Inhibition by nicardipine on endothelin-mediated inositol phosphate formation and Ca 2+ mobilization in smooth muscle cell

Abstract

We have investigated the effects of endothelin on phosphoinositide metabolism and Ca 2+ mobilization in cultured A10 cells. Endothelin stimulated a significant increase in inositol phosphate formation in a time- and dose-dependent manner. IP 3 was significantly elevated by 30 sec and reached a 2.0-fold above control at 1 min. The EC 50 for endothelin was 0.5 nM. The initiation of inositol phosphate formation was independent of extracellular Ca 2+, and the Ca 2+ ionophore, A23187, did not stimulate IP 3 formation. However, the sustained elevation of inositol phosphates was partially inhibited by incubating cells in buffer lacking Ca 2+ or in buffer containing nicardipine. Endothelin mobilized both intracellular and extracellular Ca 2+ reaching a peak intracellular concentration of 350 ± 11 nM by 1 min when cells were bathed with Ca 2+-complete buffer. Intracellular Ca 2+ remained 2-fold above baseline for at least 15 min. In contrast, when cells were exposed to endothelin in Ca 2+-free buffer, the peak value of {Ca 2+} i was 195 ± 20 nM and returned to baseline by 2 min. Nicardipine completely blocked the influx of extracellular Ca 2+ but did not interfer with the mobilization of intracellular stores. We conclude that endothelin produces a rapid and sustained elevation in inositol phosphate formation. The rapid production of IP 3 is consistent with the time course for mobilization of intracellular Ca 2+. Elevated cytosolic Ca 2+ levels are maintained by the influx of extracellular Ca 2+ through a nicardipine-sensitive Ca 2+ channel and are involved in the sustained formation of inositol phosphates. These data provide an explanation for the sustained, nicardipine-inhibitable contraction of coronary artery strips induced by endothelin.