Calcium influx in vestibular hair cells

In frog semicircular canal hair cells, recorded via whole-cell voltage-clamp, L-type and R-type Ca currents were detected. The former was non-inactivating and nifedipine-sensitive (5 microM); the latter inactivated partially, was resistant to w-conotoxin GVIA (5 microM), w-conotoxin MVIIC (5 microM), and w-agatoxin IVA (0.4 microM), but was sensitive to mibefradil (10 microM). Both currents were sensitive to Ni and Cd (>10 microM). In some cells, the L-type current amplitude increased almost twofold upon repetitive stimulation, whereas the R type current remained unaffected. Eventually, run-down occurred for both currents, but was prevented by the protease inhibitor calpastatin. The R-type current peak component ran down first, without changing its plateau, suggesting that two channel types generate the R-type current. This peak component appeared at -40 mV, reached a maximal value at - 30 mV, and became undetectable for voltages near to 0 mV, suggestive of a novel transient current: its inactivation was indeed reversibly removed when Ba was the charge carrier. The L-type current and the R-type current plateau were appreciable at -60 mV and peaked at -20 mV: the former current did not reverse for voltages up to +60 mV, the latter reversed between +30 and +60 mV due to an outward Cs current flowing through the same Ca channel. Recovery from inactivation (investigated using the standard two-pulse protocol) required times on the order of 100 ms at -120 mV and 300 ms at -70 mV. Such lengthy recovery times are comparable with the speed at which intracellular Ca is restored to its physiological level, upon returning to the holding potential. The faster recovery occurring at -120 mV when compared to -70 mV can be then explained by an acceleration of the Ca extrusion, possibly via a Na:Ca exchanger. The inactivating R-type channel may be functionally important in producing fast (synchronous) transmitter release in response to short, strong stimuli by boosting Ca entry to quickly elìcit synaptic transmission while, at the same time, preventing too large Ca influx which could be metabolically costly or even lethal to the cell. The non-inactivating R-type channels may instead sustain the ongoing spontaneous receptor activity which could also be mediated by the L-type channel. Indeed, the L-type channel can carry large, sustained Ca current, it is highly Ca selective, and is regulated by an intracellular mechanism which may be important for the response to rather weak, prolonged stimuli. The L-type channels could also cause large Ca changes throughout the cytoplasm: it is expected that strict control be exerted over the Ca transport generated by these channels, through a regulatory mechanism the sign of which is the run-up. This controlled Ca uptake, could also play an important role in replenishing the intracellular stores. The Ca influx provided by the three channels may also activate the Ca¬dependent K current which resets the system by repolarizìng the cell.