation: male intromits, holds intromission for several seconds, distinctive motor patterns Behavioral Neuroendocrinology - 127 Male Copulatory Behavior - 6 Sequence of rat copulatory behavior (male’s version) 1. Male and female ‘investigate’ each other. 2. Male mounts female and attempts to intromit. a. If successful, male dismounts quickly (< 2 seconds) and then pauses ~10-30 seconds. b. If unsuccessful, male keeps trying without waiting. 3. After 5-15 intromissions male ejaculates. 4. Male ‘plays dead’ and ‘sings’ ultrasonic song for 5-15 minutes. 5. They start all over again until male has ejaculated 510 times. 6. Male loses his interest in sex for a day or two . . . . . . . . . . unless a better offer comes along. The Coolidge Effect: Frank A. Beach One day President and Mrs. Coolidge were visiting a government farm. Soon after their arrival they were taken off on separate tours. When Mrs. Coolidge passed the chicken pens she paused to ask the man in charge if the rooster copulates more than once each day. “Dozens of times,” was the reply. “Please tell that to the President,” Mrs. Coolidge requested. When the President passed the pens and was told about the rooster, he asked “Same hen every time?” “Oh no, Mr. President, a different one each time.” The President nodded slowly, then said, “Tell that to Mrs. Coolidge.” 128 - Behavioral Neuroendocrinology Male Copulatory Behavior - 7 Rat copulatory behavior. (PG-13) Some examples of male copulatory behavior in species other than rats Dogs: 1. Single intromission, thrust 2. Lock (ouch) – Fig. 5.8 in Nelson 3. Multiple ejaculations Guinea pigs: 1. Single intromission, no thrusting 2. Multiple ejaculations Syrian hamsters: 1. Multiple intromissions, thrust 2. Multiple ejaculations Behavioral Neuroendocrinology - 129 Male Copulatory Behavior - 8 Some examples of male copulatory behavior in species other than rats Rhesus monkeys: 1. Single intromission, thrust 2. Multiple ejaculations Bonobos (pygmy chimps): 1. Single intromission, thrusting 2. Ventral - ventral mating 3. Multiple ejaculations Human beings: 1. Single intromission (?), thrust 2. Multiple positions 3. Single ejaculation! In general, predators are more leisurely when they copulate than are prey. Why? 130 - Behavioral Neuroendocrinology Male Copulatory Behavior - 9 Talk about risky sex! Raccoons 25 feet up in the air. An Introduction to Behavioral Endocrinology Fourth Edition Randy J. Nelson Cover photo by Alison Wade Behavioral Neuroendocrinology - 131 Male Copulatory Behavior - 10 Hormonal bases of male copulatory behavior 1. 2. 3. 4. 5. prepubertally (e.g., at weaning) prepubertally + TP as adult never copulate copulate normally gradually stop copulating continue copulating gradual restoration as adult + TP (immediate) as adult + TP (delayed) The effects of testosterone withdrawal and replacement on male copulatory behavior may take weeks, months or even years to be evident. This depends on: 1. The age and experience of the animal. 2. The species. 3. The individual. 132 - Behavioral Neuroendocrinology Male Copulatory Behavior - 11 Does more testosterone make you more manly (or ratly, or hamsterly, etc.)? Is male sex drive an indicator of circulating testosterone levels in males? What are some ways to test this hypothesis? Is a low male sex drive due to too little testosterone? +TP Grunt & Young, 1952 Fig. 5.22 in Nelson Behavioral Neuroendocrinology - 133 Male Copulatory Behavior - 12 In general: Individual differences in male sex drive are not due to differences in circulating testosterone levels. Providing ‘extra’ testosterone does not enhance libido. Circulating testosterone levels do not differ between animals with high and low sex drives. These individual differences in male sex drive seem to be due to differences in the ‘neural substrates’ that mediate the behavior – whatever that means. Duds and studs do not seem to differ in the quantity and distribution of neural androgen receptors. The bottom line is that we still don’t what causes ‘studliness’ or ‘dudliness.’ Contrary to what you may read in the tabloids or see in informercials, there is no magic treatment that turns duds into studs. Having said that, men suffering from profound hypogoandalism do show reductions in sexual desire, and their libidos can be enhanced with testosterone therapy. 134 - Behavioral Neuroendocrinology Male Copulatory Behavior - 13 In many, but not all, mammals, testosterone must be aromatized in the brain to stimulate male copulatory behavior. 1. Blocking aromatase activity may impair male sexual motivation. 2. Male mice without estrogen receptors (ER knockouts) exhibit impaired sexual motivation. 3. Treating castrated males with 5α-DHT does not stimulate sexual motivation. 4. Normal male copulatory behavior can be restored in castrated males by treating them with a combination of estradiol and DHT. adult testes testosterone T ar om cell nucleus ata se neuron E2 E2+ER Neural control of male copulatory behavior. Behavioral Neuroendocrinology - 135 Male Copulatory Behavior - 14 Several lines of evidence point to an important role for the preoptic area (POA) in the control of male copulatory behavior. POA The POA and male copulatory behavior. 1. Lesion studies. 136 - Behavioral Neuroendocrinology Male Copulatory Behavior - 15 Stereotaxic instrument Behavioral Neuroendocrinology - 137 Male Copulatory Behavior - 16 The POA and male copulatory behavior. 1. Lesion studies: Electrolytic lesions of the POA seriously disrupt male copulatory behavior in a number of species. Yeah, but . . . 2. Electrical stimulation studies: There are some reports that electrical stimulation of the POA elicits male copulatory behavior. Yeah, but . . . The POA and male copulatory behavior. 1. Lesion studies. 2. Electrical stimulation studies. 3. Intracerebral testosterone implants. 138 - Behavioral Neuroendocrinology Male Copulatory Behavior - 17 Testosterone implants directly into the POA stimulate copulatory behavior in castrated male rats. POA The POA and male copulatory behavior. 1. Lesion studies. 2. Electrical stimulation studies. 3. Intracerebral testosterone implants. 4. Localization of neural androgen receptors. Behavioral Neuroendocrinology - 139 Male Copulatory Behavior - 18 So is the POA the ‘male sex center’ of the brain? No way!! The brain doesn’t work that way. 1. Thinking about brain ‘centers’ is naïve and oldfashioned. 2. Neurons in the POA have multiple functions in addition to affecting male copulatory behavior, i.e., GnRH secretion, thermoregulation, parental behaviors, thirst, voluntary exercise . . . 3. The POA is just one part of an extensive circuit that controls male copulatory behavior. Ways to measure neural hormone receptors (or any other protein). Immunocytochemistry (ICC): Uses antibodies to recognize, label, and quantify specific protein molecules. Used to detect and quantify any protein in tissue, not just hormone receptors. Antibodies are made when your immune system detects something “foreign” – an antigen. They are highly specific and recognize only the antigen which triggered their production (mostly). 140 - Behavioral Neuroendocrinology Male Copulatory Behavior - 19 How immunocytochemistry (ICC) works 1. Antigen (protein) found in tissue 2. Primary antibody recognizes and binds only with this specific antigen 3. Labeled secondary antibody recognizes and binds with primary antibody 4. Detect label on secondary antibody tissue primary antibody antigen (what you want to detect) secondary antibody label Notes: 1. Antigen, primary antibody, and secondary antibody must be from different species. 2. Lots of ways to label secondary antibody – color reaction, radioactivity, etc. Fos immunostaining in rat amygdala. Behavioral Neuroendocrinology - 141 Male Copulatory Behavior - 20 Immunocytochemistry (ICC): Excellent anatomical resolution – even down to the electron microscopic level Highly antigen-specific if you have the right antibody Can be used for double or triple labeling Only ‘semiquantitative’ It’s easy to lie mislead with ICC How to lie mislead with ICC by tuning the sensitivity of the technique Making the technique less sensitive can increase the apparent magnitude of the treatment effect. Actual amount of receptor threshold 2 (insensitive) threshold 1 (sensitive) *best we can do* no treatment treatment 142 - Behavioral Neuroendocrinology Male Copulatory Behavior - 21 Treatment of ovariectomized rats with estradiol increases progestin receptors in the ventromedial hypothalamus of ovariectomized mice. A: oil vehicle B: EB + estradiol + oil placebo But not this much. J Neurosci. 18: 9556-63, 1998 Ways to measure neural hormone receptors (or any other mRNA). In situ hybridization histochemistry (ISHH) Uses mRNA or DNA probes to detect and measure a specific messenger RNA. Behavioral Neuroendocrinology - 143 Male Copulatory Behavior - 22 How in situ hybridization works: labeled ‘probe’ : RNA complementary to mRNA of interest made by experimenter + * * * = label: e.g., florescent marker or radioactive molecule message (mRNA) you want to measure How in situ hybridization works: message-probe hybrid * * probe * * probe is labeled chemically or radioactively so that it can be detected microscopically message 144 - Behavioral Neuroendocrinology Male Copulatory Behavior - 23 In situ hybridization of vasopressin mRNA in rat brain supraoptic nucleus In situ hybridization histochemistry (ISHH) Excellent anatomical resolution. Highly specific if you have the right probe. Can be used for double or triple labeling Can be combined with immunocytochemistry. Only tells you about presence of particular mRNA; does not tell you if it is translated into protein or if the protein is ‘finished’ Behavioral Neuroendocrinology - 145 Male Copulatory Behavior - 24 All methods of detecting and quantifying receptors find androgen (and estrogen) receptors in the POA – and in other parts of the brain. But is the presence of androgen receptors a requirement for concluding that a particular part of the brain participates in the control of male copulatory behavior? No – not at all. For example: AR AR So how do you assess circuits involved in a particular behavior – not just the parts containing the relevant marker? 1. Immediate-early genes. 2. Tract-tracing 146 - Behavioral Neuroendocrinology Male Copulatory Behavior - 25 Expression of immediate-early genes (IEGs), measured with immunocytochemistry (protein) or in situ hybridization (mRNA), is a very useful way to take a ‘snapshot’ of neural activation throughout the brain at any single point in time. Fos IEG expression is one of the earliest molecular events when neurons are activated. You can use this technique to look at entire circuits at the same time. Fos immunostaining in rat amygdala. You can combine IEG ICC with ICC for other cellular proteins and measure the activation of specific types of neurons. Colocalization of ERα and Fos in the ventromedial hypothalamus of mated and unmated female rats. ERα = estrogen receptor alpha, the main form From Flanagan-Cato, 1999 Behavioral Neuroendocrinology - 147 Male Copulatory Behavior - 26 Tracers can be used to plot the connections of a particular area of the brain. AAA BBB VMH retrograde tracer CCC DDD anterograde tracer Triple labeling of estrogen receptor (ERα), fos, and fluorogold (a retrograde tract-tracer, Pjn) in ventromedial hypothalamus in female rat. 148 - Behavioral Neuroendocrinology Male Copulatory Behavior - 27 Behavioral Neuroendocrinology - 149 Male Copulatory Behavior - 28 150 - Behavioral Neuroendocrinology Female Copulatory Behavior - 1 Female Copulatory Behavior Behavioral Neuroendocrinology - 151 Female Copulatory Behavior - 2 152 - Behavioral Neuroendocrinology Female Copulatory Behavior - 3 Neuroendocrinology of female copulatory behavior Very little research prior to ~1900 compared with work on male copulatory behavior. Why? 1. No economic incentive to remove ovaries 2. Ovariectomy (spaying) more difficult than castrating males 3. Sexism a. Scientists were overwhelmingly male and more interested in what made themselves tick b. Female copulatory behavior thought to be less complex than male behavior c. Females regarded as passive with males doing the interesting parts Behavioral Neuroendocrinology - 153 Female Copulatory Behavior - 4 Unlike males, sub-primate female mammals are willing to mate only at selected times. In sub-primate mammals, the occurrence of female sexual behavior is tightly controlled by the fluctuations in ovarian hormone levels and is limited to the time around ovulation. In many primates, sexual activity is at least partially ‘liberated’ from levels of ovarian hormones. Humans and most other primates have menstrual cycles. ‘Lower’ mammalian species have estrous cycles. Etymology of the word ‘estrous’: New Latin, from Latin oestrus gadfly, frenzy, from Greek oistros Estrus: noun. Estrous: adjective. Oestrus, oestrous, oestrogen, etc: British 154 - Behavioral Neuroendocrinology Female Copulatory Behavior - 5 Behaviorally, the difference between estrous and menstrual cycles is: Animals with estrous cycles only mate for a limited period around the time of ovulation. Animals with menstrual cycles will mate at all stages of the cycle – but there are changes in the female’s interest in mating. The menstrual cycle. Stages of the menstrual cycle: Follicular stage. Periovulatory stage. (peri = around) Luteal stage. Menses. Behavioral Neuroendocrinology - 155 Female Copulatory Behavior - 6 Immediately after menses, pituitary FSH and LH stimulate the growth of one Graafian follicle which secretes increasing amounts of estradiol and keeps FSH and LH levels in check via negative feedback. This is the follicular stage of the cycle. LH/FSH E2/P Ovary menses Eventually the rising levels of estradiol feed back on the hypothalamus and pituitary to evoke a LH surge – positive feedback. The LH surge acts on the mature follicle to cause ovulation – it is called the periovulatory stage (around ovulation). 156 - Behavioral Neuroendocrinology { Uterus LH/FSH E2/P Ovary Uterus Female Copulatory Behavior - 7 The ovulatory LH surge also transforms the follicle into a corpus luteum (yellow body) which produces estradiol and progesterone and supports the maintenance of the uterine lining in preparation for the implantation of a blastocyst. This is the luteal stage of the cycle. LH/FSH E2/P Ovary Uterus If the ovum isn’t fertilized, the corpus luteum eventually degenerates, and estradiol and progesterone levels plummet. Without hormonal support, the uterine lining sloughs off, and menses occurs. LH/FSH E2/P Ovary Uterus Behavioral Neuroendocrinology - 157 Female Copulatory Behavior - 8 The hamster (mouse, rat) estrous cycle. On day one of the hamster estrous cycle, reproductive hormone are quite low, although a fresh set of Graafian follicles starts to develop. ovulation es og te ne ro LH pr estrous behavior day 1 estradiol dark light 158 - Behavioral Neuroendocrinology Female Copulatory Behavior - 9 On day two of the cycle, the Graafian follicles start to secrete increasing amounts of estradiol ovulation es og te ne ro LH pr estrous behavior day 2 estradiol dark light On day three of the cycle, circulating estradiol levels rise sharply ovulation es og te ne ro LH pr estrous behavior day 3 estradiol dark light Behavioral Neuroendocrinology - 159 Female Copulatory Behavior - 10 Days 1-3 of the hamster estrous cycle are akin to the follicular phase of the menstrual cycle. Day 4 of the hamster estrous cycle is akin to the periovulatory phase of the menstrual cycle. On day four of the cycle, the high levels of estradiol induce an LH surge via positive feedback; there is a sharp peak in progesterone secretion; the animal ovulates; and she becomes sexually receptive briefly (~12 hr) ovulation es og te ne ro LH pr estrous behavior day 4 estradiol dark light 160 - Behavioral Neuroendocrinology Female Copulatory Behavior - 11 Beasts with short estrous cycles, such as rats, mice, and hamsters, do not have a spontaneous luteal phase. Unless they mate, the corpora lutea degenerate quickly, and they start a new cycle immediately. Other species, such as guinea pigs, do form functional corpora lutea in the absence of copulatory stimulation. They have a prolonged period of high progesterone secretion, and their estrous cycles are longer than those of rats, mice and hamsters (guinea pig cycle ~16 days). er st e on ovulation LH estrous behavior o pr ge estradiol dark light Three aspects of female mating behaviors (Beach). 1. Attractiveness: self explanatory 2. Proceptivity: things the female does to put her into a male’s proximity and indicate an interest in mating – ranging from ‘flirting’ to active seduction. Appetitive behavior. 3. Receptivity: the female’s willingness to copulate with the male. Consummatory behavior. Behavioral Neuroendocrinology - 161 Female Copulatory Behavior - 12 Attractiveness increases around the time of ovulation and is often enhanced by rising levels of circulating estradiol. Odors: males often find the scent of periovulatory females more attractive than those of ovariectomized females or females at other stages of their ovulatory cycle. Visual stimuli: In many nonhuman primates the genital area may become swollen or take on a brighter color (red). This drives the guys wild. Hormonal induction of female sexual behavior. 162 - Behavioral Neuroendocrinology Female Copulatory Behavior - 13 The most effective hormone regimen for inducing any behavior in female mammals is the one which most closely mimics the pattern of endogenous hormone secretion. e on er st e og pr ovulation LH estrous behavior estradiol Ovariectomized rats, mice, guinea pigs and hamsters: EB P estrous behavior e on er st e og pr ovulation LH estrous behavior estradiol Behavioral Neuroendocrinology - 163 Female Copulatory Behavior - 14 Rats and hamsters will come into heat if you give them only a really large dose of estradiol. But they show no proceptive behaviors. EB estrous behavior pr ne ro te es og ovulation LH estrous behavior estradiol So . . . is progesterone really necessary? 164 - Behavioral Neuroendocrinology Female Copulatory Behavior - 15 Yes. If you spay a female rat (hamster, guinea pig) after the peak in estradiol, but before progesterone rises, she won’t come into heat. e on estrous no estrous behavior LH o pr ge er st estradiol If you spay a female rat (hamster, guinea pig) after the peak in estradiol, but before progesterone rises, she won’t come into heat. But if you inject her with progesterone, too, she will come into heat. LH o pr ge er st e on +P estrous behavior estradiol Behavioral Neuroendocrinology - 165 Female Copulatory Behavior - 16 What happens if you give only progesterone? Nothing. Why? Because estrogen priming is necessary to increase neural concentrations of progestin receptors (among other things). Effect of estradiol treatment on progestin receptor immunoreactivity (PRIR) in the ventromedial hypothalamus of ovariectomized mice (exaggerated). A: oil vehicle B: EB 166 - Behavioral Neuroendocrinology Female Copulatory Behavior - 17 Proceptive behaviors, too, are increased by the hormonal changes that lead up to ovulation. The exact nature of those hormonal changes depends on the species. a. In some species (e.g., rhesus monkeys) rising estradiol levels alone are sufficient to stimulate proceptive behaviors, including approaching the male and ‘presenting.’ b. In other species (e.g., rats) increases in both estradiol and progesterone are required to stimulate proceptive behaviors, ear-wiggling, darting and hopping. Why this difference?? e on er st e og pr ovulation LH estrus estradiol LH/FSH E2/P Behavioral Neuroendocrinology - 167 Female Copulatory Behavior - 18 The pacing of rat mating behavior: Who’s in charge here? That depends on the physical environment. In a small, contained environment, the male exerts a great deal of control. But in a larger more naturalistic environment, the female does the pacing. In general the female will choose a slower pace than the male. Interval after sexual contact: mount < intromission < ejaculation So why put the female in charge? 