Roles of the RhoA-ROCK Signaling Pathway in the Endothelial H 2 S Production and Vasodilation in Rat Cerebral Arteries.

Cerebral endothelial H 2 S protects against cerebral ischemia-reperfusion injury through vasodilation, but its cerebral vasodilation mechanism and regulation of production are poorly understood. The RhoA-ROCK pathway plays important roles in vascular function. In this study, the roles of this pathway in the endothelial H 2 S production and vasodilation in rat cerebral arteries were investigated. Acetylcholine significantly increased H 2 S-generating enzyme cystathionine-γ-lyase (CSE) and 3-mercaptopyruvate sulfurtransferase (3-MST) protein expressions and H 2 S production in rat cerebrovascular endothelial cells (ECs), but the increases were markedly decreased by the M receptor blocker atropine or the CSE inhibitor dl-propargylglycine. Pretreatment with dl-propargylglycine or the 3-MST inhibitor l-aspartic acid markedly reduced the acetylcholine-increased H 2 S; CSE protein expression and H 2 S levels in the ECs were obviously attenuated by the RhoA agonist U 46619 but increased by the RhoA inhibitor C3 transferase. U 46619 also reduced 3-MST protein expression; Acetylcholine markedly inhibited RhoA protein expression and activity, but the inhibition was obviously reversed by atropine, dl-propargylglycine, and l-aspartic acid, respectively; Acetylcholine-induced endothelium-dependent vasodilation in rat cerebral basilar artery was significantly attenuated by pretreatment with dl-propargylglycine or l-aspartic acid or RhoA inhibitor CCG-1423 or ROCK inhibitor KD025, and was further decreased by co-pretreatment with dl-propargylglycine (or l-aspartic acid) and CCG-1423 (or KD025); NaHS significantly relaxed rat cerebral basilar artery vascular smooth muscle cells and inhibited ROCK 1/2 activities, phosphorylated myosin light chain (MLC) protein expression, and KCl-increased [Ca 2+ ] i , but these relaxation and inhibitions were markedly attenuated by pretreatment with C3 transferase or ROCK inhibitor Y27632. Our results demonstrated that endothelial H 2 S production is promoted by activation of the M receptor but inhibited by the RhoA-ROCK pathway in rat cerebral arteries; the endothelial H 2 S induces cerebral vasodilation by inhibiting this pathway to reduce phosphorylation of MLC and [Ca 2+ ] i in vascular smooth muscle cells.