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Characterization of calcium-activated potassium channels in single smooth muscle cells using the patch-clamp technique

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Abstract

Single-channel currents were recorded with the patch-clamp technique from freshly dissociated vertebrate smooth muscle cells from the stomach ofBufo marinus. Of the variety of channels observed, one displayed a large linear conductance of 250 pS (in symmetric 130 mM KCl) which in excised patches was shown to be highly K+ selective. The probability of the channel being open (P o) increased when [Ca2+]i was elevated and/or when the membrane potential was made more positive. Thus, the features of this channel resemble the large-conductance Ca2+-activated K+ channel found in a wide variety of cell types. The voltage sensitivity of the channel was studied in detail. For patches containing a single large-conductance channel a plot ofP o versus membrane potential followed the Boltzman relationship. Increasing [Ca2+]i shifted this plot to the left along the voltage axis to more negative potentials. Both the mean closed time and mean open time varied with potential as a single exponential with almost all of the voltage sensitivity ofP o residing in the mean closed time. These results were verified with a series of experiments carried out at lowP o (<0.1) in patches containing multiple (N) large-conductance channels. Here the ln (NP o) was a linear function of potential with an inverse slope of 9 mV. Almost all of the potential sensitivity lay in the mean closed time the natural log of which was also a linear function of potential with an inverse slope 11 mV in magnitude. The characteristics of this channel as well as the appearance of several of them in almost every patch suggest that they underlie the large peak outward macroscopic current found with whole-cell voltage-clamp studies.

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References

  • Almers W (1978) Gating currents and charge movements in excitable membranes. Rev Physiol Biochem Pharmacol 82:96–190

    Google Scholar 

  • Bagby RM, Young AM, Dotson RS, Fisher BA, McKinnon K (1971) Contraction of single smooth muscle cells fromBufo marinus stomach. Nature 234:351–352

    Google Scholar 

  • Barrett JN, Magleby KL, Pallotta BS (1982) Properties of single calcium-activated potassium channels in cultured rat muscle. J Physiol 331:211–230

    Google Scholar 

  • Benham CD, Bolton TB, Lang RJ, Takewaki T (1984) Calcium-dependent K+ channels in dispersed intestinal and arterial smooth cells of guinea pigs and rabbits studies by the patchclamp technique. J Pysiol 350:51P

    Google Scholar 

  • Benham CD, Bolton TB, Lang RJ, Takewaki T (1985) The mechanism of action of Ba2+ and TEA on single Ca2+-activated K+-channels in arterial and intestial smooth muscle cell membranes. Pflügers Arch 403:120–127

    Google Scholar 

  • Benham CD, Bolton TB, Lang RJ, Takewaki T (1986) Calcium-activated potassium channels in single smooth muscle cells of rabbit jejunum and guinea-pig mesenteric artery. J Physiol 371:45–67

    Google Scholar 

  • Berger W, Grygorcyk R, Schwarz W (1984) Single K+ channels in membrane evaginations of smooth muscle cells. Pflügers Arch 402:18–23

    Google Scholar 

  • Cecchi X, Wolff D, Alvarez O, Latorre R (1984) Incorporation of Ca2+-activated K+ channels from rabbit intestinal smooth muscle sarcolemma into planar bilayers. Biophys J 45:38a

    Google Scholar 

  • Chamley-Campbell J, Campbell GR, Ross R (1979) The smooth muscle cell in culture. Physiol Rev 59:1–61

    Google Scholar 

  • Colquhoun D, Hawkes AG (1977) Relaxation and fluctuations of membrane currents that flow through drug-operated channels. Proc R Soc Lond B 199:231–262

    Google Scholar 

  • Colquhoun D, Neher E, Reuter H, Stevens CF (1981) Inward current channels activated by intracellular Ca in cultured cardiac cells. Nature 294:752–754

    Google Scholar 

  • Fay FS, Hoffmann R, Leclair S, Merriam P (1982) Preparation of individual smooth muscle cells from the stomach ofBufo marinus. Methods Enzymol 85:284–292

    Google Scholar 

  • Feller W (1968) An introduction probability theory and its applications, vol I. John Wiley and Sons, New York

    Google Scholar 

  • Fenwick EM, Marty A, Neher E (1982) Sodium and calcium channels in bovine chromaffin cells. J Physiol 331:599–635

    Google Scholar 

  • Hamill OP (1983) Potassium and chloride channels in red blood cells. In: Sakmann B, Neher E (eds) Single-channel recording. Plenum Press, New York, pp 451–471

    Google Scholar 

  • Hamill OP, Marty A, Neher E, Sakmann B, Sigworth FJ (1981) Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflügers Arch 391:85–100

