Peroxynitrite resistance of sarco/endoplasmic reticulum Ca 2+ pump in pig coronary artery endothelium and smooth muscle

Abstract

We examined the effects of peroxynitrite pre-treatment on sarco/endoplasmic reticulum Ca 2+ (SERCA) pump in pig coronary artery smooth muscle and endothelium. In saponin-permeabilized cells, smooth muscle showed much greater rates of the SERCA Ca 2+ pump-dependent 45 Ca 2+ uptake/mg protein than did the endothelial cells. Peroxynitrite treatment of cells inhibited the SERCA pump more severely in smooth muscle cells than in endothelial cells. To determine implications of this observation, we next examined the effect of the SERCA pump inhibitor cyclopiazonic acid (CPA) on intracellular Ca 2+ concentration of intact cultured cells. CPA produced cytosolic Ca 2+ transients in cultured endothelial and smooth muscle cells. Pre-treatment with peroxynitrite (200 μM) inhibited the Ca 2+ transients in the smooth muscle but not in the endothelial cells. CPA contracts de-endothelialized artery rings and relaxes precontracted arteries with intact endothelium. Peroxynitrite (250 μM) pre-treatment inhibited contraction in the de-endothelialized artery rings, but not the endothelium-dependent relaxation. Thus, endothelial cells appear to be more resistant than smooth muscle to the effects of peroxynitrite at the levels of SERCA pump activity, CPA-induced Ca 2+ transients in cultured cells, and the effects of CPA on contractility. The greater resistance of endothelium to peroxynitrite may play a protective role in pathological conditions such as ischemia-reperfusion when excess free radicals are produced.