Chloride efflux inhibits single calcium channel open probability in vertebrate photoreceptors: Chloride imaging and cell-attached patch-clamp recordings

Abstract

The present study uses cell-attached patch-recording techniques to study the single-channel properties of Ca²⁺ channels in isolated salamander photoreceptors and investigate their sensitivity to reductions in intracellular Cl⁻. The results show that photoreceptor Ca²⁺ channels possess properties similar to L-type Ca²⁺ channels in other preparations, including (1) enhancement of openings by the dihydropyridine agonist, (−)BayK8644; (2) suppression by a dihydropyridine antagonist, nisoldipine; (3) single-channel conductance of 22 pS with 82 mM Ba²⁺ as the charge carrier; (4) mean open probability of 0.1; (5) open-time distribution fit with a single exponential (τ₀ = 1.1 ms) consistent with a single open state; and (6) closed time distribution fit with two exponentials (τ_c1 = 0.7 ms, τ_c2 = 25.4 ms) consistent with at least two closed states. Using a Cl⁻-sensitive dye to measure intracellular [Cl⁻], it was found that perfusion with gluconate-containing, low Cl⁻ medium depleted intracellular [Cl⁻]. It was therefore possible to reduce intracellular [Cl⁻] by perfusion with a low Cl⁻ solution while maintaining the extracellular channel surface in high Cl⁻ pipette solution. Under these conditions, the single-channel conductance was unchanged, but the mean open probability fell to 0.03. This reduction can account for the 66% reduction in whole-cell Ca²⁺ currents produced by perfusion with low Cl⁻ solutions. Examination of the open and closed time distributions suggests that the reduction in open probability arises from increases in closed-state dwell times. Changes in intracellular [Cl⁻] may thus modulate photoreceptor Ca²⁺ channels.