Intraperitoneal injection of diazepam in moderate dosage (1--10mg/kg) to rats caused a decrease in dopa and 5-hydroxytryptophan (5-HTP) formation, measured as the accumulation of these intermediates induced by inhibition of the aromatic L-aminoacid decarboxylase by means of NSD 1015 (3-hydroxybenzylhydrazine (HCl), in limbic forebrain, striatum and the remaining hemisphere portion. These effects are opposite to those induced by gamma-aminobutyric acid (gaba) and gamma-butyrolactone (100 and 750 mg/kg i.p. respectively), and the effects of the latter agents were significantly counteracted by diazepam. The effect of diazepam on dopa formation persisted after the acute transection of dopaminergic axons (transverse cerebral hemisection at the level of the caudal hypothalamus). The elevation of dopamine following hemisection was also significantly counteracted on the hemisected side of the brain, the intact side remaining unchanged. The data do not support the hypothesis that benzodiazepines act by enhancing gabaergic transmission. They rather suggest that these agents exert an inhibitory action on transmitter synthesis and utilization at the synaptic level, i.e. an action not necessarily bearing any direct relationship to gaba.