The molecular pathway of ATP-sensitive potassium channel in endothelial cells for mediating arteriole relaxation

Abstract

Aims

The endothelial molecular pathway of a new ATP-sensitive potassium channel (K_ATP) opener natakalim was investigated in mesenteric arterioles of rats.

Main methods

A DMT wire myograph was used to evaluate the vasorelaxation effects of natakalim. Ca^2 + responses of endothelial cells induced by natakalim were measured by laser confocal fluorescence microscopy. NO assay kits and Western blotting were used.

Key findings

The new K_ATP opener natakalim significantly produced endothelium-dependent arteriolar dilation and increased endothelial cell intracellular calcium concentration ([Ca^2 +]_i) as well as NO release, which could be inhibited by SB366791 and capsazepine, the specific TRPV1 blockers. Additionally, down-regulation of endothelial TRPV1 by RNA interference inhibited the Ca^2 + influx induced by natakalim.

Significance

These results suggest that endothelial K_ATP mediated natakalim-induced vasorelaxation through increasing [Ca^2 +]_i and NO production. Activation of endothelial TRPV1 channels and subsequent Ca^2 + entry, and NO release at least partly contribute to endothelium-dependent vasorelaxation induced by natakalim.

Introduction

It has been proved that activation of ATP-sensitive potassium channels (K_ATP) in the vascular endothelial cell by K_ATP channel openers (KCOs) increases intracellular calcium concentration ([Ca^2 +]_i) as a consequence of potassium currents [9], [16], and the increased [Ca^2 +]_i plays an important role in promoting the endothelial cells to release nitric oxide (NO), an endothelium-derived relaxing factor (EDRF) contributing to endothelium-dependent relaxation of vessels. Our previous studies have showed that the novel KCO iptakalim is able to enhance NO secretion of cultured bovine aortic endothelial cells and rat aortic endothelial cells associated with the increased [Ca^2 +]_i [3], [4], [5]. However, it remains unclear which pathway is responsible for mediating Ca^2 + influx in endothelial cells induced by KCOs. Recent studies from several laboratories have implicated that Ca^2 +-permeable cation channels belonging to the transient receptor potential (TRP) channel family might be involved in the increase of [Ca^2 +]_i induced by multiple stimuli [6]. Some TRP channels such as TRP vanilloid 1 (TRPV1), TRPV4 and TRP canonical 4 (TRPC4) have been reported to mediate endothelium-dependent vasodilation in particular vascular beds [7].

TRPV1 is a member of non-selective cation channels, and expressed in vascular beds. This channel can be regulated by capsaicin, resiniferatoxin, ethanol, noxious temperature and extracellular acidosis [8]. Several reports found that endothelial TRPV1 channels mediated relaxation of blood vessels [9], [10], [11]. The relaxation by activation of TRPV1 channels could be attenuated by removal of vascular endothelial cells or inhibition of endothelial nitric oxide synthase (eNOS) with N^ω-nitro-_L-arginine methyl ester (L-NAME). These studies indicated that activation of endothelial TRPV1 channels induces Ca^2 + influx and then enhances NO release, which contributes to endothelium-dependent vascular relaxation.

Natakalim is a specific opener for SUR2B/Kir6.1 channels of K_ATP [5], [12]. Previous studies have shown that natakalim improved cardiac remodeling induced by pressure overload via protecting endothelial function, and led to endothelium-dependent dilation of the isolated tail artery helical strips pre-contracted with norepinephrine [13]. However, the associated molecular mechanisms are still unclear. Based on these evidences, we hypothesized that natakalim might induce Ca^2 + influx in endothelial cells through TRPV1 channels to enhance the production of NO, which mediates endothelium-dependent vasodilation.

The objectives of the present study are to determine whether TRPV1 channels participate in Ca^2 + entry in endothelial cells induced by natakalim and to elucidate the molecular mechanism of its vasodilatory effects.