Previous studies have shown that the endogenous opioid peptides, acting at specific opiate receptor subtypes, are involved in the suckling-induced prolactin secretory response. The prolactin increase elicited by suckling is due, at least in part, to an inhibition of tuberoinfundibular dopaminergic (TIDA) neurons in the hypothalamus. We investigated the effects of immunoneutralization of dynorphin, leu-enkephalin and met-enkephalin on the suckling-induced prolactin increase and on the activity of the TIDA neurons in lactating female rats between days 7 and 12 postpartum. Rats were injected into the right lateral ventricle with antiserum specific for one of these three peptides. Control rats were administered equal amounts of immunoglobulin proteins. Suckling produced a profound and significant increase in prolactin levels, as well as a decrease in DOPA accumulation in the median eminence of lactating rats. Administration of immunoglobulin concentrations of up to 3.6 µg did not inhibit the prolactin secretory response to the suckling stimulus and did not prevent the suckling-induced inhibition of TIDA neurons. Antisera to all three endogenous opioid peptides abolished the suckling-induced prolactin increase and prevented the inhibition in DOPA accumulation in the median eminence. Thus, the endogenous opioid peptides, dynorphin, leu-enkephalin and met-enkephalin, are essential for the prolactin secretory response to suckling and inhibition of TIDA neuronal activity is at least one of the mechanisms of action utilized by these peptides.