168 - Behavioral Neuroendocrinology Female Copulatory Behavior - 19 Rats (mice, hamsters) do not form functional corpora lutea unless they receive vaginal-cervical stimulation (VCS) during mating. Without VCS, functional corpora lutea and, thus, pregnancy cannot occur. It is essential that the VCS occur in a particular temporal pattern. The female chooses a ‘more-optimum’ pattern of sexual contacts than does the male. The appropriate pattern of VCS without seminal emission causes the formation of functional corpora lutea and pseudopregnancy – the hormonal equivalent of early pregnancy. e on er st e og pr ovulation LH estrous behavior estradiol Vaginal-cervical stimulation elicits twice daily surges of prolactin, which prevent the corpora lutea from atrophying mating The female picks the optimal pacing necessary to elicit these prolactin surges and functional corpora lutea. Behavioral Neuroendocrinology - 169 Female Copulatory Behavior - 20 In species without a spontaneous luteal phase, the male provides two things needed for successful pregnancy: 1. He deposits spermatozooa 2. He provides the vaginal-cervical stimulation necessary for the formation and maintenance of corpora lutea Hormones and sexual behavior in female primates 170 - Behavioral Neuroendocrinology Female Copulatory Behavior - 21 In rhesus monkeys the distribution of male sexual behavior across the menstrual cycle depends on the social situation, including the number of males and females present, enclosure size, and social rank. !! many +many ovulation 1 +1 1 +many The likelihood that a female rhesus monkey will mate with a male depends on a number of factors: 1. The size of the enclosure: Housing one male and one female in a small enclosure ensures that they will mate often and usually without regard to the female’s menstrual cycle. 2. The stage of her menstrual cycle: In larger enclosures, females mate more often around mid-cycle – the periovulatory period. This is particularly true if you focus on sexual contacts initiated by the female. 3. Her place in the pecking order: Dominant females mate throughout their cycles, although they initiate more contacts mid-cycle. Subordinate females mate only around the time of ovulation – there is a cost to mating at other times. Behavioral Neuroendocrinology - 171 Female Copulatory Behavior - 22 Treatment with estradiol alone increases sexual interest in ovariectomized rhesus monkeys OVEX+E2 OVEX no E2 Nonpregnant (not in season) Why isn’t progesterone required? In female primates there is no sharp rise in circulating progesterone levels during the periovulatory period. LH/FSH E2/P Ovary Uterus 172 - Behavioral Neuroendocrinology Female Copulatory Behavior - 23 The most effective hormone treatments for inducing copulatory behaviors in ovariectomized animals are the ones which most closely match those of endogenous secretion leading up to ovulation in that species: Monkeys, shrews*, cats*, et al. estradiol alone. *Critters like shrews and cats are reflex ovulators, so they don’t produce significant amounts of progesterone before copulation. Rats, mice, guinea pigs, et al. estradiol + progesterone. What about androgens and female sexual behavior in primates? Some early work suggested that it might be (adrenal) androgens, rather than ovarian hormones, that stimulate sexual desire in female primates. But recent work clearly shows that concurrent treatment with an antiestrogen prevents the increase in sexual initiation induced by testosterone treatment in ovariectomized rhesus monkeys. Thus, testosterone has to be aromatized to estradiol in order to stimulate proceptivity and receptivity in monkeys. Behavioral Neuroendocrinology - 173 Female Copulatory Behavior - 24 What about women? What determines the likelihood that a woman will engage in sexual relations? Day of the week – Saturday night/Sunday morning. 174 - Behavioral Neuroendocrinology Female Copulatory Behavior - 25 There are lots of published reports that the likelihood of sexual activity varies across women’s menstrual cycles. Yeah, but . . . The field was riddled with contradictions. Confounding factors include things like risk of pregnancy and the partner’s wishes. Investigators usually just asked women to keep track of when they had sex and when they had their periods. So . . . do hormones matter for women? Behavioral Neuroendocrinology - 175 Female Copulatory Behavior - 26 Yes. Female initiated Partner initiated Women are more likely to initiate sexual contacts around the time of ovulation. total sexual outlet %h ete ros exu al 176 - Behavioral Neuroendocrinology % Women are more likely to masturbate during the periovulatory period. au tos e xu a l Female Copulatory Behavior - 27 Women report an increase in sexual desire around the time of ovulation. women (increased desire) monkeys (approach male) If women have sexual contacts with men other than their primary partners, they are more likely to do so during the periovulatory period than at other times in their cycles. Increased risk-taking? Distribution of ‘double matings’ in which women had intercourse with their primary partners and an extrapair partner within 5 days. Behavioral Neuroendocrinology - 177 Female Copulatory Behavior - 28 Women report increased sexual interest in nonprimary partners around the time of ovulation. Women ‘try to look more attractive’ around the time of ovulation. 178 - Behavioral Neuroendocrinology Female Copulatory Behavior - 29 Changes in human attractiveness across the menstrual cycle? Lots of unresolved questions: What about menopause? Age? Discomfort? Estrogens? What about hormone replacement therapy? What are the effects of antiestrogens and aromatase inhibitors (in chemotherapy)? Behavioral Neuroendocrinology - 179 Female Copulatory Behavior - 30 Neural control of female copulatory behavior. A great deal is known about the neurobiology of lordosis, but comparatively little is known about the neural control of proceptive behaviors. Measuring a simple reflex (lordosis) is simpler than measuring a complicated group of behaviors that encompass proceptivity 180 - Behavioral Neuroendocrinology Female Copulatory Behavior - 31 A simplified wiring diagram of the neural circuitry mediating lordosis - à la Pfaff (Fig. 6.21 in Nelson). Estradiol and progesterone act at several sites throughout the forebrain to induce estrous behavior, but the ventromedial hypothalamus (VMH) plays a pivotal role in this process. Behavioral Neuroendocrinology - 181 Female Copulatory Behavior - 32 Evidence that the ventromedial hypothalamus plays a pivotal role in this process: 1. Lesions of the VMH abolish estrous behavior permanently. 2. The VMH contains both estrogen and progestin receptors. 3. Treatment of ovariectomized rats with estradiol increases the concentration of progestin receptors in the VMH. Effect of estradiol treatment on progestin receptor immunoreactivity (PRIR) in the ventromedial hypothalamus of ovariectomized mice A: oil vehicle B: EB 182 - Behavioral Neuroendocrinology Female Copulatory Behavior - 33 Evidence that the ventromedial hypothalamus plays a pivotal role in this process: 1. Lesions of the VMH abolish estrous behavior permanently. 2. The VMH contains both estrogen and progestin receptors. 3. Treatment of ovariectomized rats with estradiol increases the concentration of progestin receptors in the VMH. 4. Mating stimulation induces Fos-IR in ER-IR-containing neurons in the VMH. Colocalization of ERIR and Fos-IR in the ventromedial hypothalamus of mated and unmated female rats. Behavioral Neuroendocrinology - 183 Female Copulatory Behavior - 34 Evidence that the ventromedial hypothalamus plays a pivotal role in this process: 1. Lesions of the VMH abolish estrous behavior permanently. 2. The VMH contains both estrogen and progestin receptors. 3. Treatment of ovariectomized rats with estradiol increases the concentration of progestin receptors in the VMH. 4. Mating stimulation induces Fos-IR in ER-IR-containing neurons in the VMH. 5. Implants of estradiol limited to the VMH facilitate estrous behavior in ovariectomized rats, and antiestrogens in VMH block the effects of systemic estradiol. Implants of estradiol in the VMH facilitate estrous behavior in spayed rats. E2 in VMH P test 184 - Behavioral Neuroendocrinology Female Copulatory Behavior - 35 Implanting an antiestrogen into the VMH just before giving a systemic injection of EB prevents the induction of estrous behavior. Antiestrogen in VMH P test Systemic EB injection Evidence that the ventromedial hypothalamus plays a pivotal role in this process: 1. Lesions of the VMH abolish estrous behavior permanently. 2. The VMH contains both estrogen and progestin receptors. 3. Treatment of ovariectomized rats with estradiol increases the concentration of progestin receptors in the VMH. 4. Mating stimulation induces Fos-IR in ER-IR-containing neurons in the VMH. 5. Implants of estradiol limited to the VMH facilitate estrous behavior in ovariectomized rats, and antiestrogens in VMH block the effects of systemic estradiol. 6. Implants of progesterone limited to the VMH facilitate estrous behavior in ovariectomized, estrogen-primed rats. Behavioral Neuroendocrinology - 185 Female Copulatory Behavior - 36 EB P in VMH test Implants of progesterone in the VMH facilitate estrous behavior in spayed, EB-primed rats. What happens if you stimulate the VMH electrically? The animals freak out. It is very aversive. 186 - Behavioral Neuroendocrinology Female Copulatory Behavior - 37 Although the VMH plays a pivotal role in the control of estrous behavior, it is not the ‘female sex center.’ 1. There is no such thing as centers. 2. Lesions in other parts of the brain affect estrous behavior, although not as dramatically as the VMH. Lesions of the preoptic area or lateral septum actually facilitate estrous behavior. 3. Hormone implants in other parts of the brain can facilitate estrous behavior. Estradiol in the preoptic area facilitates estrous behavior. Progesterone in the midbrain central gray facilitates estrous behavior. Behavioral Neuroendocrinology - 187 Female Copulatory Behavior - 38 Pheromones and reproduction in female mammals Pheromone: a chemical substance that is produced by an animal and serves as a signal to other individuals of the same species for one or more behavioral responses 1. Pregnancy block in female mice. Exposure of a newly pregnant (≤ 5 days) female mouse to a strange male can cause pregnancy termination, ovulation, and estrous behavior (Bruce effect). The male’s odor alone is sufficient to block pregnancy. Androgen dependent. The stranger, the better. Why? 188 - Behavioral Neuroendocrinology Female Copulatory Behavior - 39 2. Delaying puberty. Mice housed in large groups of females, particularly with their moms and sisters may exhibit delayed puberty. Estrogen dependent. 3. Advancing puberty. Exposure to the odor of an adult male can advance puberty. Androgen dependent. Why? 4. Estrus synchrony. Females rodents housed in close proximity often synchronize their estrous cycles. Olfactory cues are sufficient. Why? Behavioral Neuroendocrinology - 189 Female Copulatory Behavior - 40 Menstrual synchrony. Women living in close proximity often synchronize their menstrual cycles (McClintock, 1971). Axillary (underarm) secretions from women can shorten or lengthen cycles of other women, although they appear to be odorless. ‘Ovulatory pheromone’ lengthens cycle, and ‘follicular pheromone’ shortens cycle (Stern & McClintock, 1998). (0-2 days after LH surge) (70% isopropyl alcohol) (2-4 days before LH surge) 190 - Behavioral Neuroendocrinology Female Copulatory Behavior - 41 Boar taint. A female pig sprayed with boar taint, the musky smell of male pig which also contains androstenol, will become sexually receptive, and assume a mating posture. For this reason, androstenol sprays are manufactured on a large scale for use by pig breeders. Androstenol is also widely used in perfumes and soaps. The makers would have us believe that it has the same effect on women that it has on sows. Sprays containing androstenol can be bought in sex shops with the promise that it will attract women. Yeah, right. Should you invest your money in perfumes or colognes that will make you irresistible to the opposite sex? Yeah, right. And pigs can fly. Behavioral Neuroendocrinology - 191 Female Copulatory Behavior - 42 192 - Behavioral Neuroendocrinology Parental Behavior - 1 Parental Behavior Behavioral Neuroendocrinology - 193 Parental Behavior - 2 194 - Behavioral Neuroendocrinology Parental Behavior - 3 Parental behavior. Although courtship and mating are the fun part, reproduction certainly doesn’t end there. In fact, it’s just starting. If you want to pass your genes on to the next generation, you have to care for and nurture your offspring until they can go off and fend for themselves. Depending on your species, this can take a very long time. (But it’s all worth it.) Behavioral Neuroendocrinology - 195 Parental Behavior - 4 Because of the tremendous energetic and temporal demands of parental care, environmental conditions have to be just right for reproduction to occur. Many species use food availability or day length to determine the time of mating – so that the young will be born at a time when food is available. There is no common theme to parental behavior among vertebrates - or even mammals, for that matter. 1. Biparental versus uniparental: >90% of birds are biparental >90% of mammals are uniparental In 100% of mammals, the female is involved in parental care Why? But male mammary glands can be induced to grow and produce milk – witches’ milk 196 - Behavioral Neuroendocrinology Parental Behavior - 5 There is no common theme to parental behavior among vertebrates - or even mammals, for that matter. 2. Precocial versus altricial young: Some species give birth to precocial young: offspring that are capable of moving independently, maintaining their body temperature, hearing, and seeing shortly after birth. Examples: horses, cattle, elephants, guinea pigs But these little critters still need to be nursed, protected and cared for until they can fend for themselves. Behavioral Neuroendocrinology - 197 Parental Behavior - 6 There is no common theme to parental behavior among vertebrates - or even mammals, for that matter. 2. Precocial versus altricial young: Other species give birth to altricial young: offspring that are very immature. They may be blind, deaf, furless, unable to maintain body temperature, unable to urinate and defecate, etc. Examples: rats, mice, hamsters, dogs, cats Rat pups 198 - Behavioral Neuroendocrinology Parental Behavior - 7 Rat pup Dog pups Behavioral Neuroendocrinology - 199 Parental Behavior - 8 There is no common theme to parental behavior among vertebrates - or even mammals, for that matter. 2. Precocial versus altricial young: Most primate species aren’t clearly precocial or altricial. They can hear, see, maintain their body temperatures, urinate and defecate (can they ever!). But they can’t fend for themselves at all. 200 - Behavioral Neuroendocrinology Parental Behavior - 9 There is no common theme to parental behavior among vertebrates - or even mammals, for that matter. 3. Continuous versus intermittent care: Some species spend relatively little time on parental care. Tree shrews deposit their offspring in an arboreal nest (in a tree) and visit once every 48 hr. Rabbits spend about 3 minutes a day with their babies. ‘Inject’ some milk and then split. Other species (e.g., primates) are in nearly continuous contact with their infants. Most of what we know about the neuroendocrinology of rat maternal behavior is due to the research of Jay Rosenblatt and his academic offspring. Behavioral Neuroendocrinology - 201 Parental Behavior - 10 The basics of rat pregnancy and lactation. 1. A rat pregnancy lasts 22 days with the day after mating counting as day 1. 2. The blastocysts implant in the uterine wall around day 5, after which it becomes difficult to interrupt pregnancy. 3. The mom begins to care for her pups as they emerge from the birth canal. 4. Initially, the pups depend on their mom for everything. Newborn ~ 5 days The basics of rat pregnancy and lactation. 1. A rat pregnancy lasts 22-23 days with the day after mating counting as day 1. 2. The blastocysts implant in the uterine wall around day 5, after which it becomes difficult to interrupt pregnancy. 3. The mom begins to care for her pups as they emerge from the birth canal. 4. Initially, the pups depend on their mom for everything. 5. By day 10, they have fur and can see, hear, move around. ~ 10 days 202 - Behavioral Neuroendocrinology Parental Behavior - 11 The basics of rat pregnancy and lactation. 1. A rat pregnancy lasts 22-23 days with the day after mating counting as day 1. 2. The blastocysts implant in the uterine wall around day 5, after which it becomes difficult to interrupt pregnancy. 3. The mom begins to care for her pups as they emerge from the birth canal. 4. Initially, the pups depend on their mom for everything. 5. By day 10, they have fur and can see, hear, move around. 6. They begin to eat ‘grown-up food’ around day 12, and are weaned completely by day 25. Rats and a number of other species exhibit a postpartum estrus. 1. The hormonal changes just before parturition are similar enough to those just prior to ovulation in cycling animals. 2. Female rats mate and ovulate within 48 hours of parturition. If they copulate with a male, they become pregnant (again). 3. Implantation of the blastocysts may be delayed 5-6 days, but the females are pregnant and lactating at the same time. 4. This puts enormous energy demands on the mom. Lactation alone typically triples food intake. No postpartum estrus in hamsters. Behavioral Neuroendocrinology - 203 Parental Behavior - 12 Commonly measured components of rat maternal behavior Nestbuilding: begins before parturition; helps keep pups warm and provides a ‘central meeting place’ for other transactions Licking & grooming: begins as pups emerge from birth canal; necessary for urination and defecation; affects adult copulatory and maternal behaviors; also recycles water Pup retrieval: begins as pups are born; keeps pups together in nest; gradually declines as pups become mobile Crouching and nursing: begins as pups are born; keeps little poikilotherms warm, transfers food and drink Commonly measured components of rat maternal behavior Nestbuilding: begins before parturition; helps keep pups warm and provides a ‘central meeting place’ for other transactions Licking & grooming: begins as pups emerge from birth canal; necessary for urination and defecation; affects adult copulatory and maternal behaviors; also recycles water Pup retrieval: begins as pups are born; keeps pups together in nest; gradually declines as pups become mobile Homeotherms: animals that maintain a constant body temperature – birds and mammals Poikilotherms: animals that cannot maintain a constant body temperature – pretty much everyone else Crouching and nursing: begins as pups are born; keeps little poikilotherms warm, transfers food and drink 204 - Behavioral Neuroendocrinology Parental Behavior - 13 Appearance of maternal behaviors. Maternal behavior can be induced in rats in the absence of changes in hormone levels simply by exposing them to pups day-after-day. If you do this for approximately a week, they will begin to exhibit all components of maternal behavior. Behavioral Neuroendocrinology - 205 Parental Behavior - 14 You can induce maternal behavior in rats just by repeatedly exposing them to pups. (nulliparous = never pregnant) Median latency for female rats to respond maternally (days) Each animal tested just once (primiparous = pregnant once) Nulliparous rats are afraid of newborn pups, and that’s a major reason why they’re not immediately maternal. Based on the pups’ smell. Anosmic (can’t smell) nullipara are maternal immediately. Olfactory bulbectomy Zinc sulfate (reversible) Lesioning central projections of the olfactory bulbs (e.g., central amygdala) results in immediate maternal responses. Why should nullipara be afraid of rat pups? 206 - Behavioral Neuroendocrinology Parental Behavior - 15 Hormonal bases of maternal behavior in rats. The changes in hormone levels characteristic of mid-to-late pregnancy greatly hasten the onset of maternal responsiveness. Much of the research in this area was done by Bob Bridges at the Tufts Veterinary School. Bridges did his postdoctoral training with Jay Rosenblatt at Rutgers. Behavioral Neuroendocrinology - 207 Parental Behavior - 16 Hormone changes during pregnancy: 1. The vaginal-cervical stimulation of mating causes twice daily surges of prolactin, which induces and maintains functional corpora lutea. 2. The corpora lutea secrete increasing amounts of progesterone. 3. Around mid-pregnancy, the pituitary stops secreting large amounts of prolactin, and a placental hormone takes over. 4. Progesterone and estradiol levels continue to climb. Prolactin is an anterior pituitary hormone whose secretion is controlled by prolactin-inhibiting factor (PIF). progesterone estradiol Hormone changes during pregnancy: 5. About two days before the onset of parturition progesterone levels drop sharply, but estradiol levels remain high. These changes are necessary for parturition to occur. Progesterone inhibits uterine contractions, and estradiol facilitates them. 6. Just before parturition, prolactin levels shoot up. Prolactin stimulates the mammary glands to produce milk. 7. During parturition, oxytocin levels rise quickly. Oxytocin causes uterine contractions which expel the pups. progesterone estradiol 208 - Behavioral Neuroendocrinology Parental Behavior - 17 The hormone treatments that are most effective at inducing maternal behavior are those which mimic the endogenous changes at parturition. 