Oxygen radical-mediated reduction in basal and agonist-evoked NO release in isolated rat heart

Oxygen free radicals (OFR) play a primary role in ischemia-reperfusion-mediated vascular dysfunction and this is paralleled by a loss of endothelial nitric oxide synthase (eNOS) activity. The authors tested whether a direct exposure to OFR may affect vascular relaxation by altering nitric oxide (NO) release. Effects of electrolysis(EL)-generated OFR on basal and agonist-evoked NO release were monitored in isolated rat hearts by oxyhemoglobin assay. Electrolysis-induced changes were compared with those obtained after 30 min perfusion with NOS and cyclooxygenase (COX) inhibitors NG-nitro-L-arginine methyl ester (L-NAME, 100 microM) and indomethacin (INDO, 1 m M). Electrolysis-generated hydroxyl radical (.OH) formed by.O2-and H2O2 via the Fenton reaction as revealed by Electron Paramagnetic Resonance (EPR). After EL, basal NO release declined by 60% and coronary perfusion pressure (CPP) increased by approximately 70%. L-NAME/INDO perfusion similarly lowered NO release (-63%) but increased CPP less than EL (56+/-3%P<0.03 v post-EL). In presence of excess substrates and cofactors eNOS activity was not affected by EL. Both acetylcholine (ACh; 1 microM) and bradykinin (BK; 10 n M) had minimal effect in reversing EL-induced vasoconstriction, whereas both partially reversed L -NAME/INDO-mediated constriction. Sodium nitroprusside (SNP, 1 microM) completely reversed L-NAME/INDO constriction and partly countered that after EL (-38+/-2.5, P<0.001). Acetylcholine-evoked NO release was nearly abolished by both treatments whereas BK still elicited partial NO release after eNOS/cyclooxygenase inhibition (P<0.001) but not after EL. In conclusion, OFR severely impair NO-mediated coronary vasorelaxation affecting both basal and agonist-evoked NO release but not eNOS activity. However, EL also significantly blunts NOS/COX-independent vasodilation suggesting alteration of other vasodilatative pathways.