Objective Glucagon-like peptide-1 (GLP-1) shows beneficial effects on reduction of obesity and insulin resistance. In the previous study, we found that treatment of obese diabetic mice with GLP-1 expressing recombinant adenoviruses reduced fat mass and increased energy expenditure. Muscle is a major organ for controlling of insulin resistance and energy metabolism. Thus, we investigated whether GLP-1 improves energy metabolism through fat oxidation in muscle. Methods We examined the expression of GLP-1 receptor in mouse muscle and C2C12 myotubes by RT-PCR. We analyzed fat oxidation by measurement of oxygen consumption rate (OCR) in palmitate (with or without exendin-4) treated C2C12 myotubes using seahorse XF24 analyzer. The expression of fat oxidation related molecules was analyzed by qRT-PCR and immunoblotting in C2C12 myotubes and muscle of high fat diet fed mice injected with exendin-4. Results GLP-1 receptor expressed in both muscle and differentiated C2C12 myotubes. The basal OCR was significantly increased in exendin-4 treated C2C12 myotubes. After palmitate addition, the OCR was dramatically increased in control and exendin-4 treated group, and exendin-4 treated group further increased compared with control group. However, a GLP-1 receptor antagonist inhibited this increase. The expression of UCP1 and PPAR- α mRNA and the protein expression of p-AMPK, UCP1 and PPAR-α were significantly increased in exendin-4 treated C2C12 myotube. In addition, intra-muscular injection of exendin-4 in HFD-fed mice significantly increased the expression of p-AMPK, PPAR-α and UCP-1 in muscle. Conclusion We suggested that GLP-1 increased energy metabolism through up-regulation of fat oxidation in muscle, which may contribute to reduced obesity and insulin resistance.