Vasoactive intestinal peptide (VIP) is a neuropeptide that is present in the hypothalamus and is probably a neuroen-docrine regulator. The effect of VIP on pulsatile LH secretion in the long-term ovariectomized rat was re-examined in the light of earlier conflicting reports. VIP or saline was infused into the third ventricle at the rat of 15 µl/h and blood was sampled frequently before and during the infusion. VIP at 3.5 nmol/h significantly depressed mean LH levels (p < 0.05) and lowered pulse frequency (p < 0.05), but had no effect on LH pulse amplitude (p > 0.05). VIP at lower levels was not consistently effective, and intraventricular saline was without influence. We examined indirectly whether the site of action of VIP (3.5 nmol/h) was the brain or pituitary by injecting various doses of gonadotropin-releasing hormone (GnRH; 0.5–4.0 ng/100 g BW i.v.) during VlP-induced inhibition of LH secretion and in saline-infused controls. VIP did not alter the response of the pituitary to GnRH or the slope of the GnRH-LH dose-response curve (p > 0.05). We conclude that the inhibitory action of VIP on pulsatile LH secretion is probably exerted in the hypothalamus. To test the hypothesis that dopamine mediates the inhibitory effects of VIP (3.5 nmol/h), animals were pretreated with the dopamine receptor blocking agent pimozide (1.26 mg/kg) in an attempt to block the actions of VIP. Pimozide did not affect the response of LH to VΙP infusion (p > 0.05). We conclude that dopamine is not a likely mediator of the action of VIP.