Recent studies have provided new data on the neuroendocrine role of glutamate (the major excitatory neurotransmitter) on somatostatin release. The neuroendocrine role of gamma-aminobutyric acid (GABA) (the major inhibitory neurotransmitter) on this same secretion, is also well established. Our objective was thus to investigate whether GABA and glutamate, which have opposite neurotransmission signals, could interact in the control of hypothalamic somatostatin release. Pharmacological manipulations of the two types of receptors were performed in vitro on primary cultures of hypothalamic neurons secreting somatostatin. We found that tonic release of somatostatin was reduced by 76% in the presence of tetrodotoxin (TTX) and was regulated by endogenous secretion of glutamate and GABA. CGS 19755, a highly selective N-methyl-D-aspartate (NMDA) receptor antagonist, significantly reduced tonic somatostatin secretion whereas it was strongly increased by picrotoxin and bicuculline, two GABAA antagonists. When CGS 19755 was applied with picrotoxin, somatostatin release was the same as levels obtained in the control group with TTX. GABA reduced tonic somatostatin release (in the presence or absence of TTX), and glutamate-stimulated secretion in a dose-dependent manner. Picrotoxin stimulation of tonic somatostatin release was additive with that obtained after glutamate stimulation and was also dose-dependent. This interaction was also studied in vivo in unanesthetized rats bearing a push-pull cannula stereotaxically implanted into the median eminence. Ip injected CGS 19755 (an antagonist that can freely permeate the blood-brain barrier) completely blocked the peak secretion of somatostatin observed after ip picrotoxin administration, whereas there was no significant effect when it was injected alone. These findings corroborated our in vitro data and allow us to postulate that GABA and glutamate interact in the control of somatostatin.