Nongenomic stimulation of nitric oxide release by estrogen is mediated by estrogen receptor α localized in caveolae

Acute administration of 17 beta-estradiol (E-2) exerts antiatherosclerotic effects in healthy postmenopausal women. The vasoprotective action of E-2 may be partly accounted for by a rapid increase in nitric oxide (NO) levels in endothelial cells (ECs). However, the signaling mechanisms producing this rise are unknown, In an attempt to address the short-term effect of E-2 on endothelial NO production, confluent bovine aortic endothelial cells (BAECs) were incubated in the absence or presence of E-2, and NO production was measured. Significant increases in NO levels were detected after only 5 min of E-2 exposure without a change in the protein levels of endothelial NO synthase (eNOS). This short-term effect of estrogen was significantly blunted by various ligands which decrease intracellular Ca2+ concentration. Furthermore, plasma membrane-impermeable BSA-conjugated E-2 (E(2)BSA) stimulated endothelial NO release, indicating that in the current system the site of action of E-2 is on the plasma membrane rather than the classical nuclear receptor. The partial antagonist tamoxifen did not block E-2-induced NO production; however, a pure estrogen receptor alpha (ER alpha) antagonist ICI 182,780 completely inhibited E-2-stimulated NO release. The binding of E-2 to the membrane was confirmed using FITC-labeled E(2)BSA (E(2)BSA-FITC). Western blot analysis showed that plasmalemmal caveolae possess ER alpha in addition to well-known caveolae-associated proteins eNOS and caveolin. This study demonstrates that the nongenomic and short-term effect of E-2 on endothelial NO release is Ca2+-dependent and occurs via ER alpha localized in plasmalemmal caveolae. (C) 1999 Academic Press.