Objective To assess the roles of phosphoinositide 3 kinase/serine-threonine kinase (PI3K/Akt) signal pathway in cardioprotection of fentanyl postconditioning and limb remote postconditioning in an in vivo rat model with myocardial ischemia reperfusion injury. Methods Thirty-two anesthetized male SD rats (weighed 250 g-350 g) were randomly allocated into the four groups: group C(control), group F(fentanyl postconditioning), group R(RIPOC), and group F-R (combined fentanyl postconditioning and RIPOC). All rats were treated with left anterior descending coronary artery(LAD) occluded for 30 min, followed by a 60-min reperfusion(LAD open) in vivo. The Olingo PI3k/Akt signaling pathway microarray(SA Bioscience)and Western-blot technique were used to distinguish the expression of genes related to PI3K/Akt pathway and levels of phosphorylated Akt in myocardial tissue sample from ischemia area. Results Expressions of many detected genes relating to PI3K/Akt signal pathway were significantly different among the four groups. As compared with group C, 9 genes showed a significant up-regulated expression in group F, 2 genes showed a significant up-regulated expression in group R, and a total number of 33 genes showed a significant up-regulated expression in group F-R. The Western-blot analysis revealed that the expression of phosphorylated Akt in myocardium increased significantly in groups F, R and F-R compared with group C. The expression of phosphorylated Akt in myocardium was also stronger in group F-R than in groups F and R. Conclusions Combined fentanyl postconditioning and RIPOC can produce an enhanced activation of PI3K/Akt signal pathway.