THE MEMBRANE-POTENTIAL OF VESTIBULAR DARK CELLS IS CONTROLLED BY A LARGE CL- CONDUCTANCE

The K+ secretory epithelium of the vestibular labyrinth (dark cells) was impaled with glass microelectrodes in order to test the hypothesis that it contains a large Cl- conductance. In the first series of experiments, the short-circuited epithelium was perfused on both sides by a solution containing 150 mmol/1 Cl-. The membrane voltage (PD) was -18 +/- 1 mV (N = 101), showed a Gaussian distribution, and the estimated input resistance of the cell (R('cell')) was 17 +/- 3 MOMEGA. The PD responded to 10(-4) mol/l ouabain with a depolarization, suggesting the presence of a (Na+ + K+)-ATPase. The PD responses to Cl- steps yielded an apparent transference number t(Cl) = 0.34 +/- 0.03 (N = 65) and those to K+ steps yielded a t(K) = 0.16 +/- 0.01 (N = 48). In the second series of experiments, cells presumed to be Cl--depleted were impaled in Cl--free solutions. The distribution of the PD was not Gaussian; PDs as negative as -90 mV were observed. Cells with a highly negative PD also had a high R('cell'). With the addition of Cl- the PD collapsed to -19 +/- 1 mV and R collapsed to 16 +/- 3 MOMEGA (N = 145) which are not significantly different from values obtained in the first series of experiments when cells were impaled in a solution containing 150 mmol/l Cl-. Alternating the bath perfusate between Cl--free and Cl--containing solutions led to large PD transients. Cl--induced PD transients could be mimicked by other anions suggesting an ion selectivity of Cl- > Br- > NO3- > I - much greater than glutamate- = isethionate- = gluconate-. Cl--induced PD transients could not be abolished by a blocker of the Na+/Cl-/K+ cotransporter, 10(-4) mol/l piretanide (N = 5); by a blocker of various anion transport systems, 10(-3) Mol/l 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid (SITS; N = 2); or by several blockers of epithelial Cl-channels, 10(-4) mol/l 5-nitro-2(3-phenylpropylamino)-benzoic acid (NPPB; N = 2), and 10(-4) Mol/l dichloro-di-phenyl-amine-2-carboxylic acid (DCDPC; N = 2). In a solution containing 15 mmol/l Cl- the apparent transference numbers were t(CL) = 0.56 +/- 0.03 (N = 19) and t(K) = 0.44 +/- 0.02 (N = 66). The PD responses to K+ steps were not abolished by the K+ channel blockers barium, quinidine or TEA suggesting that the observed PD deflections were only partly due to a K+ conductance and that the apparent t(K) is overestimating the contribution of the K+ conductance. The response to piretanide was variable. However, in a solution containing 15 mmol/l Cl- hyperpolarizations up to -80 mV were observed. These results demonstrate that vestibular dark cells contain a dominating Cl- conductance and suggest that this Cl- conductance is a manifestation of a Cl- channel.