FUNCTIONAL IDENTIFICATION OF A SARCOLEMMAL CHLORIDE CHANNEL FROM BOVINE TRACHEAL SMOOTH-MUSCLE

The biophysical and pharmacological characteristics of unitary Cl- cur rents from bovine tracheal smooth muscle cells were studied after reco nstitution of microsomal vesicles into planar lipid bilayers. Two type s of currents were recorded simultaneously in KCl buffer: the well-def ined Ca2+-dependent K+ conductance [G(K(Ca))] and a much smaller Cl- c urrent, indicating that the Cl- channels under scrutiny originate from the same membrane as the G(K(Ca))-type channels, the plasma membrane of airway smooth muscle (ASM) cells. The G(K(Ca)) activities were elim inated 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 cocktai ls, including protein kinases A, G, or C, did not alter the activity o f the channel nor did changes in pH. Among a series of Cl- channel inh ibitors, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid [50% effecti ve concentration (EC(50)) = 30 mu M] and 5-nitro-2-(3-phenylpropylamin o) benzoic acid (EC(50) = 130 mu M) were the most potent blockers of t he current examined. The exact role of this surface Cl- conductance re mains unclear, and its involvement in cellular activity needs further investigation.