1. An abrupt drop in progesterone levels. If progesterone levels remain high, labor is inhibited and dams do not show maternal behavior. Used clinically to prevent premature labor. progesterone estradiol The hormone treatments that are most effective at inducing maternal behavior are those which mimic the endogenous changes at parturition. 2. High estradiol levels. A drop in progesterone levels alone does not elicit the appearance of maternal behavior. Estradiol levels must be high, too. progesterone estradiol Behavioral Neuroendocrinology - 209 Parental Behavior - 18 The hormone treatments that are most effective at inducing maternal behavior are those which mimic the endogenous changes at parturition. 3. A sharp rise in prolactin levels. Prevention of the rise in prolactin inhibits maternal behavior. Suckling stimulation from the pups increases prolactin secretion, which in turn, stimulates milk production. 4. A sharp rise in oxytocin levels. Treatment with an oxytocin antagonist delays the onset of maternal behavior. progesterone estradiol Rapid-onset maternal behavior can be induced in ovariectomized rats by: 1. Repeated daily injections of estradiol + progesterone 2. Cessation of progesterone injections with continuing treatment with estradiol 3. Treatment with prolactin 4. Treatment with oxytocin Ovarian steroids and oxytocin are needed for the rapid induction of maternal behavior but not its maintenance. High prolactin levels are needed to maintain maternal behavior during lactation. 210 - Behavioral Neuroendocrinology Parental Behavior - 19 Neural control of maternal behavior in rats Much of the research in this area was done by Michael Numan at Boston College. Numan did his postdoctoral training with Jay Rosenblatt at Rutgers. Several lines of evidence point to an important role for the preoptic area (POA) in the control of maternal behavior. Remember, it’s important in the control of male copulatory behavior, too. Behavioral Neuroendocrinology - 211 Parental Behavior - 20 The preoptic area and maternal behavior 1. Lesions of the POA eliminate nest building and pup retrieval. Crouching in a nursing posture is unaffected. Animals will retrieve non-pup items, e.g., candy. Animals will build nests in the cold. Seen in pup-induced and postpartum animals. 2. POA contains receptors for estradiol, progesterone, prolactin, and oxytocin. Levels of all these receptors rise during the course of pregnancy. 3. Implants of estradiol or prolactin in the POA facilitate maternal behavior. An oxytocin antagonist in the POA decreases maternal behavior in newly parturient rats. 4. Repeated exposure to pups induces Fos expression in the POA of nulliparous animals. Parental behavior in primates is extremely diverse. 1. The incidence of placentophagia varies with species and culture. 2. Participation by males varies with species and culture. 3. Hormonal factors less important than experiential factors. 4. Parents, step-parents, grandparents, siblings, and others can participate in infant care. 212 - Behavioral Neuroendocrinology Parental Behavior - 21 People are just beginning to study the role of hormones in human parental behaviors Much of the research in this area has been done by Alison Fleming at the University of Toronto. She also did much of the pioneering work on olfaction and pup-induced maternal behavior. Fleming did her Ph.D. with Jay Rosenblatt at Rutgers. In general: 1. Women feel increasingly positive about and attached to their fetus/baby their pregnancies progress. But these changes do not appear to be related to changes in estradiol, progesterone, prolactin or cortisol levels during pregnancy. 2. Postpartum feelings of attachment and nurturance are associated with elevated estradiol/progesterone ratios during pregnancy. 3. Postpartum feelings of attachment and nurturance are associated with increased non-stressed circulating levels of cortisol. Positive about infant odors. More sympathetic to crying. Better able to identify their baby’s odors. Better able to identify their baby’s cries. Behavioral Neuroendocrinology - 213 Parental Behavior - 22 In general: 4. Fathers with lower testosterone levels are more sympathetic and feel a greater need to respond to infant cries. 5. Fathers with higher prolactin levels feel more positive about infants. 6. Fathers show increases in prolactin, cortisol, and testosterone after hearing infant cries. What’s all this mean? ????? Testosterone levels in men: unattached men > men with partners = married men. Domestication lowers testosterone levels in men? 214 - Behavioral Neuroendocrinology Parental Behavior - 23 Non-genomic transmission of maternal behavior in rats. Much of the research in this area was done by Michael Meaney at McGill University. Meaney never worked with Jay Rosenblatt. Meaney actually interested in the effects of stress during development. Rats who were repeatedly handled as infants show less reactivity to stress as adults than unhandled animals do. Less vocalizing, defecating, and freezing Dampened corticosterone responses to stress Not due to the ‘stress’ of being handled as pups Moms lick and groom their pups when they are returned to the nest As it turns out, it is the increased maternal licking and grooming that reduces reactivity to stress in adulthood Behavioral Neuroendocrinology - 215 Parental Behavior - 24 Individual differences in maternal behavior. In the absence of any handling, rat moms differ in how much they spontaneously lick and groom their pups. Pups raised by moms who display high levels of licking and grooming show lower reactivity to stress than do the offspring of moms who show low levels of licking and grooming. Pups raised by moms who display high levels of licking and grooming lick their own pups more than do the offspring of moms who show low levels of licking and grooming. Nature (genetics)? Or nurture? Cross-fostering experiment: Raise pups born to high-licking moms with low-licking moms and vice versa and then see how pups behave as adults. + high-licking mom ‘high’ pup* ‘low’ pup* high-licking mom high-licking mom + + low-licking mom ‘high’ pup* ‘low’ pup* low-licking mom low-licking mom + *Pup from a different mom 216 - Behavioral Neuroendocrinology Parental Behavior - 25 The level of maternal behavior a female rat shows is ‘culturally’ determined. Highly maternal moms beget highly maternal moms. Less maternal moms beget less maternal moms. Due to nurture, not nature. Is this the only rat behavior that is ‘culturally’ determined? No. Mother rats transmit their food preferences to their offspring. Rat pups learn to eat what their moms eat. In general, rats strongly prefer familiar foods to novel ones. (Called neophobia – a fear of new things.) If you give weanling (just weaned) rats a choice of foods to eat, they prefer the food(s) that their moms ate during lactation. You can vary the taste, smell, color, texture, etc., of the food that the moms eat. The weanlings prefer the foods that taste and smell like what their moms ate. To make a long story short, food gets stuck to the moms’ faces when they’re out of the nest eating. The pups then sample the foods and become familiar with them. In addition, pups become familiar with tastes in the amniotic fluid and the mother’s milk. Behavioral Neuroendocrinology - 217 Parental Behavior - 26 218 - Behavioral Neuroendocrinology Affiliation and Aggression - 1 Affiliation and Aggression Behavioral Neuroendocrinology - 219 Affiliation and Aggression - 2 220 - Behavioral Neuroendocrinology Affiliation and Aggression - 3 Affiliative and aggressive behaviors. 1. Pair bonding 2. Social recognition 3. Aggression(s) Pair bonding between male and female Relatively uncommon among mammals ~3% of all mammalian species ~15% of primates (including humans) Studied most intensively in voles (field mice) Several closely related species Different social patterns Behavioral Neuroendocrinology - 221 Affiliation and Aggression - 4 The cast of characters: Meadow vole (Microtus pennsylvanicus) Prairie vole (Microtus ochrogaster) Pine vole (Microtus pinetorum) Montane vole (Microtus montanus) The cast of characters: Live in boggy grasslands in northeast US Live in grasslands – widely distributed, including prairie Meadow vole (Microtus pennsylvanicus) Prairie vole (Microtus ochrogaster) Live in deciduous forests, not pine forests, in eastern US Live in boggy grasslands in mountain West Pine vole (Microtus pinetorum) Montane vole (Microtus montanus) 222 - Behavioral Neuroendocrinology Affiliation and Aggression - 5 Some species of voles form pair bonds. 1. Male and female pair up until death do them part If partner dies, only ~20% go on to form new pair bonds 2. Share living space and hang out together 3. Share pup care tasks (although the male may not be the father) Prairie vole 1. Form pair bonds Like to ‘cuddle’ Pine vole Meadow vole 1. No pair bonds Montane vole No cuddling – even in small cages 2. Puberty delayed as long as they live at home Females ovulate, mate and pair bond within 24 hr of exposure to urine from strange male Males pair bond with females and become aggressive toward strangers within 24 hr of mating 2. No social inhibition of puberty Behavioral Neuroendocrinology - 223 Affiliation and Aggression - 6 Pair bond formation – partner preference test: Prairie voles prefer to be with partner. Meadow voles prefer to be by themselves Seen in both sexes Pair-bonded males are also aggressive toward intruders – the resident intruder test: (behavior of toward new ) 224 - Behavioral Neuroendocrinology Affiliation and Aggression - 7 Prairie vole Pine vole Meadow vole Montane vole 3. Mom and dad both parental prairie & pine parental (also pair bond) 4. Young very distressed by separation Ultrasonic vocalizations Elevated cortisol 3. Only mom parental meadow & montane maternal Mom may split after 8-14 days 4. Young not distressed by separation Hormonal control of pair bonding in voles Behavioral Neuroendocrinology - 225 Affiliation and Aggression - 8 The major players: arginine (AVP) differ in just 2 amino acids (OT) Oxytocin (OT) and vasopressin (AVP) are the two peptides which induce pair bonding in prairie voles In this case, OT and AVP are acting as neurotransmitters rather than posterior pituitary hormones. Both OT and AVP projections and receptors are widely distributed in the brain. Vasopressin in gerbil brain 226 - Behavioral Neuroendocrinology Affiliation and Aggression - 9 OT controls pair bonding in female prairie voles, while AVP plays this role in males female ICV* OT in unmated animal ICV OT antagonist just prior to mating ICV AVP in unmated animal ICV AVP antagonist just prior to mating *Intracerebroventricular : induces behavior : inhibits behavior male --- --- Neither OT nor AVP affects mating behavior in males or females. Could differences in vasopressin and oxytocin in the brain that account for species differences in vole social behavior? Behavioral Neuroendocrinology - 227 Affiliation and Aggression - 10 AVP and OT distribution in vole brains: Oxytocin distribution and projections do not differ between monogamous and polygamous species, nor between males and females. Vasopressin distribution and projections do not differ greatly between monogamous and polygamous species. Infusion of vasopressin and oxytocin or their respective antagonists do not induce pair bonding in meadow voles (which don’t normally pair bond). Thus, the distribution of oxytocin and vasopressin cannot account for the species differences in pair bond formation. However, there are differences between the monogamous and polygamous species in vasopressin and oxytocin receptor concentration and distribution in both males and females. 228 - Behavioral Neuroendocrinology Affiliation and Aggression - 11 Oxytocin receptor distribution in female prairie vole vs. female meadow vole Higher oxytocin receptor concentration in the prelimbic cortex, nucleus accumbens and caudate-putamen in prairie vole Prairie Meadow Nucleus accumbens oxytocin receptor necessary for pair bond formation in female voles Thus, it is probably the species difference in neural oxytocin receptor levels, particularly in the nucleus accumbens, that accounts for the species difference in female pair bonding in voles Behavioral Neuroendocrinology - 229 Affiliation and Aggression - 12 In male voles, it is probably a difference in vasopressin receptor distribution that accounts for the differences in pair bonding behavior. Concentration of vasopressin receptor in brains of male prairie and meadow voles: Prairie: Meadow: Pair-bonding males have a greater concentration of vasopressin receptor in the ventral pallidum (VP), medial thalamus (MDThal) and amygdala (Amyg) 230 - Behavioral Neuroendocrinology Affiliation and Aggression - 13 Thus, it is the species difference in neural vasopressin receptor levels, particularly in the ventral pallidum, that accounts for the species difference in male pair bonding in voles So – can you make male meadow voles pair bond with females by adding vasopressin receptors to the ventral pallidum? Yes. Behavioral Neuroendocrinology - 231 Affiliation and Aggression - 14 Vasopressin receptor-hit Control gene Lim, et al., used a viral vector to insert the vasopressin gene or a different (control) gene into the ventral pallidum of male meadow voles. Vasopressin receptor-miss Vasopressin receptor-miss 232 - Behavioral Neuroendocrinology Affiliation and Aggression - 15 Male meadow voles paired with a sexually receptive mate for 24 hours, then placed in a partner preference test and observed for time spent with partner versus the novel female partner novel female control gene VP receptor misses VP receptor hits Hormones and pair bonding in people?? One can only speculate, because the work hasn’t been done. Oxytocin (and vasopressin to some degree) are released during orgasm, so one can make up stories about how experiencing an orgasm with someone might contribute to pair bonding. Of course this way of thinking has its limits, since people don’t develop pair bonds with vibrators or similar implements. Behavioral Neuroendocrinology - 233 Affiliation and Aggression - 16 Social recognition Seen and studied exhaustively in multiple species, including human beings. Neuroendocrinology studied in rodents. Functions of social recognition: 1. Kin recognition 2. Pair bonding 3. Endocrine phenomena such as pregnancy block and puberty 4. Dominance relationships 234 - Behavioral Neuroendocrinology Affiliation and Aggression - 17 Ways to measure social recognition in rats and mice: Animals are more interested in strangers than in someone familiar. Social recognition paradigm Test 1 Social discrimination paradigm Habituation-dishabituation paradigm Test 2 Test 3 Test 1 Test 1 Test 4 Test 2 Test 2 Test 5 Sex differences in social recognition in rats Females spend less time investigating strangers than males do, but they discriminate just as well – habituation-dishabituation paradigm no recognition And the females have better memories. Males forget by 90 minutes, but females still remember the familiar animal 3 hours later. (Higher ratios are worse. 1 = no discrimination) Compare this with voles who remember forever. (lower scores are better) Behavioral Neuroendocrinology - 235 Affiliation and Aggression - 18 Gonadal status and social recognition in rats Castration decreases time of investigation in males but not ability to discriminate T Investigation time decreases at estrus but not ability to discriminate Ovariectomy (OVX) increases investigation time but not ability to discriminate. Reversed by estradiol. Primates, including humans, primarily use vision and hearing for social recognition, but rodents use olfaction. Simply exposing a rat or mouse to the odor of a strange animal is sufficient to reduce investigation time when they are paired. Removing the olfactory bulbs greatly reduces, but doesn’t completely prevent, social recognition. Perhaps there are nonvolatile chemical signals that are received by the vomeronasal organ and accessory olfactory bulbs. Very little is known about the neuroanatomy of social recognition, but we know something about the hormones involved. 236 - Behavioral Neuroendocrinology Affiliation and Aggression - 19 Hormones and social recognition The major players: arginine (AVP) differ in just 2 amino acids (OT) Vasopressin facilitates social recognition in rats. Extends period of social recognition to > 2 hours in males (versus < 90 minutes in untreated beasties). Effective whether given systemically or intracerebroventricularly (ICV). Effective when put directly into the dorsolateral septum or olfactory bulbs. Behavioral Neuroendocrinology - 237 Affiliation and Aggression - 20 But this effect of vasopressin is not unique to social memory. AVP improves all kinds of memory, e.g.: Active avoidance Passive avoidance Conditioned taste aversions no novel taste, plus drug or placebo wait for hours sick? yes poison! food OK Tremendously useful in behavioral neuroscience Brattleboro rats. Naturally occurring mutant with vasopressin gene absent. Peeing and drinking problems because kidney can’t resorb water*. (They have diabetes insipidus. “Diabetes” means animal urinates a lot.) Exceptionally dull creatures. You do see them at the best colleges and universities world-wide. But not because of their intellectual gifts. Treating them with vasopressin reverses their memory (and peeing/drinking) impairments. *Vasopressin also known as antidiuretic hormone (ADH) 238 - Behavioral Neuroendocrinology Affiliation and Aggression - 21 Oxytocin and social memory The rat literature on the effects of oxytocin on social memory in rats is incredibly messy. Contradictions abound. Oxytocin can either facilitate or interfere with social memory in rats depending on the dose used and where it is administered. The situation is clearer in mice where oxytocin facilitates social memory. Infusion of an oxytocin antagonist (OTA) into the medial amygdala (but not the olfactory bulb) induces a ‘social amnesia’ in wild-type mice. Infusion of oxytocin into the medial amygdala (but not the olfactory bulb) reverses the ‘social amnesia’ of OT knockout mice. wild-type males Time spent investigating ‘familiar’ odor oxytocin knockout males Behavioral Neuroendocrinology - 239 Affiliation and Aggression - 22 Exposure of wild-type male mice to a social odor (a female) increases fos expression in the medial amygdala. But exposure of oxytocin knockouts (OTKO) male mice to the same odor does not increase fos expression. wild-type, medial amygdala OTKO wild-type Summary, affiliative behaviors: 1. Oxytocin and vasopressin act as neurotransmitters to affect pair bonding and social recognition. 2. Vasopressin facilitates pair bonding and aggression toward aliens in male prairie voles. 3. Oxytocin facilitates pair bonding in female prairie voles. 4. Species differences in pair bonding are associated with differences in the neural receptors for oxytocin and AVP. 5. Vasopressin facilitates social memory in rats and mice, but it facilitates other types of memory, too. 6. Oxytocin is crucial for social memory in mice, but its role in rats is confusing and contradictory. 7. Role of oxytocin and vasopressin in humans? We can make up interesting stories, but we don’t know. 240 - Behavioral Neuroendocrinology Affiliation and Aggression - 23 Aggressions 1. Parental aggression Maternal aggression usually starts around parturition Object(s) of aggression depend on species Usually induced by hormones that elicit maternal behavior, particularly oxytocin and prolactin Paternal aggression in males of biparental species For example, vasopressin in prairie and pine voles 2. Predatory aggression Males and females – independent of hormones Can be ‘recreational’ – cats, people Can be a highly organized group activity – wolves, lions Aggressions 3. Territorial aggression In order to obtain resources necessary to attract and keep mating partners (e.g., money?). Mostly seen in males – but sometimes females, too (e.g., hamsters, hyenas) Usually depends on androgens for expression Behavioral Neuroendocrinology - 241 Affiliation and Aggression - 24 Hormones and aggression In animals there is an overwhelming association between androgens and aggression. 1. In nearly all mammalian species, males are more aggressive than females. 2. Androgen levels and aggression increase at puberty. 3. Seasonal changes in testis function are associated with seasonal changes in aggression. 