Scorpion has strong analgesic effects, but its analgesic mechanisms remain unclear. This study investigated the effects of scorpion powder on enkephalin expression in the midbrain of rats with nitroglycerin-induced migraine at mRNA and protein levels. Results confirmed that migraine rat abnormal behavior was significantly improved, and proenkephalin mRNA expression was significantly increased following treatment with scorpion. The number of methionine-enkephalin- positive cells in the migraine rats following treatment with scorpion was significantly increased, but no significant difference in the number of leucine-enkephalin-positive cells was detectable compared with migraine and normal rats. Taken together, these results show that scorpion exerts potentially beneficial effects by promoting enkephalin expression in nitroglycerin-induced migraine rats.