Abstract Objective Though observation the changes of Caspase-3 protein and reactive oxygen species in the forebrain of mice induced by isoflurane inhalation to understand the role of reactive oxygen species in neural damages of isoflurane. Method Fifty-six male C57BL/6J mice were randomly divided into isoflurane treatment (Iso, n=20), dimethylthiourea treatment (DMTU, n=8), imethylthiourea plus isoflurane treatment (DMTU+Iso, n=8), and control (Con，n=20) groups. Mice in Iso and DMTU+Iso groups were exposed to three sessions of 1.4% isoflurane, 2 h for each session. Thirty min before oxygen inhalation in DMTU group and first isoflurane inhalation in DMTU+Iso group, 50 mg/kg of radical scavenger DMTU were intraperitoneally injected to the mice in the two groups. The changes of caspase-3 expression in prefrontal cortex were observed by immunocytochemical staining. And the activities of superoxide dismutase (SOD) and the amount of MDA were detected. Result Compared with the Con group, the number of caspase-3 positive cells in Iso group was increased significantly (3.38 times, P<0.05), but DMTU+Iso (1.43 times of Con group) and DMTU group (0.74 times of Con group) were not obviously changed (P>0.05). There was no significant difference in the activities of SOD between the Iso (25±10) and Con groups(20±6, P>0.05), but lipid peroxidation malondialdehyde (MDA) content in Iso group (13.7±6.1) was significantly higher than that in Con group (5.7±2.7, P<0.05). Conclusion 1.4% isoflurane exposure significantly increase the expression of caspase-3 in the forebrain of mice and its molecular mechanism is related to the increasing of free radicals.