4. Castration usually decreases aggression, and androgen treatment reverses the effect of castration. Why do males fight? Usually for females. Indirectly. Actually for the resources necessary for reproduction More territory (resources, whatever) allows males to attract and support more females with whom to mate and to pass on their genes. 242 - Behavioral Neuroendocrinology Affiliation and Aggression - 25 For example, the red deer of Scotland are short day breeders. That is, they breed in the autumn, when days are getting shorter, so that their young will be born in the spring when food is plentiful. You can tell the stags (males), because they’re the ones with the antlers. For most of the year they are polite and well mannered. They play nicely in groups. They’re cool dudes, and all’s peaceful. Behavioral Neuroendocrinology - 243 Affiliation and Aggression - 26 But as summer progresses the stags’ testes wake up; the boys get horny – literally – and start behaving badly. No more nights out with the guys. No more male bonding. Just before they breed, males fight for the larger and better pastures – the kinds of places that draw babes (hinds in deerspeak). These fights can become quite intense and inflict serious damage. 244 - Behavioral Neuroendocrinology Affiliation and Aggression - 27 They breed, and over the winter the stags’ testes regress, and they stop fighting. In the spring, the hinds give birth, and all is again idyllic. What about females? Are they always less aggressive than males? Yeah, right. We all know better than that. Often aggression just takes a different form in females. Social rather than physical. For example, teenage or even preteen ‘alpha girls’ can inflict more damage on their peers than boys do – and without raising a finger. Behavioral Neuroendocrinology - 245 Affiliation and Aggression - 28 Is there hormone-related physical aggression in females, other than parental aggression? Sure there is. As they approach the time of ovulation monkeys become more assertive and fight more as they seek access to males. In some species of mammals, the sexual dimorphism in aggression is reversed and females are more aggressive than males. Well-known examples are Syrian (golden) hamsters and hyenas. Female Syrian hamsters do not play nicely in groups. • Solitary dwellers in the wild. • Females set up burrows in non-overlapping areas. • Defend their territories against intruders – male or female. • Female hamsters cannot be housed together in the lab. • Female are highly aggressive toward males, unless they are in heat. • Treatment with estradiol and progesterone inhibit aggression – until the female goes out of heat. • Not due to elevated androgens. 246 - Behavioral Neuroendocrinology Affiliation and Aggression - 29 Just for the record, not all kinds of hamsters are antisocial . . . for example, Siberian hamsters get along just fine. winter clothing (very toasty) summer clothing Then there are hyenas (and their friends, the vultures) Behavioral Neuroendocrinology - 247 Affiliation and Aggression - 30 • Live in large clans and defend territory at high density, but differ from all other social carnivores (e.g., wolves, coyotes): Spotted hyena • Clan members compete more and cooperate less than most social carnivores. • Females are bigger than males and dominate them. • Females compete for rank and food with one another. • Cubs are raised in communal dens, but are seldom or never provisioned or guarded by clan members; even close relatives do not cross-suckle offspring. • Males play no parental role – Spotted hyena only a privileged few are even permitted anywhere near dens, where juvenile offspring of highranking females dare to bully them. • During the few hours the female is fertile, several male hyenas can try to impregnate her. A female in heat can sometimes attract up to 15 male hyenas, who fight for the privilege to mate with her. 248 - Behavioral Neuroendocrinology Affiliation and Aggression - 31 • They make a lot of noise and show their teeth the whole time. The chosen ones approach her really carefully and show her their humbleness, with the head down and the tail between the legs to prevent them from being sent back. Forming a couple is an unknown concept for the hyenas. • Beginning only hours after birth, siblings of the same sex (particularly if they are both female) battle for dominance, biting each other and grabbing each other by the neck and shaking each other like two fighting adults. • The one that wins (firstborn has an advantage) can keep the other from nursing until it weakens and dies. This sibling rivalry kills an estimated 25% of all hyenas in their first month. • The surviving animal grows faster and is likelier to achieve reproductive dominance; the surviving female eliminates a rival for dominance in her natal clan. • There is no reproductive competition between siblings of opposite sex and consequently no killing. So why do hyenas do this? Behavioral Neuroendocrinology - 249 Affiliation and Aggression - 32 So how do female hyenas get to be so tough and aggressive? They have very high circulating androgen levels – during development and in adulthood. Is there some cost to having these high androgen levels? There sure is. Adult females have fully developed phalluses (clitorides) that are indistinguishable from males’ penises. Female hyenas copulate through their phalluses – no mean feat. 250 - Behavioral Neuroendocrinology Affiliation and Aggression - 33 Mating is challenging! Photos courtesy of Kay Holekamp, Michigan State University Female hyenas also give birth through their phalluses – yes, there’s lots of tearing. Ouch!!! Up to 10 percent of pregnant females may die in the process of delivering their first cubs. For those who survive, however, strong evolutionary benefits go to the females who can dominate other hyenas and the food supply. Behavioral Neuroendocrinology - 251 Affiliation and Aggression - 34 Androgens and aggression in human beings? As you would expect, direct evidence is hard to come by. However, if androgens did not increase aggression in humans, we would be unique among mammals. Aggression increases at puberty in boys. Surgical castration has been reported to decrease aggressiveness in men. • But this work had no controls. Sometimes sex offenders are ‘chemically castrated’ by giving them medroxyprogesterone acetate (MPA). • MPA acts as an antiandrogen by blocking androgen receptors • There are reports that MPA treatment decreases sex-related aggression but doesn’t affect other forms of aggression. • MPA is a progestin (Depo-Provera) used in contraception, but it is also an antiandrogen. • Androgens, glucocorticoids, and progestins all bind to each others’ receptors to some degree – e.g., RU486. • Estrogens pretty much only bind to estrogen receptors. 252 - Behavioral Neuroendocrinology Affiliation and Aggression - 35 There is a significant positive correlation between several measures of violence and circulating testosterone in prison inmates – male and female. • But . . . if this is causal, which is the cause, and which is the effect? Winning fights and exerting dominance increase testosterone levels. The bottom line is that androgens almost certainly increase aggression in humans, but the data are necessarily far from ideal. Treating hypogonadal men with testosterone improves their mood and sex drive, but it does not increase aggressiveness. Men who take anabolic steroids have been reported to be more aggressive/violent than non-users. • But these are cross-sectional, not longitudinal studies. • So there might be substantial self-selection. That is highly competitive, aggressive men may be more likely to choose to take anabolic steroids. • There is also the possibility of a placebo effect – the expectation that androgens will increase aggressiveness. • Just further evidence that humans make rotten experimental subjects. Behavioral Neuroendocrinology - 253 Affiliation and Aggression - 36 254 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 1 Hunger and Regulation of Energy Balance Behavioral Neuroendocrinology - 255 Hunger and Regulation of Energy Balance - 2 256 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 3 Neuroendocrinology of energy balance and food intake. “These little ones are mice. … These over here are hamsters. … Ooh! This must be a gerbil!” Obesity has become a huge public health problem in industrialized societies. It has become the most active area of research in endocrinology, including behavioral endocrinology. Weight reduction plans have become a major industry in the US. Pretty much any scheme works in the short term. But pretty much nothing works in the long term. Behavioral Neuroendocrinology - 257 Hunger and Regulation of Energy Balance - 4 The big issues for any motivated behavior: 1. What are the internal signals that elicit the behavior? 2. How/where is this information detected and converted to changes in neural activity? 3. What neural circuitry mediates the behaviors? 4. What are the signals from outside the creature that affect the behavior? Neuroendocrinologists mostly study questions 1-3, but the biggie is really question 4. The big issues for any motivated behavior: 1. What are the internal signals that elicit the behavior? 2. How/where is this information detected and converted to changes in neural activity? 3. What neural circuitry mediates the behaviors? 4. What are the signals from outside the creature that affect the behavior? 258 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 5 What are the internal signals that elicit the behavior? Something to do with the amount of energy available for oxidation and generation of ATP. So we have to know something about how the body handles metabolic fuels. An important fact of life is that, unlike grazing animals, we do not get a continuous flow of metabolic fuels, so we’re subjected to alternating periods of energy surfeit (after a meal) and deficit (before the next meal). Energy surfeit. Behavioral Neuroendocrinology - 259 Hunger and Regulation of Energy Balance - 6 Nearly all of the body’s energy comes from: Glucose: Fatty acids: Each bend is a carbon atom After being absorbed from the gut, a number of things can happen to glucose. 1. It can be taken up by cells and oxidized to generate ATP. wood + O2 glucose + O2 CO2 + H2O + a lot of heat CO2 + H2O + some heat + a bunch of ATP Biological systems are uniquely able to oxidize fuels and trap some of the energy that is released as ATP rather than losing all the energy as a bunch of heat. 260 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 7 Glucose oxidation: • all cells in the body use glucose for energy – all of them except the brain can use fatty acids, too • the brain is an obligatory oxidizer of glucose – it cannot use other metabolic fuels except under highly unusual circumstances (prolonged starvation) • insulin is required for glucose to leave the bloodstream and enter cells — except in the brain (mostly) • in the absence of insulin (diabetes mellitus) glucose cannot enter cells (except in the brain), so it builds up in the blood and causes hyperglycemia (high blood sugar) • diabetes mellitus – excessive urination – tastes sweet • type 1 diabetes – body can’t make insulin – autoimmune • type 2 diabetes – body can make insulin, but is insensitive to it (insulin resistance) – associated with obesity • diabetes insipidus – excessive urination – no taste • completely unrelated to diabetes mellitus • due to a deficit in the production of vasopressin (ADH) Behavioral Neuroendocrinology - 261 Hunger and Regulation of Energy Balance - 8 After being absorbed from the gut, a number of things can happen to glucose. 2. It can be linked with other glucose molecules and stored as glycogen. α α α β β β glycogen (starch) cellulose (wood) • the difference between glycogen and cellulose is simply the type of bond linking the glucose molecules. • glycogen is stored mostly in the liver and muscles – some glycogen is also stored in glial cells in the brain • insulin increases glucose entry into liver and muscle cells and stimulates glycogen storage After being absorbed from the gut, a number of things can happen to glucose. 3. It can be combined with other glucose molecules to form fatty acids. — lipogenesis (lipo=fat ; genesis=creation) • fatty acids are then combined with glycerol to form triglycerides – the primary storage form of fats + 3 fatty acids glycerol triglyceride • triglycerides can be stored in liver and muscle, but most are stored in adipose tissue 262 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 9 Glossary of glu- and gly- words glucose: a simple sugar that is a metabolic fuel for nearly all tissues in the body gluconeogenesis: synthesis of new glucose molecules from amino acids and other carbon sources glucoprivation: decreased availability of glucose for oxidation, e.g., hypoglycemia, 2DG treatments glycogen: storage form for glucose, many glucose molecules hooked end-to-end glycogenolysis: breakdown of glycogen and release of glucose molecules so that they can be oxidized glucagon: pancreatic counterregulatory hormone that opposes the actions of insulin After being absorbed from the gut, a number of things can happen to fatty acids. 1. They can be oxidized to produce ATP. • fatty acids + O2 CO2 + H2O + heat + a bunch of ATP • fatty acids can be oxidized anywhere in the body except the brain 2. They can be taken up by cells, combined with glycerol, and stored as triglycerides. + 3 fatty acids glycerol triglyceride Behavioral Neuroendocrinology - 263 Hunger and Regulation of Energy Balance - 10 Unlike the case with glucose, fatty acids can be taken up by cells, oxidized, and stored as triglycerides in the absence of insulin. Thus, in diabetics, fatty acids become the metabolic fuel of choice for most tissues other than brain, which is required to use glucose. Thus, when you are in positive energy balance, insulin is the most important hormone controlling metabolic fuel metabolism. When you start eating, your pancreas starts secreting insulin so that your body can handle the sudden onrush of metabolic fuels – cephalic phase response. Insulin facilitates glucose entry into cells where it can be oxidized or stored. Insulin lowers blood glucose levels. Insulin does not affect cellular fatty acid uptake, but it does inhibit release of fatty acids from fat cell storage depots. Thus, it decreases the availability of fatty acids for oxidation by other tissues. 264 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 11 Insulin secretion is controlled by: 1. Direct autonomic innervation (glucose sensors in brain). • Increased by parasympathetic output from brain – vagus nerve. • Decreased by sympathetic output from brain. 2. Glucose sensors in the pancreas. sympathetic – decreased insulin secretion parasympathetic increased insulin secretion vagus n. Behavioral Neuroendocrinology - 265 Hunger and Regulation of Energy Balance - 12 Energy deficit. So what happens when you don’t eat? • Stored metabolic fuels don’t do you any good whatsoever as long as they’re in storage. • Glycogen and triglycerides cannot be oxidized directly. They must be broken down to their constituent parts – glucose and fatty acids – and delivered to where they are needed. • A paramount principle of metabolic fuel metabolism is that glucose must be made available for the brain to use. • This breakdown and mobilization of stored fuels is mediated by a cluster of counterregulatory hormones – hormones that generally oppose the actions of insulin. 266 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 13 The counterregulatory hormones: 1. Epinephrine: • a catecholamine released from the adrenal medulla • acts on liver and muscle to break down glycogen and release glucose into the bloodstream, glycogenolysis – ‘lysis’ means to break down • acts on fat cells to break down triglycerides and release fatty acids and glycerol into the bloodstream, lipolysis The counterregulatory hormones: 2. Glucagon: • a 29-amino acid peptide released by the pancreas • glucagon from α-cells • insulin from β-cells • acts on liver and muscle to break down glycogen and release glucose into the bloodstream, glycogenolysis – like epinephrine, only slower • induces gluconeogenesis (gluco=glucose; neo=new; genesis=creation) – synthesis of glucose from amino acids – causes muscle wasting during prolonged starvation Behavioral Neuroendocrinology - 267 Hunger and Regulation of Energy Balance - 14 The counterregulatory hormones: 3. Cortisol/corticosterone: • a steroid – glucocorticoid – produced by the adrenal cortex, release controlled by ACTH • acts on liver and muscle to break down glycogen and release glucose into the bloodstream, glycogenolysis – like epinephrine, only slower • induces gluconeogenesis – synthesis of glucose from amino acids – causes muscle wasting during prolonged starvation Counterregulatory hormones protect the supply of glucose to the brain in two ways: 1. Epinephrine, glucagon, and cortisol raise blood sugar via glycogenolysis and gluconeogenesis. 2. Epinephrine mobilizes stored fatty acids from adipose tissue storage depots. • tissues which can burn either glucose and fatty acids selectively oxidize fatty acids during food deprivation • this spares glucose for utilization by the brain 268 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 15 insulin x inhibited by insulin epinephrine, cortisol, & glucagon So how do the neural circuits which control insulin, glucagon, epinephrine, cortisol, and hunger “know” whether enough calories are available for oxidation? The glucostatic hypothesis: • The brain monitors circulating blood glucose levels. When they fall below a critical level, we become hungry. After we eat and blood glucose levels return to normal, we are no longer hungry. Yeah, but what about diabetes? Diabetics are hyperglycemic, but they’re always hungry. Behavioral Neuroendocrinology - 269 Hunger and Regulation of Energy Balance - 16 • Oh . . . Well, it must be glucose utilization that’s important. Diabetics have high blood sugar levels, but it can’t get into the cells. Oh yeah. Remember . . . The brain doesn’t need insulin to burn glucose. • Hmmm . . . Well there must be brain cells that behave like liver cells and require insulin to use glucose. These cells must detect glucose utilization. Liver cells in the brain? Actually this is pretty close to the truth. So how do the neural circuits which control insulin, glucagon, epinephrine, cortisol, and hunger “know” whether enough calories are available for oxidation? The lipostatic hypothesis: • The brain monitors body fat stores. If they fall below a certain critical level it makes us hungry. After we’ve eaten and replaced the missing body fat, we’re not hungry anymore. Yeah, but four hours after I’ve had a meal, I’m hungry again, but I haven’t lost any weight. 270 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 17 • Oh, well the lipostatic hypothesis is more for the long term. Haven’t you noticed that laboratory animals’ body weights are very stable over long periods of time? Oh come on. That’s just because you give them the same boring Purina Rat (mouse, hamster, guinea pig, dog, monkey) Chow day after day. When I started giving my pet mouse, Squeaky, Twinkies every day, she got really fat. • Hmmm . . . Me too after that Krispy Kreme place opened next door. Well, if the brain isn’t paying attention solely to blood glucose levels or total body fat content, what else is a metabolic signal to eat? The short-term availability of oxidizable metabolic fuels. Short-term means minutes to hours. Fuels must be available for immediate oxidation; stored fuels don’t count. Behavioral Neuroendocrinology - 271 Hunger and Regulation of Energy Balance - 18 Pharmacological inhibition of metabolic fuel oxidation 2-deoxy-D-glucose (2DG) (glucoprivation) methyl palmoxirate (MP) (lipoprivation) Effects of metabolic inhibitors on food intake in rats vehicles 2DG MP 2DG+MP • treatment with 2DG alone increases food intake, despite the fact that it induces a severe hyperglycemia • treatment with MP alone increases food intake • treatment with 2DG+MP is far more effective than either drug alone 272 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 19 Animals don’t care where their calories are coming from carbohydrates or fats. They are just interested in the overall availability of oxidizable metabolic fuels. Hyperphagia (overeating) in diabetic rats, and what it tells us about metabolic fuel sensing. • Rats fed Purina Rat Chow are hyperglycemic and hyperphagic • In Purina Rat Chow, 90% of the non-protein calories come from carbohydrates • But diabetic rats can’t oxidize carbohydrates, so you might as well be feeding them wood chips. • If you feed diabetics a diet with a high fat content, they aren’t hyperphagic anymore, because you’re giving them a metabolic fuel that they can oxidize – fatty acids. Behavioral Neuroendocrinology - 273 Hunger and Regulation of Energy Balance - 20 Diet dilution studies with normal and diabetic rats. Normal rats diet High CHO Diabetics diet intake intake + + + (cellulose) +++++ + + (cellulose) High fat High CHO (diluted) High fat (diluted) +++ +++ +++++ +++ + + + = normal intake of food Animals don’t care where their calories are coming from carbohydrates or fats. They are just interested in the overall availability of oxidizable metabolic fuels. We don’t know how they do this at a cellular or molecular level – yet – very active area of research. Something to do with intracellular ATP levels? AMPK: AMP-activated protein kinase may function as a ‘fuel gauge’ or ‘metabolic master switch.’ When ATP is depleted, AMP builds up and activates AMPK, which phosphorylates other proteins Increased AMPK activity increases food intake and alters metabolism 274 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 21 The big issues for any motivated behavior: 1. What are the internal signals that elicit the behavior? 2. How/where is this information detected and converted to changes in neural activity? 3. What neural circuitry mediates the behaviors? 4. What are the signals from outside the creature that affect the behavior? Potential mechanisms for detecting metabolic fuel availability 1. Fuel detectors in the brain. 2. Fuel detectors in the periphery – neural communication with the brain. 3. Fuel detectors in the periphery – hormonal communication with the brain. Behavioral Neuroendocrinology - 275 Hunger and Regulation of Energy Balance - 22 Fuel detectors in the brain. • Sue Ritter from Washington State Univ. has been a leader in this area. • In order to determine where glucose availability is detected in the brain, she put tiny amounts of 5-thioglucose (5-TG)* into various parts of the brain and measured food intake and blood glucose levels. • Blood glucose levels are an index of counterregulatory hormone release, especially epinephrine. *5-TG blocks glucose oxidation, just as 2DG does. 5-TG in the CNS Everyone assumed that 5-TG implants in the hypothalamus would increase eating and blood glucose, because that was widely considered to be the center of the universe for the control of energy balance. They were wrong. 276 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 23 Instead, 5-TG implants in the caudal hindbrain increased both food intake and blood glucose levels. In addition, lesions of the area postrema and the nucleus of the solitary tract abolished glucoprivic hunger (eating in response to 5-TG or 2DG. APNTS hindbrain Therefore, the hindbrain contains cells that can monitor metabolic fuel (glucose) availability and control food intake and epinephrine release. To everyone’s surprise, the hypothalamus does not. Behavioral Neuroendocrinology - 277 Hunger and Regulation of Energy Balance - 24 Projections from the hindbrain to forebrain and spinal cord. Information about the availability of metabolic fuels gets from the hindbrain detectors to the rest of the CNS via cells which send their axons to the forebrain or spinal cord and use a catecholamine (epinephrine or norepinephrine) and neuropeptide-Y (NPY) as neurotransmitters. You can kill these cells by putting a compound called DSAP into their terminal fields. projections (axons) to spinal cord projections (axons) to forebrain other? PVN DSAP fuel detectors (CA/NPY) DSAP DSAP spinal cord MBH adrenal medulla hindbrain cells containing catecholamines (epinephrine/norepinephrine) and neuropeptide Y • Dopamine-β-hydroxylase (DβH) is necessary for the synthesis of norepinephrine and epinephrine. DβH PNMT epinephrine dopamine norepinephrine • DβH is released along with the neurotransmitter and then taken back up by presynaptic terminal button • Saporin-[anti-DβH] – an antibody to DβH coupled to the toxin, saporin or DSAP –locks onto the DβH while it is in the synaptic cleft and is taken up, too. saporin-[anti-DβH] + DβH saporin-[anti-DβH]-DβH • The saporin kills the catecholaminergic neuron once it gets inside. 278 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 25 Projections from the hindbrain to the forebrain. Infusion of anti-dopamine-β-hydroxylase (antibody to DβH) conjugated to the toxin, saporin, (DSAP) into catecholaminergic terminal fields destroys the neurons projecting to them. Taken up in synaptic buttons and transported backward to cell bodies (retrograde transport); not taken up by cell bodies. Injection of DSAP (but not saporin alone) into the spinal cord abolishes 2DG-induced hypergly-cemia but does not prevent glucoprivic eating. DSAP fuel detectors (CA/NPY) Injection of DSAP (but not saporin alone) into forebrain CA projection sites abolishes glucoprivic eating but does not prevent 2DGinduced hyper-glycemia. other? PVN DSAP DSAP spinal cord MBH adrenal medulla hindbrain cells containing catecholamines (epinephrine/norepinephrine) and neuropeptide Y Summary of DSAP experiments: Glucose-sensitive cells in the hindbrain send projections to either the forebrain or to the spinal cord. These cells use a catecholamine (epinephrine or norepinephrine) and neuropeptide Y as neurotransmitters. Destroying the cells that project to the forebrain with DSAP abolishes the increase in food intake that normally follows glucoprivation (2DG treatment). Destroying the cells that project to the spinal cord with DSAP abolishes the increase in blood glucose that normally follows glucoprivation (2DG treatment). Behavioral Neuroendocrinology - 279 Hunger and Regulation of Energy Balance - 26 Potential mechanisms for detecting metabolic fuel availability 1. Fuel detectors in the brain. 2. Fuel detectors in the periphery – neural communication with the brain. 3. Fuel detectors in the periphery – hormonal communication with the brain. The first thing that happens to food after it is absorbed is that it makes a pass through the liver. Nutrients infused directly into the hepatic portal vein decrease food intake – in doses which were ineffective when given into the general circulation Infusing 2DG directly into the hepatic portal vein increases food intake – in doses which were ineffective when given into the general circulation. 280 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 27 What happens if you induce an acute energy deficit and then selectively ‘feed’ the brain or the liver? • Animals were given a large dose of insulin to: 1) make them hungry and to 2) induce epinephrine release. • Treatment with brain foods – glucose and β-hydroxybutyrate – inhibited food intake and epinephrine release. • Treatment with liver foods – fructose and fatty acids – inhibited food intake but not epinephrine release. “brain foods” “liver foods” So the liver contains metabolic fuel detectors that are sensitive to the availability to glucose, fructose, and fatty acids – the general availability of oxidizable metabolic fuels. This information is conveyed to the brain via the vagus nerves. • Cutting the vagus nerves abolishes the effects of hepatic portal vein infusion of metabolic fuels and inhibitors. Behavioral Neuroendocrinology - 281 Hunger and Regulation of Energy Balance - 28 sympathetic parasympathetic vagus n. Potential mechanisms for detecting metabolic fuel availability 1. Fuel detectors in the brain. 2. Fuel detectors in the periphery – neural communication with the brain. 3. Fuel detectors in the periphery – hormonal communication with the brain. 282 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 29 Circulating levels of a number of hormones are highly correlated with body fat content. Most of the attention has focused on two hormones, insulin and leptin. Circulating insulin levels go up at mealtime and are also proportional to body fat content. So maybe high insulin levels feed back onto the brain to restrain appetite? Insulin receptors are widely distributed in the brain – including the hypothalamus and hindbrain. Early research showed that infusion of insulin into the cerebral ventricles decreased food intake in baboons and male rats. Thus it came to pass that the elders of feeding behavior spake unto the masses and decreed that circulating insulin levels controleth food intake. Behavioral Neuroendocrinology - 283 Hunger and Regulation of Energy Balance - 30 But then the heretics emerged. Quite a few labs failed to replicate the original findings with male rats. And eventually they started saying so in public. In early 2004, the lab that did the original work in male rats reported that the effect did not exist in females. So contrary to what you will read in most textbooks, the evidence that circulating insulin levels act directly on the brain to affect food intake is still spotty and inconclusive and may not apply to females at all. ~~~~~~~~~~~ ~~~ Leptin ~~~ ~~~~~~~~~~~ Two single-gene mutations causing extreme obesity. The Zucker fatty (fa/fa) rat. The obese (ob/ob) mouse. 284 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 31 The Zucker fatty (fa/fa) rat. The obese (ob/ob) mouse. • Hyperphagic even before weaning. • Increased body fat by the time they are weaned. • Grossly obese as adults (>50% body fat). • Multiple metabolic disorders. • Grossly obese even when not permitted to overeat. • Infertile. In 1993, Jeffrey Friedman at the Rockefeller University identified the gene responsible for the ob/ob mutation • Coded for a peptide hormone produced only in fat cells (adipocytes). • ob/ob mice failed to make the peptide. • Injection of peptide reversed the hyperphagia and reduced body weight in ob/ob animals. • Circulating levels of the peptide are directly proportional to body fat content • Friedman hypothesized that this was the long-sought-after lipostatic signal to the brain • Friedman named the peptide leptin - from the Greek leptos, meaning thin • Friedman sold the development rights to leptin to Amgen for $20,000,000 plus royalties when it came to market Behavioral Neuroendocrinology - 285 Hunger and Regulation of Energy Balance - 32 In 1995, Lewis Tartaglia at Millennium Pharmaceuticals in Cambridge identified the gene responsible for the fa/fa mutation in rats. • It was the leptin receptor. • Zucker rats actually have very high circulating leptin levels. • Receptor found throughout the body, including the hypothalamus and the hindbrain. • Gene therapy – inserting the leptin receptor gene into the brain using a viral vector – largely reverses the obesity of animals missing the leptin receptor gene. So was leptin worth the $20,000,000 that Amgen shelled out? Is it going to be the magic bullet that cures obesity? Well . . . no. Obese people and animals have very high circulating leptin levels – the fatter, the higher. Treating obese people and animals with more leptin doesn’t affect food intake and body fat content. People and animals with high leptin levels develop a leptin resistance, and they just stop responding. Only about five families have been identified world-wide with defects in the leptin gene. 286 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 33 Child with mutation of leptin gene – very rare. From: Farooqi IS, Matarese G, Lord GM, Keogh JM, Lawrence E, Agwu C, Sanna V, Jebb SA, Perna F, Fontana S, Lechler RI, DePaoli AM, O’Rahilly S 2002 J Clin Invest 110:1093–1103 So what does leptin do anyway? Look at it in an evolutionary perspective: • As mammals evolved, was obesity a major health problem? • No, but starvation was. • So if you look at leptin from that perspective, maybe it evolved in order to prevent starvation, not prevent obesity • The important signal may be low circulating leptin. • Maybe low leptin levels send the body a message: “Whoa dude, you got a major problem here, and you better do something about it.” • The body then makes the adjustments necessary to cope with the situation – increase food intake – decrease energy expenditure Behavioral Neuroendocrinology - 287 Hunger and Regulation of Energy Balance - 34 One last hormone from the periphery: Ghrelin (pronounced GREL-in) • Ghrelin is a peptide hormone that is produced and secreted by cells in the stomach. ghrelin blood ghrelin • Circulating ghrelin levels rise just before meals and fall afterward. • Injection of rats and humans with ghrelin increases hunger. time of day • There are receptors for ghrelin in the hypothalamus and hindbrain. In general, our bodies do a far better job of defending against starvation than against obesity. Indeed, a tendency to fatten and store extra calories during times of plenty was probably a distinct advantage as we evolved. It is only during very recent that this ability to store calories has become a liability. And that’s the reason that obesity is about to pass smoking as the major preventable cause of mortality. 288 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 35 The big issues for any motivated behavior: 1. What are the internal signals that elicit the behavior? 2. How/where is this information detected and converted to changes in neural activity? 3. What neural circuitry mediates the behaviors? 4. What are the signals from outside the creature that affect the behavior? Until the mid-1980s the neurobiology of food intake was dominated by the dual-center hypothesis. According to the dual-center hypothesis, there was a lateral hypothalamic ‘feeding center’ which stimulated hunger. Animals with LH lesions became aphagic. That is, they stopped eating. This LH ‘feeding center’ was in turn inhibited by a ventromedial hypothalamic ‘satiety center.’ Animals with VMH lesions became hyperphagic. That is, they ate voraciously until they became quite obese. lateral ventromedial hypothalamus hypothalamus (LH) (VMH) Behavioral Neuroendocrinology - 289 Hunger and Regulation of Energy Balance - 36 The dual-center hypothesis is a wonderful (?) example of how a simple, intuitive explanation of a phenomenon can be extremely appealing and become widely accepted by the scientific and lay communities. It is also an example of how a simple, intuitive explanation of a phenomenon can set research back by 30 years by leading people to assume that a problem is solved. Just because it makes sense doesn’t mean that something is true, because the dual-center hypothesis was dead wrong. A few of the things that are wrong with the dual-center hypothesis: Lateral hypothalamus: • Aphagia not due to lesions of the LH, per se. Actually due to interruption of nigrostriatal bundle which is just passing through. • The animals are sick as hell. Ventromedial hypothalamus: • Hyperphagia and obesity not due to lesions of the VMH, per se. Actually due to interruption of ventral noradrenergic bundle which is just passing through. • Primary deficit is not overeating; it is hypersecretion of insulin. • Animals become obese even if they don’t overeat. • Cutting the vagus nerves prevents the hyperinsulinemia and the hyperphagia. • Animals overeat because they’re becoming obese, not vice versa. 290 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 37 “It’s O.K. to sleep with an hypothesis, but you should never become married to one.” J. William Langston The New York Times, Week in Review, p. 6, October 12, 1997 OK, so what’s the latest thinking on the neurobiology of food intake? There are quite a few amine and peptide neurotransmitters involved, but the three current favorites are, neuropeptide Y, AGRP, and α-MSH. • α-MSH is a melanocortin that inhibits food intake • brain levels of α-MSH increase in overfed animals • infusion of α-MSH into the hypothalamus or hindbrain inhibits food intake • there are melanocortin receptors in the hypothalamus and hindbrain (and elsewhere) • around 5% of all human obesities appear to be due to mutations of the melanocortin or melanocortin receptor genes – and that’s a lot Behavioral Neuroendocrinology - 291 Hunger and Regulation of Energy Balance - 38 • NPY and AGRP both increase food intake. • brain levels of NPY and AGRP increase in fooddeprived animals • infusion of NPY and/or AGRP into the hypothalamus or hindbrain increases food intake • there are receptors for NPY and AGRP in the hypothalamus and hindbrain (and elsewhere) • NPY and AGRP are colocalized in the same neurons and are secreted simultaneously • NPY has its own receptors (several kinds in fact) • AGRP binds to the melanocortin receptor – the same receptor that binds α-MSH • AGRP is actually a melanocortin (α-MSH) antagonist • α-MSH is a melanocortin that inhibits food intake • NPY and AGRP both increase food intake. • AGRP is actually a melanocortin (α-MSH) antagonist 292 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 39 The neural circuitry controlling food intake and energy metabolism extends through a large part of the brain. But two areas have gotten most of the attention: • the arcuate nucleus of the hypothalamus • the paraventricular nucleus of the hypothalamus arcuate nucleus paraventricular nucleus Here’s how it’s supposed to work: + _ NPY+ AGRP (↑ FI) _ _ α-MSH (↓ FI) Behavioral Neuroendocrinology - 293 Hunger and Regulation of Energy Balance - 40 1. Circulating, ghrelin, leptin and insulin act in the arcuate nucleus to signal the status of body energy availability. 2. Arcuate neurons producing α-MSH or NPY/AGRP project to neurons in the vicinity of the paraventricular nucleus. 3. Then a miracle occurs, we adjust our food intake and energy expenditure appropriately, and maintain an ideal body fat content. + _ NPY+ AGRP (↑ FI) _ _ α-MSH (↓ FI) 294 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 41 Challenges for the central dogma: 1. It’s questionable whether insulin, per se, provides any useful information about body energy status. 2. The role of leptin is still unresolved. 3. Lesions of the arcuate nucleus have little effect on energy balance. 4. Knocking out the NPY gene has little effect on energy balance. 5. Knocking out the AGRP gene has little effect on energy balance. 6. Knocking out the AGRP and NPY genes has little effect on energy balance. 7. What about the hindbrain? NPY+ NPY AGRP AGRP (↑ FI) _ _ + _ α-MSH α-MSH (↓ FI) The system is way more complicated than any current working model – with a huge amount of redundancy built in. Behavioral Neuroendocrinology - 295 Hunger and Regulation of Energy Balance - 42 The bottom line is that we’re a long way form curing obesity 1. Natural selection has shaped us to cope with a world where calories are scarce, not plentiful. • when calories are easily available, metabolic need is not a primary determinant of food intake • similarly, the acquisition cost of food has dropped – we don’t have to exert a lot of energy to survive 2. Obesity is a large family of diseases with a common symptom: too much body fat. • we will have to come up with multiple cures, because there are so many different causes 3. Pretty much any weight-reduction plan works short term. But 90-95% fail in the long term. — in the Gonadal steroids and regulation of energy balance • Estrogens, progestins, and androgens all affect food intake, energy expenditure, and body composition • In male rats, growth curves are quite smooth, and body weight doesn’t vary much from day to day • But in females, body weight fluctuates, and growth curves are scalloped, with decreases in body weight every 4 or 5 days • These weight fluctuations are due to cyclic changes in the females’ circulating estradiol and progesterone levels • Ovariectomy eliminates the cyclic changes in body weight – and makes them fat 296 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 43 Food intake drops and running wheel activity peaks at estrus – when circulating estradiol levels are high. From Lisa Eckel, Florida State University To make a long story short: 1. Estrogens cause behavioral and physiological changes which cause weight loss. 2. Progestins have little or no effects of their own, but they oppose the effects of estrogens. Behavioral Neuroendocrinology - 297 Hunger and Regulation of Energy Balance - 44 Changes in energy balance across rats’ estrous cycles: Pregnancy Estrus Food intake Exercise Body weight Diestrus OVX Treatment of rats with estradiol (EB) alone reverses the changes in energy balance caused by OVX • a transient decrease in food intake • a lasting increase in voluntary exercise • a lasting decrease in body weight – as long as you keep giving estradiol 298 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 45 The changes in body weight after ovariectomy (OVX) and treatment with estradiol (EB) are due almost entirely to changes in body fat content. Treatment of OVX rats with progesterone alone has no effect at all on energy balance. But when it is given with estradiol, progesterone can cancel out all of the effects of estradiol. • increased food intake • decreased voluntary exercise • increased body weight and fat content Behavioral Neuroendocrinology - 299 Hunger and Regulation of Energy Balance - 46 The antiestrogen, tamoxifen, blocks the effects of estradiol on sexual behavior, but mimics its effects on body weight. vehicle estradiol tamoxifen estradiol + tamoxifen The antiestrogen, tamoxifen, blocks the effects of estradiol on uterine weight, but mimics its effects on body fat (WAT). 