Functional identification of a sarcolemmal chloride channel from bovine tracheal smooth muscle.
The biophysical and pharmacological characteristics of unitary Cl- currents from bovine tracheal smooth muscle cells were studied after reconstitution of microsomal vesicles into planar lipid bilayers. Two types of currents were recorded simultaneously in KCl buffer: the well-defined Ca(2+)-dependent K+ conductance [GK(Ca)] and a much smaller Cl- current, indicating that the Cl- channels under scrutiny originate from the same membrane as the GK(Ca)-type channels, the plasma membrane of airway smooth muscle (ASM) cells. The GK(Ca) activities were eliminated by the use of CsCl buffer. The average unitary Cl- conductance measured in 50 mM trans-250 mM cis CsCl was 77 +/- 6 pS (n = 21), and the reversal potential measured in various CsCl gradients followed the Cl- equilibrium potential as determined from the Nernst equation. In contrast with the previous reports describing the Ca2+ sensitivity of macroscopic ASM Cl- currents, this channel was found to be insensitive to cytoplasmic and extracellular Ca2+ levels. Phosphorylation cocktails, including protein kinases A, G, or C, did not alter the activity of the channel nor did changes in pH. Among a series of Cl- channel inhibitors, 4,4'-diisothiocyanostilbene-2, 2'-disulfonic acid [50% effective concentration (EC50) = 30 microM] and 5-nitro-2-(3-phenylpropylamino) benzoic acid (EC50 = 130 microM) were the most potent blockers of the current examined. The exact role of this surface Cl- conductance remains unclear, and its involvement in cellular activity needs further investigation.
Salvail
D
D
Department of Physiology and Biophysics, Faculty of Medicine, University of Sherbrooke, Quebec, Canada.
Alioua
A
A
Rousseau
E
E
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D002417:Cattle; D002458:Cell Fractionation; D002462:Cell Membrane / Q000502:physiology; D002586:Cesium / Q000494:pharmacology; D018118:Chloride Channels / Q000187:drug effects / Q000502:physiology*; D002712:Chlorides / Q000378:metabolism / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D004533:Egtazic Acid / Q000494:pharmacology; D008051:Lipid Bilayers; D008564:Membrane Potentials / Q000187:drug effects; D009130:Muscle, Smooth / Q000502:physiology*; D009579:Nitrobenzoates / Q000494:pharmacology; D015221:Potassium Channels / Q000187:drug effects / Q000502:physiology; D012508:Sarcolemma / Q000502:physiology*; D013629:Tamoxifen / Q000494:pharmacology; D014132:Trachea / Q000502:physiology*
D018118:Chloride Channels; D002712:Chlorides; D008051:Lipid Bilayers; D009579:Nitrobenzoates; D015221:Potassium Channels; D013629:Tamoxifen; D002586:Cesium; C058176:5-nitro-2-(3-phenylpropylamino)benzoic acid; D004533:Egtazic Acid; C028019:cesium chloride; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D002118:Calcium