Control of intracellular calcium by ATP in isolated outer hair cells of the guinea-pig cochlea

Abstract

1. Intracellular calcium levels were monitored in isolated outer hair cells of the guinea-pig cochlea using the calcium-sensitive dye Fura-2. 2. The calcium in the cells was studied during application of ATP externally applied from a pipette. ATP induced a rise of intracellular calcium which could be separated into two components: a rapid rise, peaking in 20 s, localized around the apical end of the cell, and a slower rise, peaking in 50-150 s but spread throughout the cell. The effects were observed with 5, 25 and 100 microM-ATP concentrations. 3. In the absence of external Ca2+, ATP was still able to trigger a rise in Ca2+, but with a longer delay. Under these conditions, the cells did not show the initial rapid Ca2+ rise. The result suggests that ATP can mobilize intracellular stores. 4. A rise in intracellular Ca2+ was also observed when 5 mM-caffeine was applied to the bath. 5. Simultaneous measurements were made of whole-cell currents and intracellular calcium. ATP activated an inward current at resting potentials of -60 mV. Internal Ca2+ levels increased during the inward current. In current-clamped cells Ca2+ levels also increased during the associated depolarization produced by ATP. 6. Adenosine (150 microM) did not produce any measurable inward current. Acetylcholine (ACh, 100 microM-1 mM) produced only a small rise in Ca2+. However, applied simultaneously with ATP, ACh suppressed the rise in intracellular Ca2+ produced by ATP, with the kinetics of a competitive antagonist. 7. Intracellular Ca2+ increased with step depolarizations of the cell above -20 mV during whole-cell clamp. Large rises in Ca2+ were also observed on depolarizing the cell with isotonic KCl. 8. Calcium levels in supporting cells of the organ of Corti were sensitive to ATP. In these cells, rises in intracellular Ca2+ did not require the presence of extracellular Ca2+. 9. It is concluded that the organ of Corti contains receptors for ATP on a variety of the cells. ATP controls a direct entry of Ca2+ through the membrane and also may mobilize intracellular stores.