Migraine is a common neurological disease. Acupuncture has been proven to be effective but the mechanisms remain unclear. A proton magnetic resonance spectroscopy imaging (MRSI) study was used to investigate biochemical changes in brain regions key for the transmission of pain in response to acupuncture treatment. Results showed acupuncture treatment was associated with a significantly increased NAA/Cr in bilateral thalamus in migraine patients. A strong significant correlation between NAA/Cr and headache intensity was found in thalami. Our data provided the first evidence suggesting a brain biochemical change in response to acupuncture therapy in migraine, in correlation with clinical outcomes.