Background: Ischemic postconditioning (IPost) and erythropoietin (EPO) have been shown to attenuate myocardial reperfusion injury using similar signalling pathways. Aims: To examine whether EPO is as effective as IPost to decrease post-ischemic myocardial injury in both Langendorff-isolated heart and in vivo ischemia-reperfusion rat models. Methods: Rat hearts were subjected to 25min ischemia followed by 30min or 2h of reperfusion in the isolated-heart study. Rats underwent 45min ischemia followed by 24h of reperfusion in the in vivo study. In both studies, control group (n=12; ischemia-reperfusion only) was compared to IPost (n=16; 3 cycles of 10 sec reperfusion/10 sec ischemia) and EPO (n=12; 1000 IU/kg) at the onset of reperfusion. Results: 1) In the isolated hearts IPost or EPO significantly improved post-ischemic recovery of left ventricular developed pressure (LVDP). EPO induced better LVDP than IPost at 30min reperfusion (73.18±10.23mmHg vs. 48.11±7.92mmHg, p<0.05). After 2h of reperfusion, infarct size was significantly lower in EPO-treated hearts as compared with IPost and control (14.36 ± 0.60%, 19.11±0.84% and 36.21±4.20% of the left ventricle respectively, p<0.05). GSK-3 phosphorylation, at 30min of reperfusion, was significantly higher with EPO as compared with IPost. PI3K and ERK1/2 inhibitors abolished both EPO- and IPost-mediated cardioprotection. 2) In vivo, IPost and EPO induced infarct size reduction as compared with control (40.5±3.6% and 28.9±3.1% respectively, vs. 53.7±4.3% of the area at risk, p<0.05). Again, EPO decreased significantly more infarct size and transmurality than IPost (p<0.05). Conclusion: Using our protocols, EPO showed better protective effects than IPost against reperfusion injury through higher phosphorylation of GSK-3.