Endothelin-mediated calcium signaling in preglomerular smooth muscle cells

This study was performed to test the hypothesis that endothelin peptides differentially influence intracellular calcium concentration ([Ca(2+)](i)) in preglomerular microvascular smooth muscle cells (MVSMC), in part through activation of endothelin (ET)(A) receptors. Experiments were performed in vitro with the use of single MVSMC freshly isolated from rat preglomerular microvessels. The effect of ET-1, ET-2, and ET-3 on [Ca(2+)](i) was measured with the use of the calcium-sensitive dye, fura 2, and standard fluorescence microscopy techniques. Baseline [Ca(2+)](i) averaged 84+/-3 nmol/L (n=141 cells from 23 dispersions). ET-1 concentrations of 1, 10, and 100 nmol/L evoked peak increases in [Ca(2+)](i) of 48+/-16, 930+/-125, and 810+/-130 nmol/L, respectively. The time course of the [Ca(2+)](i) response was biphasic, beginning with a rapid initial increase followed by a sustained plateau phase or a period during which [Ca(2+)](i) oscillated sharply. Similar responses were observed after ET-2 administration. In contrast, ET-3 stimulated monophasic increases in [Ca(2+)](i) of only 14+/-5, 33+/-16, and 44+/-19 nmol/L at peptide concentrations of 1, 10, and 100 nmol/L, respectively. These responses are significantly smaller than responses to ET-1 or ET-2, respectively. The relative contributions of calcium mobilization and calcium influx in the response to ET-1 were also evaluated. Removal of calcium from the bathing medium did not significantly alter the peak response to 10 nmol/L ET-1 but abolished the late phase elevation of [Ca(2+)](i). These data demonstrate that endothelin peptides increase [Ca(2+)](i) in preglomerular MVSMC. The concentration-response profiles are consistent with the response involving activation of ET(A) receptors. Furthermore, these results suggest that ET-1 increases [Ca(2+)](i) by stimulating both the release of intracellular calcium and the influx of calcium from the extracellular medium.