    Google Scholar 

  • Hille B (1984) Ionic channels in excitable membranes. Sinauer Associates, Sunderland, MA

    Google Scholar 

  • Horn R (1984) Gating of channels in nerve and muscle: a stochastic approach. In: Stein WD (ed) Ion channels: Molecular and physiological aspects. Academic Press, New York, pp 53–97

    Google Scholar 

  • Inoue R, Kitamura K, Kuriyama H (1985) Two Ca-dependent K-channels classified by the application of tetraethylammonium distribute to smooth muscle membranes of the rabbit portal vein. Pflügers Arch 405:173–179

    Google Scholar 

  • Lassignal NL, Singer JJ, Walsh JV jr (1986) Multiple neuropeptides exert a direct effect on the same isolated single smooth muscle cell. Am J Physiol 250 (Cell Physiol 19):C792-C798

    Google Scholar 

  • Latorre R, Coronado R, Vergara C (1984) K+ channels gated by voltage and ions. Ann Rev Physiol 46:485–495

    Google Scholar 

  • Latorre R, Vergara C, Hidalgo C (1982) Reconstitution in planar lipid bilayers of a Ca2+-dependent K+ channel from transverse tubule membranes isolated from rabbit skeletal muscle. Proc Natl Acad Sci USA 79:805–809

    Google Scholar 

  • Lus HD, Neher E, Marty A (1981) Single channel activity associated with the calcium dependent outward current inHelix pomatia. Pflügers Arch 389:293–295

    Google Scholar 

  • Magleby KL, Pallotta BS (1983a) Calcium dependence of open and shut interval distributions from calcium-activated potassium channels in cultured rat muscle. J Physiol 344:585–604

    Google Scholar 

  • Magleby KL, Pallotta BS (1983b) Burst kinetics of single calcium-activated potassium channels in cultured rat muscle. J Physiol 344:605–623

    Google Scholar 

  • Marty A (1981) Ca-dependent K channels with large unitary conductance in chromaffin cell membranes. Nature 291:497–500

    Google Scholar 

  • Marty A (1983a) Blocking of large unitary calcium-dependent potassium currents by internal sodium ions. Pflügers Arch 396:179–181

    Google Scholar 

  • Marty A (1983b) Ca2+-dependent K+ channels with large unitary conductance. Trends Neurosc 6:262–265

    Google Scholar 

  • McCann JD, Welsh MJ (1985) Voltage-gated Ca-activated K channels in isolated canine airway smooth muscle cells. Biophys J 47:135a

    Google Scholar 

  • McManus OB, Blatz AL, Magleby KL (1985) Inverse relationship of the durations of adjacent open and shut intervals for Cl and K channels. Nature 317:625–627

    Google Scholar 

  • Meech RW (1978) Calcium-dependent potassium activation in nervous tissues. Ann Rev Biophys Bioeng 7:1–18

    Google Scholar 

  • Methfessel C, Boheim G (1982) The gating of single calcium-dependent potassium channels is described by an activation/blockade mechanism. Biophys Struct Mech 9:35–60

    Google Scholar 

  • Moczydlowski E, Latorre R (1983) Gating kinetics of Ca2+-activated K+ channels from rat muscle incorporated into planar lipid bilayers. J Gen Physiol 82:511–542

    Google Scholar 

  • Moczydlowski E, Alvarez O, Vergara C, Latorre R (1985) Effect of phospholipid surface charge on the conductance and gating of a Ca2+-activated K+ channel in planar lipid bilayers. J Membr Biol 83:273–282

    Google Scholar 

  • Neher E (1983) The charge carried by single-channel currents of rat cultured muscle cells in the presence of local anaestetics. J Physiol 339:663–678

    Google Scholar 

  • Pallotta BS (1985a) Calcium-activated potassium channels in rat muscle inactivate from a short-duration open state. J Physiol 363:501–516

    Google Scholar 

  • Pallotta BS (1985b) N-Bromoacetamide removes a calcium-dependent component of channel opening from calcium-activated potassium channels in rat skeletal muscle. J Gen Physiol 86:601–611

    Google Scholar 

  • Petersen OH, Maruyama Y (1984) Calcium-activated potassium channels and their role in secretion. Nature 307:693–696

    Google Scholar 

  • Sakmann B, Neher E (1983) Geometric parameters of pipettes and membrane patches. In: Sakmann B, Neher E (eds) Single-channel recording. Plenum Press, New York, pp 37–51

    Google Scholar 

  • Scheid CR, Fay FS (1980) Control of ion distribution in isolated smooth muscle cells. J Gen Physiol 75:163–182

