Endothelin-1 is a 21 amino acid peptide originally isolated from porcine aortic endothelium and has recently been localized within the central nervous system. We have administered endothelin-1 in a dynamic perfusion system in order to study its possible effects on the rat hypothalamus and anterior pituitary. Tissue (hypothalami or quartered pituitaries) was placed into plastic chambers and was perfused with oxygenated Krebs-bicarbonate solution. After an interval to establish stable basal peptide release, endothelin-1 was administered at two doses (0.1 and 1 μM) and the release of substance P, vasoactive intestinal peptide, 7B2, and somatostatin was meas-ured, the last being detectable only in hypothalamic perfusates. Both concentrations of endothelin-1 led to a significant increase (P < 0.01) in the release of substance P from the hypothalamus and pituitary, but not of vasoactive intestinal peptide, 7B2, or somatostatin. Thus after the 0.1 μM and 1 μM endothelin-1 perfusion substance P release from the hypothalamus increased by 125 ± 5% and 215 ± 15% (mean ± SEM) of basal and from the pituitary by 168 ± 8% and 276 ± 15% (mean ± SEM). No change occurred in the output of ACTH or other pituitary hormones. The release of substance P from hypothalamus or pituitary after stimulation with endothelin-1 was not blocked when a calcium free medium was used. Endothelin-1 binding sites were identified on rat pituitary cell membranes. These findings suggest the possibility that endothelin may act as a paracrine substance, neurotransmitter, or neuromodulator in the hypothalamo-pituitary axis.