Stimulation of Ca²⁺ influx by endothelin-1 is subject to negative feedback by elevated intracellular Ca²⁺

Endothelin-1 (ET-1) has been shown to require Ca²⁺ influx for activation of vascular smooth muscle in vivo, but in vitro models show that ET-1 mobilizes intracellular Ca²⁺ and is independent of extracellular Ca²⁺. We present data that suggest ET-1 modulates cellular responses through a dual mechanism involving both phosphatidylinositol turnover and Ca²⁺ channel activation. Addition of low concentrations of ET-1 (<10^-9 M) to serum-deprived quiescent Rat-1 cells stimulated Ca²⁺ influx while having little effect on diacylglycerol (DG) release or intracellular Ca²⁺ levels. In contrast, higher concentrations of ET-1 (>10^-9 M) stimulated intracellular Ca²⁺ transients and release of inositol trisphosphate (IP₃) and DG but did not activate Ca²⁺ uptake. Stimulation of Ca²⁺ influx at low [ET-1] could not be accounted for by depletion of intracellular IP₃-sensitive pools. Neither the stimulation of Ca²⁺ influx at low [ET-1] nor the inhibitory actions of high [ET-1] could be mimicked by the activation of protein kinase C. We tested the hypothesis that elevated intracellular Ca²⁺ was inhibitory for Ca²⁺ influx. When intracellular Ca²⁺ transients were maintained below ~165 nM by chelation with BAPTA or BAPTA derivatives with altered affinity for Ca²⁺, Ca²⁺ influx was stimulated over the entire range of ET-1 concentrations. In addition, experimentally elevating intracellular Ca²⁺ levels with the tumor promoter thapsigargin abolished ET-1-stimulated Ca²⁺ influx. These data suggest that the biological consequences of ET-1 release may be determined by local concentration differences. Thus in vascular smooth muscle cells ET-1 may act either to mobilize intracellular Ca²⁺ or to promote Ca²⁺ influx, depending on the distance from the endothelial cell source in the vascular wall. The activation of different processes by low and high ET-1 concentrations may determine the physiological response to ET-1 stimulation in vivo.