Hydrogen peroxide induces vasorelaxation by enhancing 4-aminopyridine-sensitive Kv currents through S-glutathionylation

Hydrogen peroxide (H(2)O(2)) is an endothelium-derived hyperpolarizing factor. Since opposing vasoactive effects have been reported for H(2)O(2) depending on the vascular bed and experimental conditions, this study was performed to assess whether H(2)O(2) acts as a vasodilator in the rat mesenteric artery and, if so, to determine the underlying mechanisms. H(2)O(2) elicited concentration-dependent relaxation in mesenteric arteries precontracted with norepinephrine. The vasodilatory effect of H(2)O(2) was reversed by treatment with dithiothreitol. H(2)O(2)-elicited vasodilation was significantly reduced by blocking 4-aminopyridine (4-AP)-sensitive Kv channels, but it was resistant to blockers of big-conductance Ca(2+)-activated K(+) channels and inward rectifier K(+) channels. A patch-clamp study in mesenteric arterial smooth muscle cells (MASMCs) showed that H(2)O(2) increased Kv currents in a concentration-dependent manner. H(2)O(2) speeded up Kv channel activation and shifted steady state activation to hyperpolarizing potentials. Similar channel activation was seen with oxidized glutathione (GSSG). The H(2)O(2)-mediated channel activation was prevented by glutathione reductase. Consistent with S-glutathionylation, streptavidin pull-down assays with biotinylated glutathione ethyl ester showed incorporation of glutathione (GSH) in the Kv channel proteins in the presence of H(2)O(2). Interestingly, conditions of increased oxidative stress within MASMCs impaired the capacity of H(2)O(2) to stimulate Kv channels. Not only was the H(2)O(2) stimulatory effect much weaker, but the inhibitory effect of H(2)O(2) was unmasked. These data suggest that H(2)O(2) activates 4-AP-sensitive Kv channels, possibly through S-glutathionylation, which elicits smooth muscle relaxation in rat mesenteric arteries. Furthermore, our results support the idea that the basal redox status of MASMCs determines the response of Kv currents to H(2)O(2). ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00424-014-1513-3) contains supplementary material, which is available to authorized users.