    Google Scholar 

  • Schwarz W, Passow H (1983) Ca2+-activated K+ channels in erythrocytes and excitable cells. Ann Rev Physiol 45:359–374

    Google Scholar 

  • Singer JJ, Walsh JV Jr (1980a) Passive properties of the membrane of single freshly isolated smooth muscle cells. Am J Physiol 239 (Cell Physiol 8):C153-C161

    Google Scholar 

  • Singer JJ, Walsh JV Jr (1980b) Rectifying properties of the membrane of single freshly isolated smooth muscle cells. Am J Physiol 239 (Cell Physiol 8):C175-C181

    Google Scholar 

  • Singer JJ, Walsh JV Jr (1982) Single-channel currents recorded in freshly dissociated vertebrate smooth muscle cells using the patch-clamp technique. J Gen Physiol 80:23a-24a

    Google Scholar 

  • Singer JJ, Walsh JV Jr (1984) Large conductance Ca2+-activated K+ channels in smooth muscle cell membrane: Reduction in unitary currents due to internal Na+ ions. Biophys J 45:68–70

    Google Scholar 

  • Singer JJ, Walsh JV Jr (1986) Large-conductance Ca2+-activated K+ channels in freshly dissociated smooth muscle cells. Membr Biochem 6:83–110

    Google Scholar 

  • Trautmann A, Marty A (1984) Activation of Ca-dependent K channels by carbamoylcholine in rat lacrimal glands. Proc Natl Acad Sci USA 81:611–615

    Google Scholar 

  • Vivaudou M, Singer JJ, Walsh JV Jr (1986) An automated technique for analysis of current transitions in multilevel single-channel recordings. Pflügers Arch 407:355–364

    Google Scholar 

  • Walsh JV Jr, Singer JJ (1980a) Calcium action potentials in single freshly isolated smooth muscle cells. Am J Physiol 239 (Cell Physiol 8):C162-C174

    Google Scholar 

  • Walsh JV Jr, Singer JJ (1980b) Penetration-induced hyperpolarization as evidence for Ca2+ activation of K+ conductance in isolated smooth muscle cells. Am J Physiol 239 (Cell Physiol 8):C182-C189

    Google Scholar 

  • Walsh JV Jr, Singer JJ (1981) Voltage clamp of single freshly dissociated smooth muscle cells: current-voltage relationships for three currents. Pflügers Arch 390:207–210

    Google Scholar 

  • Walsh JV Jr, Singer JJ (1983a) Identification and characterization of a Ca2+-activated K+ channel in freshly dissociated vertebrate smooth muscle cells using the pathch-clamp technique. Biophys J 41:56a

    Google Scholar 

  • Walsh JV Jr, Singer JJ (1983b) Ca2+-activated K+ channels in vertebrate smooth muscle cells. Cell Calcium 4:321–330

    Google Scholar 

  • Walsh JV Jr, Singer JJ (1987) Identification and characterization of major ionic currents in isolated smooth muscle cells using the voltage-clamp technique. Pflügers Arch 408:83–97

    Google Scholar 

  • Wolff D, Cecchi X, Naranjo D, Alvarez O, Latorre R (1985) Cation selectivity and Cs+ blockade in a Ca2+-activated K+ channel from rabbit intestinal smooth muscle. Biophys J 47:386a

    Google Scholar 

  • Wong BS (1985) Single calcium-dependent potassium channels from amphibian stomach smooth muscle cells. Biophys J 47:136a

    Google Scholar 

  • Wong BS, Lecar H, Adler M (1982) Single calcium-dependent potassium channels in clonal anterior pituitary cells. Biophys J 39:313–317

    Google Scholar 

  • Yellen G (1982) Single Ca2+-activated nonselective cation channels in neuroblastoma. Nature 296:357–359

    Google Scholar 

  • Yellen G (1984a) Ionic permeation and blockade in Ca2+-activated K+ channels of bovine chromaffin cells. J Gen Physiol 84:157–186

    Google Scholar 

  • Yellen G (1984b) Relief of Na+ block of Ca2+-activated K+ channels by external cations. J Gen Physiol 84:187–199

    Google Scholar 

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  1. Department of Physiology, University of Massachusetts Medical School, 01605, Worcester, Massachusetts, USA

    Joshua J. Singer & V. Walsh Jr.

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  1. Joshua J. Singer
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  2. V. Walsh Jr.
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Singer, J.J., Walsh, V. Characterization of calcium-activated potassium channels in single smooth muscle cells using the patch-clamp technique. Pflugers Arch. 408, 98–111 (1987). https://doi.org/10.1007/BF00581337

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  • DOI: https://doi.org/10.1007/BF00581337

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