Various types of hypothalamic roof deafferentation (RD) (i.e. large anterior (LARD), large posterior (LPRD), small anterior (SARD) or middle (MRD)), electrolytic lesions of the paraventricular nucleus (PVN) or intracerebroventricular (i.c.v.) injection of alpha-helical corticotrophin-releasing factor(9-41) (alpha-helical CRF(9-41)) were performed in ovariectomized lactating rats in order to determine the afferent pathway of the suckling stimulus and whether the PVN and corticotrophin-releasing hormone (CRH) are involved in suppressing pulsatile LH secretion during lactation. Animals were ovariectomized on day 2 of lactation (day 0 = the day of parturition). Deafferentations and electrolytic lesions were made on day 7. On the following day, blood samples were taken via an indwelling atrial cannula every 6 min for 3 h. Pulsatile LH secretion with high frequency and amplitude was present in rats with LARD or LPRD despite the suckling. In rats with MRD, LH pulses with a small amplitude were observed when the cut was on or under the ventral margin of the PVN, but there were few LH pulses when the cut passed through the PVN. Electrolytic lesioning of the PVN, however, did not affect the suppression of pulsatile LH secretion during lactation. In addition, i.c.v. injection of alpha-helical CRF(9-41) (26.1 nmol/10 microliters) into the third ventricle on day 8 of lactation did not reverse the suppression of LH secretion by the suckling stimulus. These results suggest that the pathway associated with this inhibition may be rather diffuse and that the PVN region and CRH are not critical in conveying the inhibitory inputs of the suckling stimulus.