The current study aimed to investigate the effect of melatonin on liver function in rats exposed to gamma radiation. Male Wister rats were divided into 10 groups (n=5) as follows: Control group, Sham group received the melatonin solvent, two melatonin groups received melatonin for 3 and 14 days, two irradiated groups subjected to whole body gamma radiation and assed after 3 and 14 days, two treatment groups received radiation and melatonin. Hepatic injury was convinced in rats via single dose irradiation of 4 Gy gamma radiation. Irradiation was performed using Cesium-137 at a dose rate of 0.695 rad/ sec. Melatonin was administered by gavage at dose of 25 mg/Kg 30 min before irradiation. After 3 and 14 days post co-treatment, rats were euthanized and blood sample was obtained. Results showed significant decrease in body weight if irradiated rats. The degree of liver dysfunction was assessed using determination of liver function biomarkers in serum. Results showed that exposure to γ-Irradiation caused a significant increase in the activities of serum ALT, and AST, and glucose levels. Exposure to γ-radiation of 4 Gy caused a significant decrease in the content of serum total proteins, albumin and globulin with increase in the albumin/globulin ratio. Liver injury following melatonin treatment showed remarkable improvement reflected by normalization of liver function biomarkers in serum and increased body weight. In conclusion, current study showed that treatment with melatonin following radiation exposure protects rats against liver dysfunction, which might have therapeutic role for radiation induced liver injury and dysfunction.