Hydrogen sulfide dilates rat mesenteric arteries by activating endothelial large-conductance Ca2+-activated K+ channels and smooth muscle Ca2+ sparks

Abstract

We have previously shown that hydrogen sulfide (H2S) reduces myogenic tone and causes relaxation of phenylephrine (PE)-constricted mesenteric arteries. This effect of H2S to cause vasodilation and vascular smooth muscle cell (VSMC) hyperpolarization was mediated by large-conductance Ca2+-activated potassium channels (BKCa). Ca2+ sparks are ryanodine receptor (RyR)-mediated Ca2+-release events that activate BKCa channels in VSMCs to cause membrane hyperpolarization and vasodilation. We hypothesized that H2S activates Ca2+ sparks in small mesenteric arteries. Ca2+ sparks were measured using confocal microscopy in rat mesenteric arteries loaded with the Ca2+ indicator fluo-4. VSMC membrane potential ( Em) was measured in isolated arteries using sharp microelectrodes. In PE-constricted arteries, the H2S donor NaHS caused vasodilation that was inhibited by ryanodine (RyR blocker), abluminal or luminal iberiotoxin (IbTx, BKCa blocker), endothelial cell (EC) disruption, and sulfaphenazole [cytochrome P-450 2C (Cyp2C) inhibitor]. The H2S donor NaHS (10 μmol/l) increased Ca2+ sparks but only in the presence of intact EC and this was blocked by sulfaphenazole or luminal IbTx. Inhibiting cystathionine γ-lyase (CSE)-derived H2S with β-cyano-l-alanine (BCA) also reduced VSMC Ca2+ spark frequency in mesenteric arteries, as did EC disruption. However, excess CSE substrate homocysteine did not affect spark activity. NaHS hyperpolarized VSMC Em in PE-depolarized mesenteric arteries with intact EC and also hyperpolarized EC E m in arteries cut open to expose the lumen. This hyperpolarization was prevented by ryanodine, sulfaphenazole, and abluminal or luminal IbTx. BCA reduced IbTx-sensitive K+ currents in freshly dispersed mesenteric ECs. These results suggest that H2S increases Ca2+ spark activity in mesenteric artery VSMC through activation of endothelial BKCa channels and Cyp2C, a novel vasodilatory pathway for this emerging signaling molecule.