This experiment was conducted to investigate the effects of different exogenous progesterone sources administrated after artificial insemination (AI) on serum progesterone (P4) concentration and pregnancy rates in Holstein lactating cows. Sixty-four lactating Holstein dairy cows were allocated to four different treatments (n = 16 per treatment): the cows 1) were injected with physiological saline on days 5 and 13 after AI (control group); 2) were injected with progesterone on days 5 and 13 after AI (P group); 3) received controlled internal drug releasing device (CIDR) for a period from day 5 to 19 after AI (CIDR group); and 4) were injected with gonadotropin-releasing hormone (GnRH) agonist on days 5 and 13 after AI (GnRH group). Blood samples were collected on days 0 (AI day), 5, 13, 16 and 19 after AI to determine serum P4 concentration. The results revealed a significant difference among treatment groups for serum P4 concentration on days 13, 16 and 19 with the lowest concentration of serum P4 for the control group. The pregnancy rate was also positively affected by all the treatments with CIDR having the greatest effect on pregnancy rate. Overall, the results indicated that CIDR has the greatest effect on serum P4 concentration and pregnancy rate, although the administration of P and GnRH during days after AI increased serum P4 concentration in lactating dairy cows as well.