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Unformatted text preview: Bio sat 25.10.2008 Patch Clamps Permit Measurement of Ion Movements Through Single Channels The technique of patch clamping هعقرلا تيبث¡ enables workers to investigate the opening, closing, regulation, and ion conductance of a single ion channel. ► In this technique, the inward or outward movement of ions across a patch of membrane is quantified from the amount of electric current needed to maintain the membrane potential at a particular “clamped” value (Figure 7-17a, b) . To preserve electroneutrality and to keep the membrane potential constant, the entry of each positive ion (e.g., a Na + ion ) into the cell through a channel in the patch of membrane is balanced by the addition of an electron into the cytosol through a microelectrode inserted into the cytosol; an electronic device measures the numbers of electrons (current) required to counterbalance the inflow of ions through the membrane channels. Conversely, the exit of each positive ion from the cell (e.g. , a K + ion ) is balanced by the withdrawal of an electron from the cytosol. The patch-clamping technique can be employed on whole cells or isolated membrane patches to measure the effects of different substances and ion concentrations on ion flow (Figure 7-17c) .-------------------------- EXPERIMENTAL FIGURE 7-17 Current flow through individual ion channels can be measured by patch- clamping technique . (a) Basic experimental arrangement for measuring current flow through individual ion channels in the plasma membrane of a living cell. - The patch electrode , filled with a current-conducting saline solution, is applied, with a slight suction, to the plasma membrane. 1- The 0.5-m-diameter tip covers a region that contains only one or a few ion channels. - The second electrode is inserted through the membrane into the cytosol. A recording device measures current flow only through the channels in the patch of plasma membrane. (b) Photomicrograph of the cell body of a cultured neuron and the tip of a patch pipette touching the cell membrane. He didn’t illustrated in his lec (c) Different patch-clamping configurations. Isolated, detached patches are the best configurations for studying the effects on channels of different ion concentrations and solutes such as extracellular hormones and intracellular second messengers (e.g., cAMP). ------------------------------------- The patch-clamp tracings in Figure 7-18 illustrate the use of this technique to study the properties of voltage-gated Na + channels in the plasma membrane of muscle cells. As we discuss later,- these channels normally are closed in resting muscle cells and - open following nervous stimulation. - Patches of muscle membrane, each containing one Na + channel, were clamped at a voltage slightly less than the resting membrane potential. Under these circumstances, transient pulses of current cross the membrane as individual Na + channels open and then close....
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- Spring '10
- Cellular Respiration, Adenosine triphosphate