300 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 47 Currently, multiple pharmaceutical companies are working to develop selective estrogen receptor modulators (SERMS). These are compounds which block harmful effects of estrogens (such as tumor growth) but mimic the beneficial effects (such as preventing hot flashes and osteoporosis). Estradiol acts at multiple sites throughout the body to affect energy balance • Acts in brain to alter food intake and energy expenditure • Exact sites of action unclear • Estrogen receptor in all the right places • Acts in liver to alter carbohydrate and lipid metabolism • Liver contains high levels of estrogen receptor • Acts in adipose tissue to alter fatty acid uptake, storage, and release • Adipose tissue contains estrogen receptor Behavioral Neuroendocrinology - 301 Hunger and Regulation of Energy Balance - 48 What about ovarian hormones and energy balance in women? • There are certainly changes in hunger, food preferences, and fat storage during pregnancy • During the first trimester of pregnancy, when estradiol levels are elevated, women may experience nausea and (sometimes dramatic) changes in food preferences • Later in pregnancy, when progesterone levels are elevated, appetite and fat storage increase What about ovarian hormones and energy balance in women? • There are changes in energy levels, hunger, and food cravings across the menstrual cycle. • Much of the literature is sloppy, poorly controlled, and contradictory. • There are a few well-controlled studies that report slight decreases in appetite and increases in energy expenditure during the follicular stage of the cycle, but these effects are pretty subtle. • There is no credible evidence that athletic performance varies as a function of the menstrual cycle. 302 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 49 What about ovarian hormones and energy balance in women? • After ovariectomy or menopause women tend to gain weight and body fat. • The most dramatic change is that women change from being “pears” to “apples.” menopause Postmenopausal hormone replacement therapy? • The literature on the effects of postmenopausal hormone replacement therapy is messy, because everyone uses different hormone treatments. • Estrogen alone vs. estrogen + progestin • Naturally occurring vs. synthetic steroids • Differing doses • The best available data indicate that treatment with estrogens alone is effective in preventing the menopausal changes in body fat distribution. • Concurrent treatment with progestins seems to undo the beneficial effects of estrogen replacement therapy on body weight. Behavioral Neuroendocrinology - 303 Hunger and Regulation of Energy Balance - 50 Androgens are anabolic steroids — they increase muscle mass and body weight. • Skeletal muscle contains androgen receptors, and androgens cause muscle growth • In rats, it is testosterone, not an aromatized or 5αreduced metabolite, that causes muscle growth • Androgen treatment is used therapeutically in muscle-wasting diseases • Synthetic androgens are widely misused by athletes to gain a performance advantage, but there’s a significant placebo effect, too • Aromatized metabolites of testosterone act directly on adipose tissue to decrease body fat content Placebo effect of ‘anabolic steroids” ‘anabolic steroid’ group Performance start ‘treatments’ ‘placebo’ group baseline Time (weeks) 304 - Behavioral Neuroendocrinology Hunger and Regulation of Energy Balance - 51 Behavioral Neuroendocrinology - 305 Hunger and Regulation of Energy Balance - 52 306 - Behavioral Neuroendocrinology Nutrition and Fertility - 1 Nutrition and Fertility Behavioral Neuroendocrinology - 307 Nutrition and Fertility - 2 308 - Behavioral Neuroendocrinology Nutrition and Fertility - 3 Nutritional infertility: the effects of food availability on reproduction. Why bother to look at the effects of food availability on reproduction? Behavioral Neuroendocrinology - 309 Nutrition and Fertility - 4 “Of the many environmental factors that can influence a mammal’s reproduction, food availability must be accorded the most important role.” F. H. Bronson, Mammalian reproductive biology. Chicago: University of Chicago Press, 1989, p. 88. Nutrition and fertility in women: People have known for a long time that there is an association between adequate nutrition and fertility. Name: Venus of Willendorf Hometown: Willendorf, Austria Age: ~30,000 years Occupation: Fertility Goddess 310 - Behavioral Neuroendocrinology Nutrition and Fertility - 5 Without sufficient calories, women become amenorrheic – they no longer exhibit menstrual cycles and do not ovulate. Nutritional amenorrhea is common in the Third World, where calories may be hard to come by. But it also happens in developed societies where ample supplies of food are available: • Eating disorders. • Endurance athletes. • Distance runners. • Athletes who limit their food intake. • Ballerinas. • Gymnasts. • Figure skaters. • Military basic training. Nutritional infertility can be seen in both sexes, but it is far more common among females than males. Why? Even among biparental species, the female expends far more energy than the male during a complete cycle of reproduction – from mating to weaning the offspring. She has far more to lose from a botched reproductive attempt due to insufficient calories – the loss of the litter/baby, or even her life. Behavioral Neuroendocrinology - 311 Nutrition and Fertility - 6 In and of itself, nutritional infertility is not a pathology. • It is a perfectly reasonable adaptation to the prevailing environmental conditions. • It allows the female to avoid wasting time and energy on a futile, and possibly dangerous, activity. • It is completely reversible. There is no harm whatsoever to the reproductive system. So what’s the fuss about nutritional amenorrhea? Osteoporosis and metabolic disorders. When food is abundant, animals can maintain all physiological processes at optimal levels, but when calories are in short supply, animals are forced to set priorities. Some processes can’t be compromised Others can be reduced Reproduction and fat storage have a very low priority 312 - Behavioral Neuroendocrinology Nutrition and Fertility - 7 In hamsters, as in other species, both ovulatory cycles and estrous behavior are affected by metabolic fuel availability. Ovulatory cycles: Lordosis duration (sec) Estrous behavior: 200 150 100 50 Fed ad lib. Fooddeprived The primary change in nutritional suppression of ovulatory cycles is a decrease in GnRH release from the hypothalamus. The pituitary works just fine. • treatment with exogenous GnRH elicits normal release of LH The ovaries work just fine. • treatment with exogenous LH elicits normal follicular growth and ovulation Behavioral Neuroendocrinology - 313 Nutrition and Fertility - 8 GnRH, and consequently LH, is secreted in a pulsatile fashion. • The ovaries will not respond to a sustained level of LH – that is, without pulses. • In general, it is the LH pulse frequency, not the amplitude, that determines ovarian function. • Ovulation can be prevented by giving females a steady, non-pulsatile, treatment with an LH analog – contraceptive potential. The inhibition of ovulatory cycles is reflected in a suppression of pulsatile luteinizing hormone (LH) secretion. Rhesus monkeys: 314 - Behavioral Neuroendocrinology Nutrition and Fertility - 9 It’s the same in women, too. Four days of food restriction decreases LH pulse frequency in women. Subject 1: 45 kcal/kg LBM Subject 2: 10 kcal/kg LBM This inhibition of GnRH secretion is due in part to increased negative feedback sensitivity to estradiol. The inhibition of estrous behavior by food deprivation is due in part to decreased neural responsiveness to estradiol. Which may be secondary to decreases in VMH estrogen receptor. Estrous behavior: Lordosis duration (sec) 200 150 100 50 Fed ad lib. Fooddeprived Behavioral Neuroendocrinology - 315 Nutrition and Fertility - 10 So how does this work? The big issues for any motivated behavior: 1. What are the internal signals that elicit the behavior? 2. How/where is this information detected and converted to changes in neural activity? 3. What neural circuitry mediates the behaviors? 4. What are the signals from outside the creature that affect the behavior? 316 - Behavioral Neuroendocrinology Nutrition and Fertility - 11 So how do the neural circuits controlling GnRH secretion and female copulatory behavior ‘know’ whether fuels are available? What is the metabolic signal? One hypothesis was that female mammals had to maintain a critical level of body fat or fat-to-lean ratio in order to remain fertile, and the problem was thought to have been solved, circa 1970. This critical body fat hypothesis is simple, intuitive, elegant, and wrong much like the lipostatic hypothesis of body weight regulation. And it set research in the field back by ~ 20 years. Behavioral Neuroendocrinology - 317 Nutrition and Fertility - 12 A number of lines of evidence cast doubt on the critical body fat hypothesis. • Continued cycling in very lean women and animals. • The immediate resumption of pulsatile LH secretion after refeeding. • The original data which led to the critical body fat hypothesis were flawed. If body fat levels are not the critical signal, then what is? What is the alternative? 318 - Behavioral Neuroendocrinology Nutrition and Fertility - 13 Two papers, both published in 1986, were crucial to deciphering the answer to this question One was the paper by Tordoff and Friedman on metabolic inhibitors and food intake. Maybe reproduction is like food intake, and the critical signal is the short-term availability of oxidizable metabolic fuels. So what happens if you give metabolic inhibitors instead of food depriving the animals? Behavioral Neuroendocrinology - 319 Nutrition and Fertility - 14 But the metabolic inhibitors were either very expensive (2DG), or they couldn’t be bought at all – you had to go to drug companies and grovel in order to get some (MP). So giving then for weeks at a time was out of the question. The second paper was by Larry Morin and showed that just two days of food deprivation, starting immediately after estrus, prevented the next expected estrus and ovulation in female hamsters. 320 - Behavioral Neuroendocrinology Nutrition and Fertility - 15 Estrous Behavior: Food deprivation inhibits estrous behavior in ovariectomized hamsters – and so does treatment with metabolic inhibitors. (Li et al., 1994) Intravenous treatment with 2DG inhibits pulsatile release of LH in sheep and other species. Behavioral Neuroendocrinology - 321 Nutrition and Fertility - 16 Some of the reasons why body fat content can’t be the metabolic signal controlling fertility: • Extremely lean animals and women can continue to ovulate. • When food-deprived animals are refed, LH pulses are restored within hours – before body fat content changes. • When metabolic fuel availability is restricted rapidly using metabolic inhibitors, LH pulses are inhibited immediately – before body fat content changes. The big issues for any motivated behavior: 1. What are the internal signals that elicit the behavior? 2. How/where is this information detected and converted to changes in neural activity? 3. What neural circuitry mediates the behaviors? 4. What are the signals from outside the creature that affect the behavior? 322 - Behavioral Neuroendocrinology Nutrition and Fertility - 17 Well, again we learned something from the work on the neuroendocrinology of eating. Sue Ritter’s work showed that 5TG acted in the hindbrain to increase food intake, and lesions of the area postrema and nucleus of the solitary tract abolish glucoprivic eating. APNTS hindbrain The same appears to be true for nutritional control of ovulatory cycles and female copulatory behavior. Hamsters with lesions of the area postrema show perfectly normal estrous behavior when deprived of metabolic fuels – no inhibition. 2DG+MP treatment: Food deprivation: (Li et al., 1994) Behavioral Neuroendocrinology - 323 Nutrition and Fertility - 18 The fact that AP lesions prevent the inhibition of female copulatory behavior by multiple nutritional challenges (food deprivation, 2DG/MP, etc.) rules out the possibility that metabolic fuel deprivation has its effects simply by making the animals ill. The effects of metabolic inhibitors on LH secretion are also mediated by the hindbrain. Infusion of 2DG into the fourth ventricle inhibits pulsatile LH secretion. 324 - Behavioral Neuroendocrinology Nutrition and Fertility - 19 What about peripheral fuel detectors and neuronal transmission to the brain? It can’t work that way, because metabolic challenges do not affect reproduction in animals with AP lesions, and AP lesions wouldn’t interfere with peripheral fuel detection. What about peripheral fuel detectors and hormonal transmission to the brain? • Insulin? • Leptin? • Ghrelin? Insulin? Untreated diabetics on high-carbohydrate diets are infertile. • There are plenty of metabolic fuels around, but they’re not available for oxidation. • Insulin treatment restores fertility in diabetics, but it does so by making glucose available for oxidation – not via direct action in the brain. • Insulin treatment doesn’t affect LH release in foodrestricted animals, but it should if it were a signal of adequate nutrition. Behavioral Neuroendocrinology - 325 Nutrition and Fertility - 20 Leptin? Animals without leptin or leptin receptors are infertile – ob/ob mice and fatty Zucker rats. • Treatment of ob/ob mice, but not fatty Zucker rats restores fertility. • The ob/ob mice don’t make leptin, whereas fatty Zucker rats don’t make functional leptin receptors. • Leptin has permissive effects on reproduction – some is necessary, but normal fluctuations in the physiological range seem to be unimportant. Intravenous treatment with 2DG inhibits pulsatile release of LH in sheep and other species – too fast to be explained by leptin. 326 - Behavioral Neuroendocrinology Nutrition and Fertility - 21 Food deprivation or treatment with metabolic inhibitors inhibits estrous behavior in ovariectomized hamsters, but 2DG+MP treatment does not reduce leptin levels (Li et al., 1994) In hamsters that are refed following 48 hours of food deprivation, sexual receptivity is restored more rapidly than circulating insulin or leptin levels. Behavioral Neuroendocrinology - 327 Nutrition and Fertility - 22 Ghrelin? Ghrelin increases hunger, so one would predict that it should inhibit reproduction. But it doesn’t – it actually stimulates sexual receptivity in female hamsters. The big issues for any motivated behavior: 1. What are the internal signals that elicit the behavior? 2. How/where is this information detected and converted to changes in neural activity? 3. What neural circuitry mediates the behaviors? 4. What are the signals from outside the creature that affect the behavior? 328 - Behavioral Neuroendocrinology Nutrition and Fertility - 23 What if NPY and catecholamines convey metabolic fuel information from the visceral hindbrain to the forebrain for both reproduction and food intake? Stimulates food intake and inhibits reproduction??? If this is the case, then infusion of NPY or catecholamines into the forebrain should inhibit estrous behavior. Forebrain infusion of catecholamines does not inhibit estrous behavior, so they can’t be involved. But NPY agonists suppress lordosis in Syrian hamsters Behavioral Neuroendocrinology - 329 Nutrition and Fertility - 24 If NPY mediates the effects of food deprivation on LH release, then infusing NPY into the brain should decrease circulating LH levels, and it does. (NPY infused into the third ventricle) Infusion of saporin conjugated to an antibody to DβH (DSAP) into the terminal fields of catecholaminergic neurons destroys those neurons without causing damage at the injection site. So infusion of DSAP into the forebrain should block the effects of metabolic inhibitors on reproduction. 330 - Behavioral Neuroendocrinology Nutrition and Fertility - 25 Treatment with 2DG does not inhibit ovulation in rats previously given DSAP in the forebrain. Comparable experiments looking at sexual behavior have not been done. Vehicle 2DG Thus, NPY projections from the hindbrain to the forebrain transmit information about food availability to the circuits that control estrous behavior and GnRH secretion. So what happens next? ? Behavioral Neuroendocrinology - 331 Nutrition and Fertility - 26 What does NPY do after it is released in the forebrain? Juli Jones: (smart grad student): Maybe it acts on neurons that produce corticotropin-releasing hormone (CRH) as a neurotransmitter, and then CRH inhibits reproduction. • NPY projections from the hindbrain synapse on forebrain CRH neurons. • CRH neurons send axons to parts of the forebrain that play crucial roles in the control of both estrous behavior and LH secretion. George: (old geezer professor): Yuck! CRH is one of those stress hormones. You don’t to mess with that. Juli: Thanks for your input; I’m going to do it anyway. If NPY inhibits female copulatory behavior by activating neurons producing CRH, then infusion of CRH into the forebrain should mimic the effects of food deprivation or NPY and suppress lordosis. 332 - Behavioral Neuroendocrinology Nutrition and Fertility - 27 Infusion of CRH into the lateral ventricles causes a transient (< 4 hr) inhibition of estrous behavior in hamsters. It also inhibits pulsatile LH secretion. (Jones et al., 2002) If food deprivation is detected in the hindbrain, and this information is conveyed to the forebrain by NPY which then acts via stimulation of CRH neurons, then infusion of a CRH antagonist into the brain should block the effects of food deprivation or NPY and restore lordosis. Behavioral Neuroendocrinology - 333 Nutrition and Fertility - 28 Infusion of a CRH antagonist, astressin, into the lateral ventricles blocks the effects of CRH estrous behavior in hamsters. (Jones et al., 2002) Infusion of astressin, a CRH receptor antagonist, prevents the suppression of estrous behavior by NPY – and reverses the effects of food deprivation. Lordosis duration (sec) 150 100 50 (Jones et al., 2002) 334 - Behavioral Neuroendocrinology Nutrition and Fertility - 29 Similarly, infusion of the CRH antagonist, α-helical CRH, into the forebrain reverses the suppression of LH pulses by food deprivation. Reproduction shuts down when calories are scarce. Here’s how we think it works, at least for estrous behavior: Then a miracle occurs. Behavioral Neuroendocrinology - 335 Nutrition and Fertility - 30 Is putting CRH into the brain just an elaborate way of stressing the animals, and it’s actually stress that inhibits sexual behavior? Three hours of physical restraint raises cortisol levels in female hamsters, but it doesn’t affect estrous behavior. 336 - Behavioral Neuroendocrinology Nutrition and Fertility - 31 There is no evidence in the literature that psychological stress affects copulatory behavior in any species. • May even be a way to cope with stress or grief for some. • May act as an aphrodisiac – risky sex. Treatment with astressin ‘cures’ nonresponders. Behavioral Neuroendocrinology - 337 Nutrition and Fertility - 32 Summary: 1. A decrease in the availability of oxidizable metabolic fuels inhibits both ovulatory cycles and copulatory behavior. • Absolute levels of intake and expenditure do not matter – it is the balance between the two that is crucial • Stored calories, such as fatty acids, are useless – unless they can be mobilized and oxidized 2. The availability of fuels for oxidation is monitored by neurons in the hindbrain. Summary: 3. This information is conveyed to the forebrain circuits controlling GnRH secretion and estrous behavior synaptically – probably by NPY-producing cells. 4. NPY could inhibit GnRH release via direct synapses on the GnRH neurons or indirectly via CRH. 5. NPY inhibits estrous behavior via CRH . 