The measurement of steady state level and accumulation rate of GABA and the activities of GAD and GABA-T in brain regions of adult male albino rats treated with diazepam (diazepam, 5 mg/kg/day, i.p.) and/or exposed (2 h/day) to higher environmental temperatures (higher environmental temperature, 40°C) under long-term conditions (7-30 consecutive days) shows that long-term treatment with diazepam (5 mg/kg/day, i.p.,for 15-30 consecutive days) reduced the GABAergic activity in the hypothalamus, corpus striatum (CS) and cerebellum (CM) without any alteration in the cerebrocortical region (CC). Multiple exposures to higher environmental temperatures for 7 consecutive days, on the other hand, enhanced GABAergic activity in hypothalamus, corpus striatum and cerebrocortical region of the rat. This higher environmental temperature-induced increase in GABAergic activity was further enhanced in the hypothalamus, but it was attenuated in the corpus striatum and became normalized in the cerebrocortical region upon the prolongation of exposure (15-30 consecutive days) to higher environmental temperature. The long-term diazepam-induced inhibition in GABAergic activity in hypothalamus and corpus striatum (but not in the cerebellum) disappeared and was finally activated following exposures (2 h/day) to higher environmental temperatures (40°C) along with diazepam-treatment (5 mg/kg day, i.p.)for 7, 15 and 30 consecutive days. In the cerebellum, multiple exposures to higher environmental temperature did not significantly alter the long-term diazepam-induced inhibition of GABAergic activity or GABAergic activity in the normal rat. Multiple higher environmental temperature-induced characteristic changes in the GABA system of the cerebrocortical region was not significantly altered when diazepam was administered along with long-term higher environmental temperature exposures. Thus, these results suggest that the changes in brain regional GABAergic activity with multiple higher environmental temperature exposures and/or with long-term diazepam treatment are specific to brain regions and dependent on the conditions of the treatment/exposure.