Aim To explore the improvement effects of Urocortin-I on myocardial monophasic action potential and oxidative inflammatory response in ischemia-reperfusion (I/R) myocardium by activatingAkt/GSK-3 β pathway. Methods Langendorff perfusion model of isolated myocardium was prepared and then divided into control group, I/R group, Urocortin-I group and Urocortin-I+LY group. The control group was given routine perfusion, I/R group was given routine preconditioning, Urocortin-I group was given Urocortin-I preconditioning, Urocortin-I+LY group was given Urocortin-I and LY2942 preconditioning before ischemia-reperfusion. The differences of myocardial enzymes, infarct size, monophasic action potential, inflammatory factors, oxidative stress products and Akt/GSK-3 β pathway molecules among the four groups were compared. Results Compared with the control group, the contents of LDH, CK-MB, TNF-α, IL-6, ICAM-1, ROS, MDA and the size of myocardial infarction significantly increased, while the levels of APA, APD50, APD90 and the expression of p-Akt, p-GSK-3β significantly decreased in the I/R group (P<0.05). Compared with the I/R group, the contents of LDH, CK-MB, TNF-α, IL-6, ICAM-1, ROS, MDA and the size of myocardial infarction significantly decreased, while the levels of APA, APD50, APD90 and the expressions of p-Akt, p-GSK-3β significantly increased in the Urocortin-I group (P<0.05). Compared with the Urocortin-I group, the contents of LDH, CK-MB, TNF-α, IL-6, ICAM-1, ROS, MDA and the size of myocardial infarction significantly increased, while the levels of APA, APD50, APD90 and the expressions of p-Akt, p-GSK-3β significantly decreased in the Urocortin-I+LYgroup (P<0.05).Conclusion Urocortin-I improves monophasic action potential and oxidative inflammation in I/R myocardium by activating Akt/GSK-3 β pathway.