Autocrine action and its underlying mechanism of nitric oxide on intracellular Ca2+ homeostasis in vascular endothelial cells

The rise in cytosolic Ca2+ concentration (Ca-i(2+)) in vascular endothelial cells (ECs) activates the production and release of nitric oxide (NO). NO modifies Ca-i(2+) homeostasis in many types of nonendothelial cells. However, its effect on endothelial Ca-i(2+) homeostasis at basal and excited states remains unclear. In the present study, to elucidate the effect of NO on basal Ca-i(2+) inositol 1,4,5-trisphosphate-induced Ca-i(2+) release (IICR) was blocked by expressing an antisense against type-1 inositol 1,1,B-trisphosphate receptors or by microinjecting heparin to individual ECs, and the effects of NO that was released by and diffused from adjacent IICR-intact ECs were recorded. After ATP or bradykinin stimulation, IICR-inhibited ECs showed a marked reduction of basal Ca-i(2+), which was abolished by NG-monomethyl-L-arginine monoacetate pretreatment. The reduction disappeared in sparsely seeded ECs. Exogenous NO gas mimicked the effect of ATP or bradykinin to reduce basal Ca-i(2+). Blocking plasma membrane Ca2+-ATPase Ca2+-ATPase (PMCA), but not Na+-Ca2+ exchange or sarcoplasmic/ endoplasmic reticulum Ca2+-ATPase, suppressed the reduction, indicating that the reduction resulted from a NO-dependent potentiation of PMCA. To elucidate the effect of NO on elevated Ca-i(2+), ATP-, bradykinin-, or thapsigargin-evoked Ca-i(2+) response in the presence and absence of NO production was compared in adjacent BOB-intact ECs. NO was found to potentiate PMCA, which, in turn, greatly attenuated agonist-evoked Ca2+ elevation. NO also potentiated Ca2+ influx, which markedly increased the sustained phase of Ca2+, elevation and possibly NO production. NO did not affect other Ca-i(2+)-elevating and Ca-i(2+)-sequestrating components. Thus, NO-dependent potentiation of PMCA is crucial for Ca-i(2+) homeostasis over a wide Ca-i(2+) range.