To investigate the effect of thiopental sodium on the release of glutamate and γ-aminobutyric acid (GABA) from synaptosomes in the prefrontal cortex, synaptosomes were made, the spontaneous release and the evoked release by 30 mmol/L KCl or 20 μmol/L veratridine of glutamate and GABA were performed under various concentrations of thiopental sodium (10-300μmol/L), glutamate and GABA concentrations were determined by reversed-phase high-performance liquid chromatography. Our results showed that spontaneous release and evoked release of glutamate were significantly inhibited by 30μmol/L, 100 μmol/L and 300 μmol/L thiopental sodium, IC50 of thiopental sodium was 25.8±2.3 μmol/L for the spontaneous release, 23.4±2.4 μmol/L for KClevoked release, and 24.3±1.8 μmol/L for veratridine-evoked release. But GABA spontaneous release and evoked release were unaffected. The study showed that thiopental sodium with clinically related concentrations could inhibit the release of glutamate, but had no effect on the release of GABA from rats prefrontal cortical synaptosomes.