Insulin resistance (IR) is a common pathophysiological feature of Type 2 diabetes. Although the mechanisms leading to IR are still elusive, evidence has shown that aerobic exercise can reverse this process. To investigate the effects of aerobic exercise on IR, the authors created an IR animal model by feeding C57BL/6 mice a high-fat diet for 8 wk. They then compared the effect of 6 wk of treadmill training (60 min/d) at 75% VO2max on mice in normal-diet (NE) and high-fat-diet (HE) groups with their sedentary control groups. Levels of skeletal-muscle AMPKα (AMP-activated protein kinase α), ACC (acetyl-CoA carboxylases), and CPT1 (carnitine palmitoyltransferase 1) mRNA and AMPKα, pAMPK-Thr172, ACC, pACC-Ser79, and CPT1 protein expressions were analyzed. In addition, fasting serum levels of insulin, triglyceride, and cholesterol were measured. The results demonstrate that 6 wk of exercise increased AMPKα mRNA expression by 11% and 25 % (p < .01) in the NE and HE groups, respectively, and AMPKα protein expression by 37.9% and 20.1% (p < .01) in NE and HE compared with their sedentary control. In addition, ACC mRNA and protein expressions declined, whereas CPT1 mRNA and protein expressions were elevated in both exercise groups compared with sedentary control groups. In addition, pAMPK-Thr172 and pACC-Ser79 expression increased significantly in the NE and HE groups compared with sedentary control groups. In conclusion, our results demonstrate that 6 wk of aerobic exercise can effectively ameliorate IR by increasing the expression of AMPKα and pAMPK-Thr172, thereby activating the key enzymes that facilitate lipid metabolism.