338 - Behavioral Neuroendocrinology Nutrition and Fertility - 33 Behavioral Neuroendocrinology - 339 Nutrition and Fertility - 34 340 - Behavioral Neuroendocrinology Biological Rhythms and Seasonal Reproduction - 1 Biological Rhythms and Seasonal Reproduction Behavioral Neuroendocrinology - 341 Biological Rhythms and Seasonal Reproduction - 2 342 - Behavioral Neuroendocrinology Biological Rhythms and Seasonal Reproduction - 3 Biological rhythms and seasonal reproduction. Many species breed only at certain times of year – in order to give birth when ample food is available Hamsters: Spontaneous recovery in the spring the summer Breed during the summer Stop breeding when days get short in fall Behavioral Neuroendocrinology - 343 Biological Rhythms and Seasonal Reproduction - 4 Quite a few seasonal rhythms are cued by photoperiod (day length), because photoperiod is the single most reliable environmental index of seasonal change. Days are longer in the summer than in the winter, and animals can detect these differences in photoperiod. The further you live from the equator, the greater the difference between summer and winter. Maine NC Peru Some seasonal rhythms cued by photoperiod (day length): Reproduction – maybe humans, too. Before artificial lighting Depression – seasonal affective disorder. Energy balance Some species fatten in preparation for winter, others lose weight 344 - Behavioral Neuroendocrinology Biological Rhythms and Seasonal Reproduction - 5 Animals use circadian rhythms to keep track of day length. (circa = around, dies = day) The two important parts of any rhythm are the amplitude (how much it changes) and the period (time required for a full cycle, τ ). For circadian rhythms, τ ≈ 24 hours. Level of some physiological variable τ( Properties of circadian rhythms: They are endogenous – generated from within the animal – not by external events. They are approximately (not exactly) 24 hours in length, τ ≈ 24 hr. Behavioral Neuroendocrinology - 345 Biological Rhythms and Seasonal Reproduction - 6 Measuring daily activity patterns in hamsters. Hamsters are nocturnal – active at night. Animals that are active during the day are diurnal. DAY NIGHT If running wheel activity reflects a circadian rhythm, then it should persist in constant conditions. And it does. If running wheel activity reflects a circadian rhythm, then the period should be roughly, but not exactly, 24 hours. And it is. day & night constant conditions constant darkness = 24 τ τ > 24 346 - Behavioral Neuroendocrinology Biological Rhythms and Seasonal Reproduction - 7 Another example: Reentrainment Resetting the light-dark cycle phase-shifts the running rhythm τ >24.1 Activity free runs in constant In constant conditions, conditions, and is always τ is activity free-runs andclose to 24 hr, .g., 23.9 < >24.1 alwayseclose to 24 hr, e.g., 23.9 < τ >24.1 Humans have daily rhythms, too. And some of them are circadian. Behavioral Neuroendocrinology - 347 Biological Rhythms and Seasonal Reproduction - 8 OK we have this internal endogenous clock . . . So where is it? It turns out that we have little clocks all over our bodies, but the biggie (the master clock) is in the suprachiasmatic nucleus (SCN) of the hypothalamus. What’s the evidence that the SCN is the master clock? 1. If you lesion the SCN, animals become arrhythmic. They run at all different times of day with no organized pattern. free-running SCN-X arrhythmic 348 - Behavioral Neuroendocrinology Biological Rhythms and Seasonal Reproduction - 9 2. There is a 24-hour rhythm in fos expression in the SCN. CT=clock time But the SCN could just be driven by the “real clock” rather than being the clock itself. clock SCN behavior 3. Yeah, but if you remove the animal’s brain, take a slice containing just the SCN, and do electrical recording in vitro, there is still a circadian rhythm in electrical activity. Behavioral Neuroendocrinology - 349 Biological Rhythms and Seasonal Reproduction - 10 SCN lesion 4. If you lesion an animal’s SCN, it becomes arrhythmic. Then if you transplant the SCN from another animal, the lesioned creature becomes rhythmic again. transplant 0 24 48 5. And if the host and donor had different τ’s to start with, the host takes on the donor’s τ. Tau mutant (τ ≈ 18 hr) Tau mutant with wildtype SCN transplant (τ ≈ 24 hr) Wild-type (τ ≈ 24 hr) Wild-type with Tau mutant SCN transplant (τ ≈ 18 hr) 350 - Behavioral Neuroendocrinology Biological Rhythms and Seasonal Reproduction - 11 The SCN is the body’s master clock and can generate circadian rhythms in the absence of any external input. But under normal conditions, all of our circadian rhythms are coordinated with (entrained to) the lightdark cycle. Re-entrainment happens when the environment is light at a time when the hamster expects it to be dark. Normal: runs just in dark Starts early - starts later next day Runs late - stops earlier next day If you put animals in 23.5-hour (or 24.5-hour) photoperiods, their rhythms will entrain to that schedule. This works just fine as long as the photoperiod doesn’t deviate too much from 24 hours. So how does the SCN know whether the lights are on or off? Behavioral Neuroendocrinology - 351 Biological Rhythms and Seasonal Reproduction - 12 Information about the light-dark cycle gets to the SCN via a direct projection from the retina – the retinohypothalamic tract. The retinohypothalamic tract is completely separate from the pathways mediating vision. optic nerve eye (optic chiasm-where the two optic nerves cross and enter the brain) (retinohypothalamic tract – nothing to do with vision) Seasonal reproduction. 352 - Behavioral Neuroendocrinology Biological Rhythms and Seasonal Reproduction - 13 OK, so all nature’s creatures have got rhythm in the form of these endogenous ~24 hour rhythms. But how does this result in yearly rhythms of reproduction? Do the creatures just count days until they get to 365? No, they don’t, but a very few species (e.g., marmots, ground squirrels) actually have endogenous circannual (~ a year) rhythms. • If you put them into constant conditions they free-run with a period of around 11 months. • Their circannual rhythms become entrained to seasons just as circadian rhythms become entrained to the daily light-dark cycle. • But this is pretty rare Most seasonal breeders use circadian timing mechanisms to measure photoperiod and turn the reproductive system on or off at the appropriate time of year. Behavioral Neuroendocrinology - 353 Biological Rhythms and Seasonal Reproduction - 14 For example, Syrian (golden) hamsters need more than 12 hours of light per day to remain reproductively active. But Siberian hamsters, which live further to the north, need more than 15 hours of light per day to remain reproductively active. Why? Syrian hamster Siberian hamster (hypothetical) The pineal gland mediates the effects of photoperiod via secretion of its hormone, melatonin. melatonin tryptophan serotonin melatonin 354 - Behavioral Neuroendocrinology Biological Rhythms and Seasonal Reproduction - 15 In nonmammalian vertebrates, the pineal is actually a third eye. It contains photoreceptors and is sensitive to light. This is not the case in mammals, but they still respond to light . . . but photic information gets to the pineal via a circuitous neural pathway from the eyes. NE sympathetic nerve (active during light – inhibits melatonin) (superior cervical ganglion) If you pinealectomize hamsters, they no longer become reproductively inactive in short photoperiods – they assume that they’re always in long days. pinx Behavioral Neuroendocrinology - 355 Biological Rhythms and Seasonal Reproduction - 16 On the other hand, if you house intact hamsters in long days and give them daily injections of melatonin within a couple of hours before lights-out, their gonads will regress. But if you give the melatonin at other times of day, it has no effect. This led to a great deal of confusion for a while. inhibits reproduction: long day short night no inhibition: long day short night The pineal secretes melatonin only at night. So animals in short days (long nights) produce more melatonin than animals in long days (short nights). So giving exogenous melatonin at the right time effectively turns the long-day pattern into a shortday pattern and inhibits reproduction. - Summer melatonin - Winter 356 - Behavioral Neuroendocrinology Biological Rhythms and Seasonal Reproduction - 17 So how do melatonin injections make animals think that they are in short days when they are actually in long days? • more melatonin? • longer duration? - Summer melatonin - Winter More melatonin? Pinealectomize animals and infuse melatonin in long-day (short night) pattern– but in different doses. If it is the amount of melatonin that is critical, then the higher dose should inhibit reproduction. But dose doesn’t matter, so it isn’t the amount of melatonin. no inhibition infusion no inhibition infusion short duration short duration Behavioral Neuroendocrinology - 357 Biological Rhythms and Seasonal Reproduction - 18 Longer duration of melatonin? Pinealectomize animals and infuse melatonin in long-day pattern and short day (long night) pattern – giving the same total amount. If it is the duration of melatonin that is critical, then longer infusions should inhibit reproduction. And they do, so it is melatonin duration that counts. no inhibition infusion reproduction inhibited infusion short duration long duration (but low amplitude And that’s why giving melatonin at other times of day doesn’t inhibit reproduction in hamsters housed in long days. It doesn’t increase the duration of melatonin exposure, because it is cleared rapidly. melatonin melatonin 358 - Behavioral Neuroendocrinology Biological Rhythms and Seasonal Reproduction - 19 Thus, it is the duration of the daily rise in melatonin levels, as measured by circadian timing mechanisms, that tells seasonal breeders what time of year it is. This work was independently in two labs. • By Bruce Goldman at UConn using Siberian hamsters. • Long-day breeders. • Breed when melatonin duration is short. • By Eric Bittman at the University of Michigan (now of the UMass Biology Dept.) in sheep. • Short-day breeders. • Breed when melatonin duration is long. You can trick an animal housed in short days into acting like it is in long days simply by exposing it to a brief (≥ 10 seconds) light pulse in the middle of the night. This works by suppressing melatonin levels and shortening the period of nocturnal elevation. light pulse Tricks short-day animal into thinking it’s in long days. Behavioral Neuroendocrinology - 359 Biological Rhythms and Seasonal Reproduction - 20 In hamsters, recovery of reproductive function in the spring actually begins well before day length reaches 12 hr. After ~12 weeks in short days, animals become unresponsive (refractory) to melatonin, and their gonads recover spontaneously. 12-hr days So where is the nocturnal melatonin signal detected? There are quite a few parts of the brain that contain melatonin receptors, but there is substantial variation between species. This leaves the possibility that melatonin acts on different parts of the brain to affect reproduction in different species. 360 - Behavioral Neuroendocrinology Biological Rhythms and Seasonal Reproduction - 21 Seasonal change in neural responses to negative feedback In hamsters, short photoperiods (long melatonin durations) inhibit reproduction by suppressing LH (GnRH) pulses and by inhibiting (male and female) copulatory behavior simultaneously. The inhibition of LH secretion is due to increased sensitivity of the hypothalamus to the negative feedback actions of steroid hormones. The inhibition of sexual behavior is due to decreased sensitivity of the hypothalamus to the actions of steroid hormones. So the hypothalamus is simultaneously more sensitive and less sensitive to the actions of gonadal hormones. The same thing happens when reproduction is inhibited by underfeeding. Photoperiod, melatonin, and body weight Behavioral Neuroendocrinology - 361 Biological Rhythms and Seasonal Reproduction - 22 Syrian (golden) hamsters fatten when placed in short (winter) photoperiods (< 12 hr light) or treated with melatonin. And they lose this extra weight when they are returned to long (summer) photoperiods or melatonin is withdrawn. spontaneous reversal On the other hand, Siberian hamsters lose weight when placed in short photoperiods (< 9 hr light). If you displace their body weights by under- or overfeeding, they return to the level appropriate for that time of year. 48 Body weight (g) 44 40 36 0 2 long day short day 4 Weeks 6 8 10 12 362 - Behavioral Neuroendocrinology Biological Rhythms and Seasonal Reproduction - 23 The bottom line is that many species – maybe including humans, too – show seasonal changes in body weight and regulation of energy balance. But not everyone fattens in preparation for winter. Behavioral Neuroendocrinology - 363 Biological Rhythms and Seasonal Reproduction - 24 364 - Behavioral Neuroendocrinology Stress - 1 Stress Behavioral Neuroendocrinology - 365 Stress - 2 366 - Behavioral Neuroendocrinology Stress - 3 Stress. So, what is stress, anyway? Defining stress is somewhat like trying to define pornography. “I shall not today attempt further to define the kinds of material [pornography] . . . but I know it when I see it.” US Supreme Court Justice Potter Stewart, 1964 Behavioral Neuroendocrinology - 367 Stress - 4 Physiologists make the distinction between a stressor and the stress response. A stressor is anything that disturbs physiological balance. • It can be a physical insult, like being bitten by a dog, or a psychological insult, like a growling dog – or even worrying that a dog might growl at you. The stress response is the body’s physiological adaptations used to cope with the stressor. So we’ll admit that we can’t come up with a rigorous definition of stress – although we each know it when we see it. Instead, we’ll talk about the stress response and how that affects us. 368 - Behavioral Neuroendocrinology Stress - 5 This way of thinking of stress is derived from the physiological concept of homeostasis (Claude Bernard, 1865) Homeostasis is the process by which the body maintains a constant internal environment. • Examples include things like blood glucose, body temperature, intra- and extracellular fluid volumes, blood pH, etc. So when physical or psychological stressors throw the body out of whack, the stress response is an attempt to restore homeostasis. There are many physiological responses to stressors. They are pretty much the same, regardless of the nature of the stressor, for example: • Chasing after prey. • Being chased by a predator. • Worrying about being chased by a predator. Their purpose is to prepare the body for ‘fight or flight.’ 1. Increased mobilization of metabolic fuels for energy. 2. Increased oxygen supply so fuels can be oxidized. 3. Faster delivery of fuels an oxygen to where they’re needed. 4. Inhibition of anything that interferes with any of the above. Behavioral Neuroendocrinology - 369 Stress - 6 A hallmark of the stress response is an activation of the adrenal glands – both the cortex and the medulla. • Release of corticosterone (or cortisol) from the cortex. • Release of epinephrine from the medulla. • Glucocorticoids and epinephrine are commonly referred to as “stress hormones” This fact has led to some confusion, because it became a circular definition. Circular definition. Meaning: The definition includes the term being defined as a part of the definition. Examples: • An animal is human if and only if it has human parents. (The term being defined is “human”. But in order to find a human, we would need to find human parents. To find human parents we would already need to know what a human is.) • A book is pornographic if and only if it contains pornography. (We would need to know what pornography is in order to tell whether a book is pornographic.) • You are experiencing stress if you release stress hormones. (We need to know what stress is before hormone we can define stress hormones.) 370 - Behavioral Neuroendocrinology Stress - 7 The stress response actually involves an activation of the entire sympathetic nervous system. The net effect is to cope with an emergency – actual or perceived. increases vasopressin release decreases insulin release increases glucagon release OK, so the stress response is a great thing. It prepares me to do battle with the forces of evil and survive to tell about it. But I thought that stress was a bad thing. It makes people sick, doesn’t it? Yeah … well … if you have to be picky about it … nothing’s perfect. If you’re gonna get all technical, long-term stress makes you more vulnerable to other things that can make you sick. Stress itself doesn’t make you sick. Behavioral Neuroendocrinology - 371 Stress - 8 There is a difference between humans and animals here. • In both humans and animals acute stress is beneficial. • After the stressor has abated, animals can put it behind them and move on. (What, me worry?) • The problem with people is that they often fail to put it behind them. • We worry and stew and fret about things in our lives – things that don’t pose an immediate threat to life and limb – things that might not even happen. • Some of this can be beneficial if it helps us prepare for upcoming events in our lives, e.g., exams. • But chronic activation of the physiological stress responses can be damaging. Some of the bad stuff that chronic stress does to you: • Muscle wasting and fatigue. • Cardiovascular disease. • Gastrointestinal disorders – maybe. • Inhibition of growth. • Inhibition of reproduction. • Increased vulnerability to pathogens (germs). • Brain damage and cognitive dysfunction 372 - Behavioral Neuroendocrinology Stress - 9 Muscle wasting and fatigue. The stress response does a great job of mobilizing stored metabolic fuels for oxidation – epinephrine. But if you do this over and over again, eventually you run out of fuel, and your body starts to do what it can to save calories – and causes fatigue. More important, corticosteroids are very effective at inducing gluconeogenesis – converting amino acids from protein into glucose. Basically, your body starts eating it’s own proteins. It is likely that this paradox is related to the different roles of catecholamines (epinephrine and norepinephrine) and the glucocorticoids in the stress response. The catecholamines act immediately at the time of the stressor to make metabolic fuels available. In contrast, the glucocorticoid response is delayed – peaking 1-2 hours after the stressor and can last a long time. The purpose of the glucocorticoid-induced gluconeogenesis is to replenish the glucose used up in the acute response. And that ends up ‘eating’ muscle. Behavioral Neuroendocrinology - 373 Stress - 10 Cardiovascular disease. The stress response does a great job of increasing heart rate and force of contraction and raising blood pressure in order to deliver fuels and oxygen to where they’re needed. But if you do this over and over again, it causes heart disease and atherosclerosis. Gastrointestinal disorders – maybe. • Historically, chronic stress has been associated with gastric ulcers, and it became accepted as dogma that chronic stress was the leading cause of ulcers. • But acute stress inhibits gastric function, including acid secretion • In the 1980’s John Warren and Barry Marshall proposed that ulcers were actually caused by a bacterium, Helicobacter pylori. Not surprisingly, this hypothesis was widely ridiculed. • But they were right, and now the notion that stress causes ulcers is generally dismissed by physicians. (From one extreme to another.) 374 - Behavioral Neuroendocrinology Stress - 11 • In 2005, Warren and Marshall were awarded the Nobel Prize for Physiology or Medicine for their discovery. • The most parsimonious hypothesis at this time is that chronic stress decreases resistance to H. pylori. Behavioral Neuroendocrinology - 375 Stress - 12 Inhibition of growth. Acute stress stimulates growth hormone release. But in the long term, stress actually inhibits growth hormone, resulting in disorders of growth. Prolonged stress in childhood can lead to dwarfism. J.M. Barrie is a good example of this. Decreased growth hormone in adulthood can lead to decreased tissue repair and osteoporosis. (Rodney Harrison of the Patriots took growth hormone to help him recover from off season surgery.) Inhibition of reproduction. • Chronic stress inhibits ovulatory cycles via suppression of pulsatile LH release. • The literature is kind of confusing because of differing definitions of stress. • Insulin-induced decreases in metabolic fuel availability. • Psychological stressors such as restraint. • The two seem to act via different neural circuitry but both may involve CRH. • Intracerebroventricular (ICV) infusion of CRH inhibits pulsatile release of LH. • ICV infusion of a CRH antagonist can prevent the inhibition of LH pulses by various stressors. 