The synthesis and release of nitric oxide (NO) by the vascular endothelium is known to be a critical process in the regulation of arterial tone and the interaction of hematopoietic cells with the vessel wall. Recent studies have identified both small and intermediate conductance, Ca-activated K⁺ channels (SK_Ca and IK_Ca channels) in vascular endothelium, which become active in response to Ca-mobilizing agonists that stimulate NO production. Pharmacologic studies have demonstrated that selective toxin inhibitors (ie, apamin and charybdotoxin) of these channels interfere with agonist-induced vasordilation in intact resistance arteries; however, the cellular mechanism(s) underlying this phenomenon remains poorly defined. We have hypothesized that agonist-induced activation of endothelial SK_Ca and IK_Ca channels, leading to membrane hyperpolarization, is essential for promoting the entry of external Ca that is ultimately responsible for stimulating membrane-bound NO synthase. To examine this hypothesis, we have directly followed NO synthesis in cultured endothelial cells from human umbilical cord loaded with the NO-sensitive fluorescent dye DAF-2DA. Our results indicate that selectively blocking SK_Ca and IK_Ca channels with apamin and charybdotoxin, respectively, abolishes stimulated rises in NO, as well as elevations in intracellular free calcium, in response to the receptor agonists histamine or ATP. Preventing agonist-induced membrane hyperpolarization by raising the external [K⁺] to 80 mM also inhibits both stimulated elevations in cytosolic Ca and NO production. These findings, along with others, thus suggest a mechanistic model describing a critical role for SK_Ca and IK_Ca channels in supporting agonist-stimulated NO synthesis, leading to vasodilation.

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