We have previously demonstrated that the inhibition of neurotransmitter uptake at dopamine (DA) terminals stimulates the hypothalamo-pituitary-adrenal axis. In the present study we investigated the role of central DA neuronal systems in the regulation of this axis. Administration of the DA uptake inhibitor GBR12909 (3-30 micrograms) into the third ventricle dose-dependently elevated serum adrenocorticotropin hormone (ACTH) levels in rats. GBR12909 (10 micrograms) elevated serum ACTH levels following administration into the paraventricular nucleus of the hypothalamus but not into the lateral ventricle. The administration of 6-OHDA into the third ventricle significantly decreased DA content in the hypothalamus and striatum and significantly attenuated the ACTH response to GBR12909 (10 micrograms, third ventricle or 10 mg/kg, i.p.). Conversely, 6-OHDA lesions of the medial forebrain bundle, which depleted 99% of DA in the caudate but did not decrease DA content in the hypothalamus, and did not attenuate the ACTH response to i.p. GBR12909. Measurement of GBR12909-induced inhibition of [3H]DA uptake into synaptosomal preparations indicates the presence of GBR12909-sensitive DA transporters in the region of the hypothalamus surrounding the third ventricle. The present findings suggest that an endogenous DA neuronal system terminating in the hypothalamus mediates the effects of stimulants on hypothalamo-pituitary-adrenal function and might play a role in the ongoing regulation of hypothalamo-pituitary-adrenal activity.