MCB 32 F08 Chapter 6, Membrane transport of solutes, water and ions by diffusion, mediated transport, active transport, secondary active transport, filtration and osmosis. Generation of voltages. Outline Diffusion in solution Permeation across a membrane Water transport: filtration and osmosis Carrier mediated transport: <a href="/keyword/facilitated-diffusion/" >facilitated diffusion</a> Exchangers Active transport: primary and secondary <a href="/keyword/ion-channels/" >ion channels</a> and development of membrane voltages Body fluids As noted in previous lecture, intracellular and <a href="/keyword/extracellular-fluid/" >extracellular fluid</a> s have similar total salt concentrations (about 150 mM salt), but the specific compositions are quite different. Thus, <a href="/keyword/extracellular-fluid/" >extracellular fluid</a> is high in [Na +] and low in [K +] and protein while the intracellular fluid is low in [Na +], [Cl ] and 2+ ] compared to the <a href="/keyword/extracellular-fluid/" >extracellular fluid</a> . Intracellular fluid also contains about 1 mM ATP, which is [Ca found outside cells in much lower amounts. Membrane transport of ions and water The plasma membrane with its lipids and proteins determines what goes into and out of the cell. The movements occur by either diffusion through the lipids or mediated transport through the various specific proteins that are embedded in the membranes. Diffusion: property associated with any dissolved molecule, which has its own kinetic energy and vibrates randomly in solution, leading to the random movement throughout the solution. Diffusion of solutes occurs to equalize the concentration throughout the solution, from high to low concentration. Example: drop of dye at the bottom of a beaker. The rate of diffusional flux from one region to another in solution is directly proportional to the difference in concentration between two regions (C1 and C2) and the area available for exchange (A) and the diffusion coefficient of the solute (D, inversely proportional to the size of the molecule) and inversely proportional to the distance ( x) between the two points. Net diffusional flux = D x (C1 C2) x A x ( x) 1 The rate of diffusional flux of lipid soluble substances like gases across a cell membrane is directly proportional to the difference in concentration between inside and outside the cell (Co and Ci) and the area of the cell membrane available for exchange (A) and the permeability coefficient of the solute (P, directly proportional to the lipid solubility of the molecule). Overall, most important determinants of flux into cells is permeability of the cell membrane to the molecule and the area of membrane. flux across a cell membrane = P x (Co Ci) x area available for exchange Filtration and osmosis are processes determining movements of water across membranes. Filtration involves there being water pressure in one region being higher than in another region, and water flows from high to low pressure. Examples: hose with high pressure at one end, low pressure at the other. U tube, push on the water, displace the fluid until it reaches a certain height. Called <a href="/keyword/hydrostatic-pressure/" >hydrostatic pressure</a> in which the height of the column of fluid equals the pressure forcing the fluid in one tube down. Pushing pressure. pressure inside the cardiovascular system, blood pressure, that is important for moving the blood around the circulatory system, but the capillaries are very thin and permeable to water. Thus, water is forced out across the capillaries by this pushing pressure. Called filtration. Osmosis is a pulling force or pulling force that is determined by the number of dissolved molecules in a solution compared to another solution, with the two solutions separated by a membrane that is impermeant to the solute molecule. Example: U tube with a membrane. Put in some solute on one side, the water will move across the membrane, pulled by the so called osmotic pulling pressure. Height of water will be determined by the number of dissolved, impermeant molecules. Thus, osmotic pressure is determined by the number of dissolved molecules in solution. When equal number of impermeant solutes on each side, no water flow, i.e., no osmosis. Example: proteins dissolved in the blood, much less in the tissue spaces. This difference in concentration of proteins leads to an osmotic pulling pressure that offsets the filtration pressure such that there is balance between the filtration force outward and the osmotic force inward. Carrier mediated transport refers to the movement of water soluble (mostly) molecules across the cell membrane. Important substances that need to get into and out of cells must go through the membrane in association with proteins in the membrane. <a href="/keyword/facilitated-diffusion/" >facilitated diffusion</a> refers to the movement of substances like glucose that traverse the membrane from high to low concentration driven solely by this difference in concentration, but do so associated with a membrane transport protein. In the case of glucose the carrier is called Glut short for glucose transporter. Characteristics of <a href="/keyword/facilitated-diffusion/" >facilitated diffusion</a> : Specific for only glucose and similar looking molecules because the Glut protein has a particular shape, accepting only sugars with 6 C. Exhibit competition in which movement of glucose can be reduced by adding another, similar molecule like galactose, also with 6 C. They also can be blocked by inhibitory drugs that bind to the site at which glucose binds. The transporters work only so fast, and this occurs when the transport sites all contain glucose (at high concentration of glucose) and the transporters are therefore saturated. Active transport (also called primary active transport) is similar to <a href="/keyword/facilitated-diffusion/" >facilitated diffusion</a> in that proteins in the membrane mediate the movement of the molecules, but this also involves input of energy in the form of ATP, such that there can be an accumulation of substances inside the cell to concentrations higher than outside the cell. Specific, competition, inhibitors, saturation Also, break down ATP, couple the hydrolysis of ATP to the movement of substances. Example: Na/K pump or ATPase (hydrolysis of ATP) Secondary active transport occurs when the diffusive movement of one substance is coupled to the active, energy requiring movement of a second substance. This type of transport can lead to the energy requiring accumulation of substances by cells. This involves the so called down hill, diffusive movement of one molecule, for example Na, coupled to the so called uphill, energy requiring movement of another molecule, for example glucose, into the cell. This is called secondary active transport because it does not directly involve