Received for publication April 23, 2008.
Revised May 19, 2008.
Accepted for publication June 4, 2008.

Endothelin-1 Regulates Cardiac L-Type Calcium Channels via NAD(P)H Oxidase-Derived Superoxide

Abstract

It has been shown that reactive oxygen species (ROS) are involved in the intracellular signaling response to G-protein coupled receptor stimuli in vascular smooth muscle cells and in neurons. In the present study, we tested the hypothesis that NAD(P)H oxidase-derived ROS are involved ET-1-induced L-type calcium channel activation in isolated cardiac myocytes. ET-1 (10 nM) induced a 2-fold increase in L-type calcium channel open state probability (NPo). This effect of ET-1 was abolished by the ET_A receptor antagonist, BQ-123 (1 µM) but was not altered in the presence of an ET_B receptor antagonist, BQ-788 (1 µM). Pretreatment of cells with the ROS scavenger tempol (100 µM), superoxide dismutase (PEG-SOD 25 units/ml), or the NAD(P)H-oxidase inhibitor gp91ds-tat (5 µM) significantly attenuated ET-1-induced increases in calcium channel NPo. Tempol, SOD, and gp91ds-tat alone had no effect on basal calcium channel activity. In addition, ET-1 significantly increased NAD(P)H oxidase activity and elevated intracellular superoxide levels in cultured cardiac myocytes. The superoxide generator, xanthine-xanthine oxidase (10 mM, 20 mU/ml), also increased calcium channel NPo in cardiac myocytes, mimicking the effect of ET-1. These observations provide the first evidence that ET-1 induces the activation of L-type Ca²⁺ channels via stimulation of NAD(P)H-derived superoxide production in cardiac myocytes.

Key words: Endothelin receptors, Endothelin-1, Heart, calcium channel, cardiomyocyte, superoxide