376 - Behavioral Neuroendocrinology Stress - 13 Acute restraint stress inhibits pulsatile LH release in female rats, and this effect is blocked by a CRH antagonist. (vehicle) (CRF antag.) Marshall et al., 2005 Inhibition of reproduction. • Chronic stress also seems to inhibit LH pulsatility by decreasing anterior pituitary sensitivity to GnRH. • This is mediated in part by vasopressin. • Posterior pituitary release of vasopressin helps increase blood volume during stress. • Hypothalamic neurons that produce CRH also produce vasopressin and release it into the hypophyseal portal system – where it can inhibit LH release by acting in the anterior pituitary • Also due to glucocorticoid feedback on the anterior pituitary. Behavioral Neuroendocrinology - 377 Stress - 14 Inhibition of reproductive behaviors? • Despite what you read in Cosmo and the supermarket tabloids, there is almost no literature whatsoever on the effects of stress on sexual behaviors. • Nothing in primates. • One paper in hamsters. • Restraint stress just before behavioral testing raised cortisol levels but didn’t affect lordosis duration. Human sexual behavior? • Despite what you read in Cosmo and the supermarket tabloids, there is almost no literature whatsoever on the effects of stress on sexual behaviors. • Anecdotal reports suggest that acute stress might even have aphrodisiac properties – e.g., fear of getting caught, having just escaped from a threat, a family tragedy, etc. • Sexual desire, and even fertility, persist in the face of terrible stressors – e.g., concentration camps. 378 - Behavioral Neuroendocrinology Stress - 15 Increased vulnerability to pathogens (germs). Chronic stress suppresses the immune system. Primarily due to the actions of corticosteroids (cortisol and corticosterone). This fact can be put to use clinically to combat autoimmune diseases such as lupus, rheumatoid arthritis, and multiple sclerosis. Type I diabetes is an autoimmune disease, so why wouldn’t you treat diabetes with corticosteroids? So why would you want stress to suppress the immune system? • Lots of clever ideas – but they have all fallen by the wayside. • To make a long story short, the immediate effect of stress is to enhance immune function – good. • The delayed, glucocorticoid-mediated, response functions to bring immune activity back down to ‘normal’ levels – good. • Unless you put the brakes on the immune response you are at risk for autoimmune diseases – bad. • But then chronic elevation of glucocorticoid levels ends up suppressing immune activity to below optimal levels – bad. Behavioral Neuroendocrinology - 379 Stress - 16 So what are the implications of all this for human disease? • It turns out to be pretty confusing. • Compared with something like AIDS, glucocorticoid-mediated immunosuppression is pretty modest. • So you might be more likely to get a cold after finals week, but the best available evidence indicates that you are unlikely to be highly vulnerable to infection. • There is no evidence that stress-induced immunosuppression plays a role in cancer. The work suggesting this link is sloppy and poorly controlled. • The notion that a positive attitude is of value in fighting off cancer is most likely erroneous, too. • Physicians can do their patients a serious disservice by erroneously suggesting that they can help control their cancer this way. Brain and cognitive dysfunction • As with pretty much everything else with stress, the effects on the brain and cognitive function depend on the duration of exposure. Short-term is good, and long-term is bad. • In the short term, stress improves cognitive function. • This is beneficial if it enables an animal to remember some threatening event better. • Both catecholamines and glucocorticoids contribute to this improvement in memory. 380 - Behavioral Neuroendocrinology Stress - 17 Short-term effects of catecholamines on cognition • Adrenalectomy impairs performance on both positivelyand negatively-reinforced tasks. • Treatment of adrenalectomized animals with epinephrine partly reverses this impairment – indicating that it was partly due to removal of adrenal medulla. • Moderate doses are more effective than either high or low doses. • Treatments are most effective when given immediately after the reinforcer. • Not clear whether action is on memory formation directly or is secondary to effects on arousal and attention. • Effects of epinephrine on cognitive function are probably mediated via the amygdala. • But there’s a problem here. Peripherally released epinephrine from the adrenal medulla doesn’t cross the blood-brain barrier. • It turns out that the epinephrine is acting at peripheral adrenergic receptors. • Which communicate with the nucleus of the solitary tract in the hindbrain via the vagus nerves. • Which then signals the amygdala using norepinephrine as a neurotransmitter. adrenal medulla Epi peripheral adrenergic receptors vagus NTS NE amygdala Behavioral Neuroendocrinology - 381 Stress - 18 Finally, stressor-induced peripheral catecholamine release probably facilitates cognitive function by increasing glucose availability to the brain. Hypoglycemia can cause cognitive impairment and confusion. Short-term effects of glucocorticoids on cognition • Adrenalectomy impairs performance on both positivelyand negatively-reinforced tasks. • Treatment of adrenalectomized animals with corticosteroids partly reverses this impairment – indicating that it was partly due to removal of adrenal cortex. • Moderate doses are more effective than either high or low doses. 382 - Behavioral Neuroendocrinology Stress - 19 In this case it turns out that this phenomenon is mediated by two types of glucocorticoid receptors – cleverly named type I and type II. Receptor occupancy Increasing occupation of type I receptors improves performance Increasing occupation of type II receptors improves, then impairs, performance With cognitive function, the relevant glucocorticoid receptor targets seem to lie in the amygdala and hippocampus. As is the case with epinephrine, it isn’t clear whether glucocorticoids affect cognitive performance by altering learning directly or by changing attention and arousal. Behavioral Neuroendocrinology - 383 Stress - 20 Long-term effects of glucocorticoids on cognition Repeated or prolonged exposure to high levels of corticosteroids impairs cognitive performance and damages the brain. Rodents, primates, and humans. This is the case whether the high circulating corticosteroid levels are due to repeated stress or to therapeutic use of synthetic corticosteroids. – Prednisone. Three weeks of exposure to elevated corticosteroid levels causes a reversible atrophy of the dendrites of pyramidal cells in the hippocampus in rats. This means that they make fewer synaptic connections, and this leads to impairments in spatial ability and short-term memory tasks. Due to increased release of excitatory amino acids, glutamate and aspartate, which can be toxic. Can be prevented by treatment with Dilantin, an antiepileptic drug which blocks receptors for the excitatory amino acids. Repeated seizures can have the same effects on the hippocampus and cognitive performance as prolonged stress. 384 - Behavioral Neuroendocrinology Stress - 21 Prolonged exposure to elevated corticosteroids, whether from stress or therapeutic use, also reduces the number of granule cells in the hippocampus – neurons critical for normal cognitive function. Dogma used to be that adults could not produce new neurons – we could only lose them as we age. We now know that this is not true. Some parts of the nervous system are constantly generating new cells (neurogenesis) and turning them over by a process called apoptosis, or programmed cell death. High levels of corticosteroids inhibit neurogenesis, the production of new neurons, in less than an hour, and prolonged elevations can result in measurable cell loss and cognitive decline. Individual differences in the stress response Regardless of the species, individuals differ in the frequency and intensity of stress responses. Even when they are exposed to the same stressors. Example: Two monkeys are deprived of food for a day, but they exhibit different levels of circulating corticosteroids. • Stress physiologists would look for a physiological explanation – e.g., availability of metabolic fuels, pituitary sensitivity to CRH, clearance of ACTH, etc. • But if two monkeys are food deprived, and one of them is given access to a non-nutritive flavored solution, that animal will not show the expected rise in corticosteroids. This means that you have to take into account “psychological factors,” whatever those are. Behavioral Neuroendocrinology - 385 Stress - 22 Psychological factors The same event is not always equally stressful in a single individual, nor is it necessarily stressful in different individuals. What kinds of things affect the magnitude of the stress response? • Control: being able to exert some control over an aversive event lessens the stress response. • Predictability: being able to predict the occurrence of an aversive event lessens the stress response – even if it is inevitable. • Outlet for frustration: being able to react to the aversive event lessens the stress response. Early experience and the stress response Briefly removing rat pups from their mothers dampens the magnitude of the corticosteroid response to stress in adulthood. • Also decreases emotionality in stressful situations. • Actually due to increased maternal licking of pups when they are returned – not to infantile stress. Late prenatal stress (stressing the mom) or prolonged neonatal stress of the pups increases the magnitude of the adult stress response. • Also inhibits adult male copulatory behavior. 386 - Behavioral Neuroendocrinology Stress - 23 Behavioral Neuroendocrinology - 387 Stress - 24 388 - Behavioral Neuroendocrinology Thirst - 1 Thirst Behavioral Neuroendocrinology - 389 Thirst - 2 390 - Behavioral Neuroendocrinology Thirst - 3 Neuroendocrinology of water balance and thirst. “Dogs that drink from the toilet bowl – right after this message” Obviously, we get water from drinking, but where does it go? Urine and feces. Sweat (both water and salt). Insensible water loss through evaporation from mouth and skin (just water). (Don’t give your dog Gatorade.) Behavioral Neuroendocrinology - 391 Thirst - 4 Thirst (water balance) is more tightly regulated than other motivated behaviors such as mating and eating. Why? You’re much more likely to die from dehydration than from not having sex. Unlike calories, it is impractical to carry large stores of water around with you. Physiological controls of thirst: what is regulated: Body water is found in two compartments. • About two-thirds of the body’s water is found inside cells – intracellular fluid* • The rest is found outside cells – extracellular fluid • Interstitial fluid – in tissue surrounding cells • Intravascular fluid – in the bloodstream* • Cerebrospinal fluid *critical regulated compartments 392 - Behavioral Neuroendocrinology Thirst - 5 The amount of fluid in the two compartments is regulated separately. The body carefully monitors and controls both the intracellular fluid and the blood volume Too little water in cells disrupts chemical reactions; too much can cause the cell to burst Too little water in the blood stream interferes with circulation – low blood pressure Too much water in blood stream – high blood pressure Interstitial fluid volume is not tightly regulated, e.g., edema Intracellular volume is controlled mainly by the concentration of dissolved substances (solutes) in the interstitial fluid When the concentration of solutes dissolved in the interstitial fluid is the same as inside the cell, it is said to be isotonic. (iso = same as) When the concentration of solutes dissolved in the interstitial fluid is higher than inside the cell, it is said to be hypertonic. (hyper = greater than) When the concentration of solutes dissolved in the interstitial fluid is lower than inside the cell, it is said to be hypotonic. (hypo = less than) Behavioral Neuroendocrinology - 393 Thirst - 6 The laws of physics and chemistry require that the tonicity of the interstitial fluid and the intracellular fluid be the same – isotonic. This is accomplished by having water, not solutes, move in and out of the cell. Why? Because water can pass freely across the cell membrane, but most solutes cannot – barriers, ion pumps, etc. – it is semipermeable. A hypertonic interstitial fluid will draw water out of the cells. A hypotonic interstitial fluid will force water into the cells. Here’s how it works isotonic solutions dissolved molecules (solutes) 394 - Behavioral Neuroendocrinology Thirst - 7 Here’s how it works add solute (salt, for example) dissolved molecules (solutes) Here’s how it works hypotonic solution hypertonic solution Membrane won’t allow solutes to move from one side to the other, so water has to move. dissolved molecules (solutes) Behavioral Neuroendocrinology - 395 Thirst - 8 Here’s how it works hypotonic solution hypertonic solution H2O dissolved molecules (solutes) Here’s how it works hypotonic solution hypertonic solution isotonic solutions dissolved molecules (solutes) 396 - Behavioral Neuroendocrinology Thirst - 9 It’s the same in the body inside of cell outside of cell inside of cell outside of cell eat salt Isotonic solutions osmosis osmosis movement of a solvent through a semipermeable membrane (as of a living cell) into a solution of higher solute concentration that tends to equalize the concentrations of solute on the two sides of the membrane osmotic pressure the pressure that develops in a solution separated from a solvent by a membrane permeable only to the solvent Behavioral Neuroendocrinology - 397 Thirst - 10 There are two kinds of thirst: Osmotic (cellular) thirst is a response to the loss of water from within the cells – cell shrinkage Just need to replace water Volemic (extracellular) thirst is a response to the loss of blood or sweat – loss of water and solutes Need to replace both water and solutes Osmotic (cellular) thirst: Osmotic thirst occurs when you: a) lose only water, as through evaporation from mouth or skin, or b) consuming a solute which cannot enter the cell, e.g., sodium salts. Each of these increases the tonicity of the interstitial fluid – makes it hypertonic. The common feature is that water is drawn out of cells, and they shrink. 398 - Behavioral Neuroendocrinology Thirst - 11 salty water Drinking salty water increases the tonicity of the extracellular fluid, which in turn, draws water out of the cells and causes them to shrink. You can also increase the tonicity of the extracellular fluid by losing just water – e.g., by breathing or drooling. Osmoreceptors, which detect cell shrinkage, are located in a number of tissues, including the small intestine and the liver. The most important for regulation of fluid balance and thirst are located in the organum vasculosum of the lamina terminalis (OVLT) in the brain. Behavioral Neuroendocrinology - 399 Thirst - 12 Dehydration (shrinkage) of cells in the OVLT increases neuronal firing. This information is conveyed to the median preoptic nucleus and then the supraoptic and paraventricular nuclei of the hypothalamus. Neurons in the SON and PVN increase their rate of firing, which causes release of antidiuretic hormone (a.k.a. vasopressin) from the posterior pituitary. Antidiuretic hormone acts on the kidneys to increase water retention. median preoptic nucleus supraoptic & paraventricular nuclei posterior pituitary OVLT ADH kidney Increased neuronal firing in the OVLT increases thirst at the same time it increases ADH secretion. So we react to cellular dehydration by decreasing water loss in the urine and by drinking more. lateral hypothalamus median preoptic nucleus supraoptic & paraventricular nuclei thirst OVLT posterior pituitary ADH kidney 400 - Behavioral Neuroendocrinology Thirst - 13 Volemic thirst: You can lose a tremendous amount of body water without causing cellular dehydration, simply by losing salt at the same time. Hemorrhage and sweating, both induce thirst without causing cellular dehydration. So how does this work? The body constantly monitors the volume of the blood independent of cell hydration (volume). Losing isotonic fluid – both water and solutes – depletes extracellular fluids and causes hypovolemia. Blood loss and sweating both cause hypovolemia. Behavioral Neuroendocrinology - 401 Thirst - 14 There are pressure receptors in the atria of the heart. Information from these baroreceptors is relayed to the hindbrain via the vagus nerves. The vagus nerves terminate in the nucleus of the solitary tract (NTS), which in turn, projects to the median preoptic nucleus. The median preoptic nucleus then stimulates ADH release and thirst, just like cellular dehydration. lateral hypothalamus thirst posterior pituitary vagus nerves NTS median preoptic nucleus supraoptic & paraventricular nuclei ADH kidney But if you’re hypovolemic from the loss of salt and water, just drinking water isn’t enough. Two-thirds of the water in the body is inside cells, so that’s where two-thirds of the water you drink will end up. Just drinking water will cause the cells to swell. (Remember they were not dehydrated (shrunken) before the volemic challenge.) Thus, drinking just water when you’ve lost both water and salt can do more harm than good. (As in the Boston Marathon.) 402 - Behavioral Neuroendocrinology Thirst - 15 In addition to inducing thirst, hypovolemia induces a hunger for sodium (salt). Reduced blood flow to the kidneys activates renal baroreceptors (pressure detectors), and the kidneys respond by secreting renin. Renin is an enzyme that is active in the bloodstream. Renin catalyzes the conversion of the circulating protein, angiotensinogen, to angiotensin I. Angiotensin I is then converted to angiotensin II. Angiotensin II acts as a hormone to affect multiple aspects of salt and water balance. I ACE inhibitors Angiotensin II Behavioral Neuroendocrinology - 403 Thirst - 16 Angiotensin II acts directly on the brain – the subfornical organ – to stimulate thirst and ADH secretion (H2O retention). lateral hypothalamus angiotensin II SFO median preoptic nucleus supraoptic & paraventricular nuclei thirst posterior pituitary ADH kidney Angiotensin II also acts on the adrenal cortex to cause the secretion of the mineralocorticoid, aldosterone. Aldosterone, is a steroid hormone which: Acts in the brain to induce a sodium hunger Acts on the kidney to increase sodium retention brain angiotensin II adrenal cortex aldosterone kidney sodium hunger sodium retention 404 - Behavioral Neuroendocrinology Thirst - 17 Renal hypertension. Reduced blood flow to the kidneys triggers renin release, which then stimulates thirst, sodium hunger, and water and sodium retention. This raises blood pressure. I Angiotensin II Practical advice: 1. If you’re thirsty because the air is dry or because you drool a lot: Drink water, because you’re experiencing cellular dehydration (osmotic thirst). 2. If you’re thirsty because you sweat profusely or because you’re hemorrhaging: Drink saline or Gatorade, because you’re experiencing extracellular dehydration (volemic thirst). Behavioral Neuroendocrinology - 405 Thirst - 18 Yeah, but what I really want to know is why partying just makes me thirstier. That’s easy – you don’t buy beer, you just rent it. Ethanol acts directly on the supraoptic and paraventricular nuclei of the hypothalamus to inhibit ADH secretion. The resulting diuresis dehydrates the body and induces osmotic thirst. 406 - Behavioral Neuroendocrinology Thirst - 19 Behavioral Neuroendocrinology - 407 Thirst - 20 408 - Behavioral Neuroendocrinology ACRONYMS OFTEN USED IN THE COURSE 2DG: 2-deoxy-D-glucose 5TG: 5-thio-glucose ACTH: adrenocotropic hormone ADH: antidiuretic hormone AGRP: agouti-related peptide α-MSH: alpha-melanocyte-stimulating hormone AP: area postrema ARC: arcuate nucleus of the hypothalamus ATP: adenosine triphosphate AVP: argenine vasopressin CAH: compensative adrenal hypertrophy CRH: corticotropin-releasing hormone DHT (5α-DHT): dihydrotestosterone DNA: deoxyribonucleic acid DSAP: saporin conjugated to dopamine beta hydroxlyase E: estradiol (or epinephrine) E2: estradiol EB: estradiol benzoate ER (ERα): estrogen receptor FSH: follicle-stimulating hormone GDX: gonadectomized GH: growth hormone GnRH: gonadotropin-stimulating hormone ICC (IHC): immunocytochemistry ICV: intracerebroventricular IR: immunoreactivity ISSH: in sity hybridization histochemistry (in situ) IV: intravenous KO: knockout L:D: light:dark LH: luteinizing hormone (or lateral hypothalamus) MeA: medial amygdala MP: methyl palmoxirate mRNA: messenger ribonucleic acid NE: norepinephrine NPY: neuropeptide Y NTS: nucleus of the solitary tract OB: olfactory bulb OT: oxytocin OVX: ovariectomized P: progesterone PIF: prolactin-inhibitory factor POA (mPOA): preoptic area PRL: prolactin PVN: paraventricular nucleus of the hypothalamus RNA: ribonucleic acid SCN: suprachiasmatic nucleus TP: testosterone propionate TRH: thyrotropin-releasing hormone TSH: thyroid-stimulating hormone VMH: ventromedial hypothalamus Behavioral Neuroendocrinology - 409 410 - Behavioral Neuroendocrinology ...
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