Cl^- channels in basolateral renal medullary memnbranes: III. Determinants of single-channel activity

We evaluated the effects of vawrying aqueous Cl^- concentrations, and of the arginyl- and lysyl-specific reagent phenylglyoxal (PGO), on the properties of Cl^- channels fused from basolaterally enriched renal medullary vesicles into planar lipid bilayers. The major channel properties studied were the anion selectivity sequence, anionic requirements for, channel activity. and the efects of varying Cl^- concentrations and/or PGO on the relation between holding voltage V_H-mV) and open-time probability (P_o). Reducing cis Cl^- concentrations, in the range 50-320 mm, produced a linear reduction in fractional open time (P_v) with a half-maximal reduction in P_o at cis Cl^-≈170 mM. Channel activity was sustained by equimolar replacement of cis Cl^- with F^-, but not with impermeant isethionate. For trans solutions, the relation between Cl^- concentration and P₀ at 10m m Cl^-. Reducing cis Cl^- had no effect on the gating charge (Z) for channel opening, but altered significantly the voltage-independent, energy (δG) for channel opening. Phenylglyoxal (PGO) reduced Z and altered δG for Cl^- channel activity when added to cis, but not trans solutions, Furthermore, in the presence of cis PGO, reducing the cis Cl^- concentration had no effect on Z but altered δG. Thus we propose that cis PGO and, cis Cl^- concentrations affect separate sites determining channel activity at the extracellular faces of, these Cl^- channels.