A patch-clamp study of potassium channels and whole-cell currents in acinar cells of the mouse lacrimal gland.

摘要

Individual acinar cells were isolated enzymatically from the mouse exorbital lacrimal gland. Their electrical characteristics were studied by the patch-clamp methods of single-channel and whole-cell recording as described by Hamill, Marty, Neher, Sakmann & Sigworth (1981). Recording from cell-attached and excised inside-out patches of acinar membrane with quasi-physiological ion gradients demonstrated large outward current events that correspond to single-channel openings. The amplitude, frequency and duration of channel events increased as the membrane patch was depolarized and were reduced by hyperpolarization of the patch membrane. The reversal potential for these channel events is more negative than -40 mV. In excised inside-out patches exposed to quasi-physiological ion gradients single-channel events were abolished when K+ was replaced by Rb+. Since there was no Cl- gradient the channel is clearly highly selective for K+. In excised inside-out patches, when the free Ca2+ concentration bathing the physiological inside of the membrane was raised from less than 10(-9) M to 10(-8) M the frequency and duration of opening of the K+ channel was increased. The channel was almost continuously open when the membrane was exposed to 10(-7) M-free Ca2+. 'Whole cell' recording of lacrimal acinar cells containing 140 mM-KCl and 1 mM-EGTA (with no added Ca2+) provided cell resting membrane potentials of -30 to -40 mV. Depolarizing voltage jumps from the resting membrane potential evoked large outward currents. Hyperpolarizing voltage jumps only evoked small inward currents. Whole-cell recording where RbCl replaced KCl in the pipette provided resting membrane potentials of -20 to -30 mV, reduced the amplitude of outward currents evoked by cell-depolarizing voltage steps by 60% and slowed the time course of the currents. Isolated cells containing 140 mM-KCl and 1 mM-EGTA were voltage clamped at their resting membrane potentials. Acetylcholine (ACh) was applied locally and immediately evoked a strong outward current which rapidly declined to a steady-state level. Sustained agonist responses were obtained by exposing the isolated cell to a solution containing 10(-6) M-ACh. In both K+- and Rb+-filled cells, where the intracellular Ca2+ concentration was buffered by the inclusion of 1 mM-EGTA, 10(-6) M-ACh evoked sustained outward currents that corresponded to cell hyperpolarizations of 5-15 and 10-20 mV, respectively. Increasing intracellular Ca2+ buffering by including 10 mM-EGTA abolished secretagogue-induced outward current in both K+- and Rb+-filled cells. It is concluded that the lacrimal acinar cell membrane contains voltage- and Ca2+-activated K+ channels.(ABSTRACT TRUNCATED AT 400 WORDS)