Existence of voltage-dependent Ca²⁺ channels in vestibular dark cells: cytochemical and whole-cell patch-clamp studies

Abstract

To determine whether functional Ca²⁺ channels are present in vestibular dark cells, changes in intracellular Ca²⁺ concentration ([Ca²⁺]_i) due to K⁺ applications were measured using the Ca²⁺-sensitive dye (fura-2) and patchclamp whole-cell recordings were made in dark cells isolated from the ampullae of the semicircular canal of the guinea pig. Exchange of the external solution with a buffer medium containing a high K⁺ concentration (80 mM K⁺ or 150 mM K⁺) caused a concentration-dependent increase in [Ca²⁺]_i in vestibular dark cells. Application of 1 μM nifedipine as a Ca²⁺ channel antagonist completely blocked the increase in [Ca²⁺]_i. Further treatment with 10 μM BAY K 8644 as a Ca²⁺ channel agonist caused an increase in [Ca²⁺]_i. In the patch-clamp whole-cell recordings a 1-s depolarizing pulse given into the dark cell in the presence of a high barium concentration (50 mM Ba²⁺) induced an inward current. In determining the current-voltage relationship, a current was detected at a potential that depolarized at −50 mV and was maximal at +10 mV. This inward current was completely blocked by 1 mM La³⁺ as a Ca²⁺ channel antagonist. These findings suggest the presence of voltage-dependent Ca²⁺ channels in dark cells, which have a presumed function in the regulation of [Ca²⁺]_i in the vestibular endolymph.