Properties and regulation of chloride channels in cystic fibrosis and normal airway cells

Abstract

The present study examines the properties of Cl⁻channels in cultured respiratory cells of cystic fibrosis (CF) patients and normal (N) individuals. In excised membrane patches the conductances for CF and N Cl⁻ channels were larger at positive as compared to negative clamp voltages (V_c): 74±2.6 (V_c > 0) and 47±2.0 pS (V_c < 0) for CF (n= 57) and 69±3.6 (V_c > 0) and 45±2.3 pS (V_c < 0) for N (n=35). The open probability (P_o) of the channel increased markedly with depolarization. Both the voltage dependence of the conductance and of P_o contribute to the outward rectification of the channel. The time histogram analysis reveals two open and two closed time constants. The selectivity of the channel was Cl⁻=Br⁻ =I⁻ > NO ₃ ⁻ ≫ gluconate. The channel was inhibited reversibly by 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB) at 10⁻⁷ mol/l to 10⁻⁵ mol/l. While Cl⁻ channels were present in cell attached patches of N cells, they were absent in those of CF cells. The mean conductance for cell attached (N) Cl⁻ channels was 76±3.2 pS for positive clamp voltages (V_c) and 46±3.9 pS for negative V_c (n=8). When the membrane patches were excised from CF cells Cl⁻ currents appeared spontaneously (n=19). The immediate appearance (within 1 s) of Cl⁻ channels after excision was observed at positive (n=6) as well as at negative clamp voltage (n=13). “Excision activation” of CF Cl⁻ channels was observed at low (< 10⁻⁹ mol/l) or high (10⁻³ mol/l) calcium activities on the cytosolic side of the excised patch. Variation of the Ca⁺ activity (< 10⁻⁹–10⁻³ mol/l) or pH (6.5–8.5) on the cytosolic side exerted no effects on these Cl⁻ channels. These results suggest that Cl⁻ channels are present in the apical membrane of CF and N respiratory cells but they seem to be inhibited in intact CF cells. Excision of the patch and hence removal of the cytosolic “inhibitor” leads to an activation of Cl⁻ channels. The Cl⁻ channels in excised patches of N and CF cells have identical properties.