We developed a method for the determination of γ-aminobutyric acid (GABA) in brain tissue using a gas-liquid chromatograph equipped with a flame ionization detector. This method is based on the formation of a GABA derivative by treatment with isobutyloxycarbonyl chloride and diazomethane, under mild conditions. This method proved to be useful for the determination of GABA in the whole mouse brain. We thus studied the effect of nitrazepam, one of the benzodiazepines (BDZs), on the GABA levels in the mouse brain and found that the concentration of GABA in the nitrazepam treated group was decreased markedly compared with values in the untreated controls.