Objective To investigate the effect of exercise preconditioning on serum level of tumor necrosis factor α (TNF-α), interleukin (IL) -1β and IL-6 in rats after cerebral ischemia-reperfusion (I/R). Methods 24 male Sprague-Dawley rats were randomly divided into exercise preconditioning group (n=8), model group (n=8) and sham group (n=8). The middle cerebral arteries were occluded for 120 min and re-perfused. All the rats were evaluated with neurological deficit score 2 hours, 24 hours after I/R. The serum levels of TNF-α, IL-1β and IL-6 was detected with enzyme-linked immunosorbent, and the pathology was observed with HE staining 24 hours after I/R. Results The neurological deficit score decreased in the exercise preconditioning group compared with that in the model group, as well as the serum TNF-α, IL-1β and IL-6 (P<0.05) 24 hours after I/R. The pathological damage and interstitial edema alleviated in cerebral ischemia cortical, and the degeneration and necrosis of the neurons in the ischemic area significantly reduced in the exercise preconditioning group. Conclusion Exercise preconditioning may inhibit inflammatory response in I/R rats to protect neurological function from impairment.