Endogenous Peroxynitrite Generation Causes a Subsequent Suppression of Coronary Arterial Contraction to Serotonin

Abstract

High levels of exogenous peroxynitrite (ONOO−) have been reported to cause coronary vascular relaxation by a mechanism that appears to involve the subsequent generation of nitric oxide (NO). In this study, we examined if endogenous vasoactive levels of ONOO−are formed from endogenous superoxide anion (O2⨪) upon exposure of isolated endothelium-removed bovine coronary arteries (BCA) to biological levels of NO. During exposure of BCA to ∼50 nmNO for 2 min, the level of endogenous O2⨪detected by lucigenin-dependent chemiluminescence (CL) was markedly decreased and an increase in luminol-dependent CL was observed, consistent with the detection of ONOO−generation. NO treatment caused a decrease in contraction of BCA to 0.1–3 μmserotonin (5-HT). This suppression of contraction to 5-HT was completely prevented by preincubation prior to NO exposure with agents that prevent endogenous O2⨪production (10 μmdiphenyliodonium) or trap intracellular O2⨪(10 mmTiron) or ONOO−(0.1 mmurate), and by post-NO treatment with an agent that traps NO (1 μmoxyhemoglobin) or prevents the stimulation of cGMP production by NO (10 μmmethylene blue). The NO treatment caused a subsequent release of NO (measured in the head space after a 5-min equilibration with 95% N2–5% CO2), and this subsequent release of NO was reduced by the presence of urate during NO exposure and by depletion of endogenous tissue glutathione (by pretreatment with 7 mmdiethyl maleate). Thus, exposure of BCA to elevated physiological levels of NO causes a prolonged suppression of contraction to 5-HT which appears to result from endogenous ONOO−formation and a thiol-dependent process that traps and subsequently releases vascular relaxant levels of NO.