Recent studies have emphasized an important role for Oxytocin in protecting against myocardial ischemia-reperfusion injury. In this study, we assessed whether phosphatidylinositol-3-kinase (PI3K-AKT) pathway was involved in Oxytocin mediated cardioprotection. Isolated perfused rat hearts were subjected to 35minutes of ischaemia and 120minutes of reperfusion. Hearts underwent triphenyl-tetrazolium staining for infarct size assessment, or frozen for Western blot analysis. Oxytocin (1nM, 10 nM, 100nM) was administered throughout reperfusion in the presence and absence of the Oxytocin receptor antagonist of L371,527 (10nM), PI3K inhibitor Wortmannin (100nM). Data was analysed using one way ANOVA followed by Tukey's test (n = 4-6). Oxytocin (1nM, 10nM, 100nM) when administered during reperfusion significantly reduced infarct size when compared to control (22.7 ± 1% (1nM); 30.2 ± 71% (10nM); 37.3 ± 1.5% (100nM); vs. 56.1 ± 2.2% (control), p < 0.05). The protective effect of Oxytocin (1nM) was abrogated by administration of either L371,527 (10nM) (49.2 ± 3.1%, p < 0.001), Wortmannin (100 nM), (59.1 ± 4.0%, p < 0.001). Western blot analysis further demonstrated that Oxytocin receptor activation during reperfusion induced a significant increase in p-Akt(Ser473) compared to non-treated control hearts. Oxytocin dependent phosphorylation of Akt was abrogated by Wortmannin. This is the first study to show that Oxytocin receptor activation can protect the ischaemic-reperfused myocardium via recruitment of the PI3K-AKT cell Survival pathway.