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

Sang Woong Park,Jeong Min Kim,Bokyung Kim,Hyun Ju Noh,Shin-Young Ryu,Hana Cho,Young Min Bae,Dong Jun Sung,Jae Gon Kim,Kyeongjin Kang

doi:10.1007/s00424-014-1513-3

Abstract

Hydrogen peroxide (H2O2) is an endothelium-derived hyperpolarizing factor. Since opposing vasoactive effects have been reported for H2O2 depending on the vascular bed and experimental conditions, this study was performed to assess whether H2O2 acts as a vasodilator in the rat mesenteric artery and, if so, to determine the underlying mechanisms. H2O2 elicited concentration-dependent relaxation in mesenteric arteries precontracted with norepinephrine. The vasodilatory effect of H2O2 was reversed by treatment with dithiothreitol. H2O2-elicited vasodilation was significantly reduced by blocking 4-aminopyridine (4-AP)-sensitive Kv channels, but it was resistant to blockers of big-conductance Ca2+-activated K+ channels and inward rectifier K+ channels. A patch-clamp study in mesenteric arterial smooth muscle cells (MASMCs) showed that H2O2 increased Kv currents in a concentration-dependent manner. H2O2 speeded up Kv channel activation and shifted steady state activation to hyperpolarizing potentials. Similar channel activation was seen with oxidized glutathione (GSSG). The H2O2-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 H2O2. Interestingly, conditions of increased oxidative stress within MASMCs impaired the capacity of H2O2 to stimulate Kv channels. Not only was the H2O2 stimulatory effect much weaker, but the inhibitory effect of H2O2 was unmasked. These data suggest that H2O2 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 H2O2.Electronic supplementary materialThe online version of this article (doi:10.1007/s00424-014-1513-3) contains supplementary material, which is available to authorized users.

Highlights

Reactive oxygen species (ROS) are detrimental to biological processes and contribute to disease conditions such as inflammation, ischemia–reperfusion injury, atherosclerosis, diabetes mellitus, and hypertension
The present study resolves some of this complexity by providing direct evidence for an effector molecule that can mediate H2O2-induced vasodilation through the Kv channel and proposes that S-glutathionylation underlies the stimulatory effect of H2O2 on Kv channels
H2O2 relaxed the rat mesenteric artery that was precontracted with an agonist

Summary

Introduction

Reactive oxygen species (ROS) are detrimental to biological processes and contribute to disease conditions such as inflammation, ischemia–reperfusion injury, atherosclerosis, diabetes mellitus, and hypertension. Several studies have identified KCa channels as putative targets that are activated in the process of H2O2induced vasodilation [81], while some other groups indicate that H2O2 induced a vasorelaxation through opening of ATPdependent K+ (KATP) channels [74]. Studies using cloned Kv1.5, a major component of Kv current in coronary arteries, show that H2O2 increases Kv1.5 current for voltages < +20 mV but decreases it for high depolarizing voltages [12]. It is still uncertain whether H2O2 acts as a vasodilator. These differences may depend on experimental design and the specific vascular bed or vessel being studied [42, 47]

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Conclusion