Calcium homeostasis in rabbit ventricular myocytes. Disruption by hypochlorous acid and restoration by dithiothreitol

Abstract

Hypochlorous acid (HOCl) is a toxic oxidant produced by neutrophils at sites of cardiac inflammation. To examine the effect of this oxidant on Ca2+ homeostasis in the heart, isolated rabbit ventricular myocytes were iontophoretically loaded with the Ca2+ indicator fura 2 and superfused with 100 μM HOCl under voltage-clamp conditions. Ca2+ transients and the corresponding Ca2+ currents were elicited by 300-msec depolarizing pulses from -40 to 0 mV. Within 200 seconds after HOCl addition, the amplitude of the Ca2+ transients was reduced from 402±89 to 82±29 nM (p<0.01) while intracellular free Ca2+ concentration ([Ca2+](i)) increased from 78±16 to 265±48 nM (p<0.01). During this time, the amplitude of the slow inward currents increased by 10%, while steady-state holding current remained stable. This sustained steady-state rise in [Ca2+](i) occurred even in the absence of extracellular Ca2+ but was virtually abolished by a 20-second preexposure to 10 mM caffeine, suggesting that the major source of this Ca2+ was the sarcoplasmic reticulum. Although washout of HOCl failed to induce recovery, subsequent exposure to the dithiol reducing agent dithiothreitol caused a rapid restoration of both the steady-state [Ca2+](i) and Ca2+ transient amplitude. We conclude that 1) HOCl caused a rise of [Ca2+](i) by inducing the release of Ca2+ from internal stores and impairing cellular extrusion mechanisms and 2) these effects occur through alteration of protein thiol redox status.