The aim of this study was to examine whether increased frequency of luteinizing hormone (LH) pulses influences luteal progesterone (P4) secretion by measuring progesterone concentrations at the secreted (caudal vena cava) and circulating levels (jugular vein) in lactating dairy cows. Cows received six intravenous administrations of 2.5 μg of GnRH (gonadorelin acetate, n=4) or 2 ml saline (n=3) at 1-h intervals on 12.4 ± 0.4 (mean ± SE) days after ovulation. Blood samples were collected from the caudal vena cava and jugular vein every 12 min for 12 h (6 h before and after treatment). During the 6 h after treatment, frequency of LH pulses (5.3 ± 0.3 and 3.0 ± 0.0 pulses/6 h) and mean LH concentration (0.50 ± 0.06 and 0.38 ± 0.05 ng/ml) were greater (P<0.05) in GnRH-treated cows than in saline-treated cows. Mean P4 concentration and amplitude of P4 pulses in the caudal vena cava during the 6 h after treatment were greater (P<0.05) in GnRH-treated cows than in saline-treated cows, but the frequency of P4 pulses was not different between the groups. Mean P4 concentration in the jugular vein during the 6 h after treatment was also higher (P<0.05) in GnRH-treated cows than in saline-treated cows (7.0 ± 1.3 and 5.4 ± 0.9 ng/ml). These results indicate that the increased frequency of LH pulses stimulates progesterone secretion from the functional corpus luteum and brings about higher P4 concentrations in the circulating blood in lactating dairy cows.