THE CEREBROPROTECTIVE EFFECTS of mild hypothermia have been extensively studied in various animal models of ischemia, but the mechanism by which mild hypothermia diminishes ischemic injury is not well understood. Nitric oxide (NO) has been implicated as a mediator of glutamate excitotoxicity in primary neuronal cultures, and its synthesis is acutely increased during focal ischemia in vivo. To evaluate possible mechanisms of hypothermic neuroprotection, we measured markers of NO synthesis–nitrite and cyclic guanosine monophosphate (cGMP) levels and NO synthase activity–during right middle cerebral artery occlusion (MCAO) in the rat under normothermic (36.5°C) and mild hypothermic (33°C) conditions. There was a significant increase in nitrite concentration in the right hemisphere versus the left under normothermic conditions at 10 and 20 minutes after MCAO (P< 0.01), with a return to baseline levels by 60 minutes. The increase in cortical nitrite levels in the right hemisphere versus the left was not observed with mild hypothermia. There was a threefold increase in cGMP synthesis in the normothermic right cortex 10 minutes after MCAO (P< 0.05). This rise in cGMP did not occur in hypothermic animals, and the right to left cortical disparity in cGMP production was abolished. Finally, the significant increase in NO synthase activity seen in the normothermic ischemic cortex was absent in hypothermic rats (P< 0.05). These results suggest that mild hypothermia (33°C) modulates the burst of nitric oxide synthesis during cerebral ischemia and may account, at least partially, for its cerebroprotective effects.