We investìgated the possibility that, ìn hair cells mechanically isolated from frog semicircular canals, Ca2+ extrusìon occurs via a Na+:Ca2+ (cardìac type) or a Na+:Ca2+,K+ (retinal type) exchanger. Cells concurrently imaged during whole-cell patch-clamp recordings using the Ca2+ sensitive fluorescent dye Oregon Green 488 BAPTA-1 (100 microM) showed no voltage dependence of Ca2+ clearance dynamics following a Ca2+ load through voltage-gated Ca2+ channels. Reverse exchange was probed in hair cells dìalysed with a Ca2+- and K+-free solution, containing a Na+ concentration that saturates the exchanger, after zeroing the contribution to the whole-cell current from Ca2+ and K+ conductances. In these conditions, no reverse exchange current was detected upon switching from a Ca2+-free external solution to a solution containing concentrations of Ca2+ alone, or Ca2+K+ that saturates the exchanger. By contrast, the same experimental protocol elicited peak exchange currents exceeding 100 pA in gecko rod photoreceptors, used as positive controls. In both cell types, we also probed the forward mode of the exchanger by rapidly increasìng the intracellular Ca2+ concentration using flash photolysis of two novel caged Ca2+ complexes, Calcium 2,2- [1— (2 — Nitrophenyl) ethane-1,2-diyl]bis(oxy) bis(acetate)] and Calcium 2,2-[1 — (4,5 — Dimethoxy — 2 — Nitrophenyl) ethane — 1,2 — diyl] bis(oxy) bìs(acetate)], in the presence of internal K+ and external Na+. No currents were evoked by UV-triggered Ca2+ jumps in hair cells, whereas exchanger conformational currents up to 400 pA, followed by saturating forward exchange currents up to 40 pA, were recorded in rod photoreceptors subjected to the same experimental conditions. We conclude that no functional electrogenic exchanger ìs present in this hair cell population, which leaves the abundant plasma membrane Ca2+-ATP-ases as the primary contributors to Ca2+ extrusion.
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