This study investigated the effects of neuroactive steroids, which have been reported to modulate GABA-ergic transmission, on the secretion of somatostatin (SRIH) and also dopamine (DA) from primary rat hypothalamic cell cultures, where the release of both substances is regulated by a GABAA receptor-mediated inhibitory tone. Pregnenolone sulphate (PS), a negative allosteric modulator at the GABAA receptor, enhanced SRIH secretion in a time and dose-dependent manner (10−12–10−8 M). This effect was reversed by muscimol (10−8 M) and enhanced by bicuculline (10−6 M), thus supporting an action of PS at the GABAA receptor. The release of endogenously synthesized dopamine (DA) was, however, unaffected by PS. A number of other steroids were also tested for their potential actions on SRIH and DA secretion. Allopregnanolone had slight but significant stimulatory actions on SRIH secretion, whereas tetrahydro-deoxycorticosterone (TH-DOC) markedly stimulated SRIH secretion with a bell-shaped dose response curve resembling that found for PS. The release of DA was unaffected by these neuroactive steroids but, unlike SRIH, DA release was stimulated by dehydroepiandrosterone sulphate (DHEAS). The results support the view that neuroactive steroids may play an important role in regulating some aspects of neuroendocrine function and they also provide the first demonstration of differential activities of neuroactive steroids within the hypothalamus at low, physiologically relevant concentrations. The results also raise the possibility that certain hypothalamic neuronal populations may posses uniquely different GABAA receptors and that such mechanisms may contribute to the functional development of the neuroendocrine system.