the input of energy in the form of ATP, but secondarily there is an ATP requirement because the concentration gradient for Na must be maintained through the input of ATP energy (primary active transport). Example: uptake of glucose out of the intestinal lumen by the small intestinal cells. Requires both Na and glucose, both of which enter the cell together. Called Na glucose cotransport. This transporter is used in the treatment of cholera: secondary active accumulation of Na and glucose at the apical membrane, primary active transport of Na in exchange for K at the basal membrane, and <a href="/keyword/facilitated-diffusion/" >facilitated diffusion</a> of glucose into the blood. Overall, there is accumulation of both Na and glucose, and because these form osmotically active molecules, there is water that follows. This prevents dehydration associated with cholera. Diffusive flux of ions through <a href="/keyword/ion-channels/" >ion channels</a> is similar to <a href="/keyword/facilitated-diffusion/" >facilitated diffusion</a> of non electrolytes because the ions associate with a protein that has specificity and can exhibit competition and inhibition and saturation, but there is a big difference in that the ions have charge associated with them. The presence of charge on the ion means that the movement of the ion can alter the charge or voltage across the membrane and also that the ion s movement is affected by voltage across the membrane. Voltage: separation of small amount of charge across the very thin cell membrane. Measured in volts or millivolts (10 3 volts). In biology, due to the presence of different ion concentrations across a membrane and the presence of <a href="/keyword/ion-channels/" >ion channels</a> that are selectively permeable to one or another ion. The ion <a href="/keyword/concentration-gradients/" >concentration gradients</a> that we are most concerned with are those for Na and K. These concentration differences for Na and K are in turn established by the activity of the Na/K ATPase or pumps present in every cell of the body. Example: K concentration difference across a membrane causes diffusive flux of K, and if no other ion is permeant, this leads to separation of small amount of charge across the membrane, with side with low concentration becoming the same sign voltage as the ion that is permeating. Magnitude of the voltage in this non physiological, but useful, situation can be calculated using the Nernst equation (which is useful for making predictions): Vin out = {RT/zF} x {ln([Kout]/[Kin])} Vin out is the voltage across the membrane, inside with respect to outside, and R is the gas constant, T is the absolute temp, z is the valence on the ion and F is the Faraday constant. For all practical purposes, this equation reduces to; Vin out = 60 log [Kout]/[Kin] Vin out (measured in mV) = 60 mV x log [5 mM]/[120 mM] Because the ratio of the concentrations is smaller than one, the log of this number will be negative, so the inside is negative with respect to the outside, approximately 80 mV. This is the voltage expected for a cell in which the ratio of the K concentrations is as given and the membrane was permeable only to K. If the cell were permeable to other ions, then the voltage would be expected to be smaller, more depolarized (less polar) on the inside with respect to the outside. Example: what if the concentration of K were the same inside and outside the cell? Membrane voltage would be zero (log 1 = 0). Example: what if the membrane were permeable only to Na instead of to K and the concentration gradient for Na across the membrane were as shown at the beginning of the lecture? Inside would be positive because [Naout]/[Nain] &gt; 1, and log of anything larger than 1 will be a positive number. Overall, the Nernst equation is useful for predicting the magnitude and orientation of a membrane s voltage, if you know the most permeant ion and its concentrations inside and outside a cell. Membrane voltages are generated by: Selective <a href="/keyword/ion-channels/" >ion channels</a> in the membrane Concentration gradient across the membrane Study Guide: Distinguish between: diffusion and osmosis; filtration and osmosis; primary and secondary active transport; cotransporter and exchanger; electronegativity and electroneutrality 1. T F? Cell voltage is negative inside because there are more cations (positively charged ions) outside the cell than inside. 2. T F? Carrier proteins have multiple similarities to enzymes. 3. A poison like cyanide that blocks ATP production would prevent the action of which of the following: Na K pump, glucose transporter, ion channel, Na glucose uptake mechanism in the small intestine? 4. In the following condition, which direction does water flow, from 1 2 or from 2 1 or no flow? Which direction do Na and Cl flow, from 1 2 or from 2 1 or no flow? Would there be a membrane voltage established? What is its orientation, side 1 or side 2 positive or no voltage? Membrane that is permeant to Na and Cl and water but impermeant to protein separates two fluids. Side 1 contains 100 mM NaCl + 2 mM protein Side 2 contains 150 mM NaCl + 1 mM protein Answer the same questions for the situation in which the membrane is permeant to Na and water but impermeant to Cl and protein. 5. Which ion is the most important in determining the voltage across the membranes of most cells? In hyperkalemia there is increased [K] in the blood and interstitial fluids, but [K] in the cells remains roughly constant. What would you expect to occur in hyperkalemia to the voltage across the membranes of cells that had very high permeability to K and little permeability to other ions? 6. Cholera is caused by a bacterial toxin that causes the cells in the intestine to actively secrete Cl and Na. Explain why this loss of salt leads to loss of fluids and dehydration and why the treatment for cholera involves drinking fluids that contain both salt and glucose. 7. Explain why cooling cells (e.g., putting them in a refrigerator) often causes them to lose K and gain Na, and rewarming the cells causes them to regain K and lose Na. Hint: <a href="/keyword/ion-channels/" >ion channels</a> can often function relatively normally at low temperature but active transporters can stop working at low temperatures. 8. Capillary membranes are permeable to water but not to proteins. Under this condition, the <a href="/keyword/hydrostatic-pressure/" >hydrostatic pressure</a> developed by the heart inside the blood vessels tends to elicit filtration of fluid in which direction? Into the tissues or into the blood? What would happen to the magnitude of this filtration if protein concentration in the blood were increased or decreased? 9. T F? Gases like O2 and CO2 are charged and lipid insoluble and therefore require specific transporters to move between the blood and the insides of cells.