A study was conducted to determine the effect of the milk-ejection reflex on exogenous gonadotropin releasing hormone (GnRH)-induced release of luteinizing hormone (LH) after short-term calf removal. Twenty-four postpartum multiparous beef cows were assigned randomly to groups arranged in a 2(3) factorial arrangement. Factors consisted of two levels of suckling [suckled (S) or nonsuckled (NS)], treatment with GnRH [saline (C) or 200 micrograms GnRH] and days postpartum (d 1 and 14). Dams were isolated from their calves for 4 h on d 1 and 14 postpartum. At the end of 4 h dams were reunited with their calves in S + C and S + GnRH groups, while dams of calves in NS + C and NS + GnRH groups remained separated an additional 2 h. Cows were injected iv with saline or GnRH following the 4-h isolation period, 5 min after calves had begun suckling or nuzzling the udder. Sera from jugular blood samples collected 15 min prior to the end of the 4-h isolation period, immediately prior to injection (0 h) and at 15 min intervals thereafter for 120 min were analyzed for LH. Serum concentrations of LH in control cows did not differ due to suckling or stage of the postpartum period and averaged 2.3 +/- .1 ng/ml. Pituitary response to GnRH was determined by computing the rate of LH release. Rate of LH release (ng LH.ml-1.min-1) in response to GnRH on d 14 was greater (P less than .001) than on d 1 in both suckled and nonsuckled groups (S + GnRH, 37.1 +/- 3.9 vs 18.3 +/- 5.0; NS + GnRH, 34.7 +/- 5.9 vs 14.5 +/- 1.1). However, GnRH-induced release of LH did not differ between suckled and nonsuckled cows on either d 1 or 14 postpartum. These data indicate that response of the bovine pituitary to GnRH during the postpartum period is not influenced by the act of suckling but is enhanced with time after parturition.