DIRECT MEASUREMENTS OF CA2+-ACTIVATED K+ CURRENTS IN INNER HAIR-CELLS OF THE GUINEA-PIG COCHLEA USING PHOTOLABILE CA2+ CHELATORS

Intracellular photorelease of Ca2+ from caged Ca2+(DM-nitrophen or nitr5) and the patch-clamp technique in the whole-cell configuration were used to investigate Ca2+-activated currents in inner hair cells (IHCs) of the mammalian cochlea. Photoliberation of intracellular Ca2+ activated outward currents with a mean amplitude of 260 +/- 110 pA when IHCs were voltage-clamped, near the resting membrane potential, at - 50 mV. The photoactivated currents were reversibly blocked by extracellular application of tetraethylammonium (TEA, 10 mM), neomycin (1 mM) and charybdotoxin (1 mu M), but not by apamin. The voltage dependence of membrane currents activated by photolysis of DM-nitrophen demonstrated a reversal potential near the K+ equilibrium potential (E(k)) and saturation near 0 mV. The presence of Ca2+-activated currents was further confirmed by the effects of extracellular adenosine 5'-triphosphate (ATP, IO mu M) and the Ca2+ ionophore ionomycin (10 mu M). Both agents raised intracellular Ca2+ and simultaneously activated outward currents when IHCs were voltage-clamped near the resting membrane potential. In experiments where currents were activated by depolarizing voltage steps, nifedipine (50 CIM) and Cd2+ (1 mM) reduced significantly (20-50%) the whole-cell outward currents, suggesting the presence of L-type Ca2+ currents activating Kf currents. These results are the first direct evidence for Ca2+-activated K+ currents in mammalian IHCs, these currents being potentially important for cell repolarization during sound-induced depolarization and synaptic transmission.