Objective The purpose of the present study was to observe the muscle glucose metabolism by determining the changes in sarcolemma GLUT4 protein content and muscle AMPK activity in overtraining rats.Methods Twenty-seven SD rats were randomly divided into group A(sedentary),group B(underwent endurance exercise 60 minutes/day)and group C(underwent endurance exercise 120 minutes/day).Both exercise groups trained 6 days/wk for 9 weeks.Muscle glycogen,sarcolemma GLUT4 contents and muscle AMPK activity were determined after the 9-week training.Results Sarcolemma GLUT4 content and muscle AMPK activity increased,muscle glycogen contents tended to rise in group B as compared with that in group A.Sarcolemma GLUT4 content and muscle AMPK activity decreased,but muscle glycogen contents were equivalent in group C as compared with that in group B.Conclusion Muscle AMPK activity deceased,and the translocation of GLUT4 protein was inhibited in the overtraining rats.The results could not support a glycogen depletion hypothesis.