Aim To study the protection mechanism of mild hypothermia on cerebral ischemia-reperfusion(I/R) injury. Methods 36 male SD rats were randomly divided into sham operation group, recombinant tissue plasminogen activator(rt-PA) group, model group, I/R+mild hypothermia group, I/R+ rt-PA group, and I/R+mild hypothermia+rt-PA group, with 6 rats in each group. By building the rat model of ischemia reperfusion, the semi quantitative neurological deficit score was made. By the experiment of blood brain barrier penetration, the difference of blood-brain barrier protection was compared among every groups. Immunohistochemistry was used to detect the expressive level of t-PA and PAI-1 in the tissues of rats; Western blot was used to test the expressive level of p-MEK1/2 and p-ERK1/2. Results Neural injury was lower in two groups of mild hypothermic rats than that in experimental control group(P<0.05). Mild hypothermic treatment can reduce the permeability of the blood-brain barrier. The expression of t-PA and PAI-1 in rat brain tissue significantly increased in rt-PA group and I/R+rt-PA group, while the expression of other groups was lower. The p-MEK1/2 and p-ERK1/2 expressions were significantly increased in the rt-PA group and the I/R+rt-PA group, which were significantly reduced after mild hypothermic treatment. Conclusion The protective effect of mild hypothermia on cerebral ischemia reperfusion injury is related to reducing the permeability of blood-brain barrier, reducing the expression of t-PA and PAI-1 after ischemia reperfusion in rat brain tissue, and reducing the expression of p-MEK1/2 and p-ERK1/2.