Recent evidence suggests that postischemic myocardial dysfunction (“stunning”) may be mediated by oxygen free radicals, but the exact time window during which the critical radical-mediated damage develops remains unknown. Furthermore, the evidence for the oxyradical hypothesis is indirect and, therefore, inconclusive. Thus, the potent and cellpermeable antioxidant JV-(2-mercaptopropionyl)-glycine (MPG) was administered as an intracoronary infusion (8 mg/kg/hr) to three groups of open-chest dogs undergoing a 15-mlnute coronary occlusion followed by 4 hours of reperfusion. In group I (n=8), the infusion of MPG was started 15 minutes before occlusion and ended 2 hours after reperfusion; in group II (n=9), MPG was started 1 minute before reperfusion and ended 2 hours thereafter; in group III (n= 10), MPG was started 1 minute after reperfusion and ended 2 hours and 15 minutes thereafter. Control dogs (group IV) (n= 10) received vehicle. Recovery of contractile function (assessed as systolic wall thickening) was equivalent in groups I and II, and in both groups it was substantially greater than in controls (p<0.005 at 4 hours). In contrast, in group III recovery of function was indistinguishable from controls. To determine whether the protection afforded by MPG was due to inhibition of free radical reactions, myocardial production of free radicals was directly assessed by intracoronary infusion of the spin trap o-phenyl JV-tert-butyl nitrone (PBN). In control dogs (group VII,n=6), radical adducts of PBN were released in the coronary venous blood after reperfusion, with a burst occurring in the first 5 minutes. MPG given as in group II (group V,n-5) markedly suppressed myocardial production of PBN adducts (Δ = − 98% over 3 hours,p<0.01 vs. controls); this effect was evident immediately after reperfusion. MPG given as in group III (group VI,n=5) also suppressed PBN adduct production (Δ =−83% over 3 hours,p<0.025 vs. controls), but this effect was delayed. Hence, the radicals important in myocardial stunning appear to be those generated immediately after reperfusion. In vitro studies demonstrated that MPG is an exceptionally powerful scavenger of • OH (rate constant=8.1x109M−1sec−1by pulse radiolysis) but has no significant effect on • O2(rate constant < 103M−1sec−1), H2O2(rate constant=1.6 M−1sec−1), or non- • OH-initiated lipid peroxidation, suggesting that removal of • OH is the major mechanism of the beneficial effects of MPG. These results demonstrate that a substantial portion (apparently most) of the damage responsible for myocardial stunning develops in the initial seconds of reperfusion and can be prevented by antioxidant therapy started just before reftow. The results further demonstrate that attenuation of this reperfusion-induced damage is associated with attenuation of free radical reactions in vivo, thereby providing direct evidence for a causal role of reactive oxygen species. It is proposed that myocardial stunning is, at least in part, a form of oxyradicalmediated "reperfusion injury."