Na<SUP>+</SUP>/Ca<SUP>2+</SUP> exchange facilitates Ca<SUP>2+</SUP>-dependent activation of endothelial nitric-oxide synthase

作     者：Teubl, M Groschner, K Kohlwein, SD Mayer, B Schmidt, K 

作者机构：Karl Franzens Univ Graz Inst Pharmakol & Toxikol A-8010 Graz Austria Graz Univ Technol Inst Biochem & Lebensmittelchem A-8010 Graz Austria 

出 版 物：《JOURNAL OF BIOLOGICAL CHEMISTRY》 (生物化学杂志)

年 卷 期：1999年第274卷第41期

页      面：29529-29535页

核心收录：

学科分类：0710[理学-生物学] 071010[理学-生物化学与分子生物学] 07[理学] 

主　　题：阿米洛利/药理学 缓激肽/药理学 钙/药理学 钙通道阻滞药/药理学 窖蛋白1 细胞质膜微囊蛋白 细胞 培养的 瓜氨酸/代谢 内皮 血管 酶激活/药物作用 荧光抗体技术 Fura-2/化学 咪唑类/药理学 膜蛋白质类/代谢 莫能星/药理学 一氧化氮合酶/代谢 钠/药理学 钠钙交换蛋白/代谢 猪 动物 

摘      要：Recent evidence suggests the expression of a Na+/ Ca2+ exchanger (NCX) in vascular endothelial cells, To elucidate the functional role of endothelial NCX, we studied Ca2+ signaling and Ca2+-dependent activation of endothelial nitric-oxide synthase (eNOS) at normal, physiological Na+ gradients and after loading of endothelial cells with Na+ ions using the ionophore monensin. Monensin-induced Na+ loading markedly reduced Ca2+ entry and, thus, steady-state levels of intracellular free Ca2+ ([Ca2+](i)) in thapsigargin-stimulated endothelial cells due to membrane depolarization. Despite this reduction of overall [Ca2+](i), Ca2+-dependent activation of eNOS was facilitated as indicated by a pronounced leftward shift of the Ca2+ concentration response curve in monensin-treated cells. This facilitation of Ca2+-dependent activation of eNOS was strictly dependent on the presence of Na+ ions during treatment of the cells with monensin, Na+-induced facilitation of eNOS activation was not due to a direct effect of Na+ ions on the Ca2+ sensitivity of the enzyme. Moreover, the effect of Na+ was not related to Na+ entry-induced membrane depolarization or suppression of Ca2+ entry, since neither elevation of extracellular K+ nor the Ca2+ entry blocker 1-(beta-[3-(4-methoxyphenyl)-propoxy]-4-methoxyphenethyl)-1H-imidazole hydrochloride (SK&F 96365) mimicked the effects of Na+ loading. The effects of monensin were completely blocked by 3 ,4 -dichlorobenzamil, a potent and selective inhibitor of NCX, whereas the structural analog amiloride, which barely affects Na+/Ca2+ exchange, was ineffective. Consistent with a pivotal role of Na+/Ca2+ exchange in Ca2+-dependent activation of eNOS, an NCX protein was detected in caveolin-rich membrane fractions containing both eNOS and caveolin-1. These results demonstrate for the first time a crucial role of cellular Na+ gradients in regulation of eNOS activity and suggest that a tight functional interaction between endothelial NCX and eNOS may tak