Abstract. The present study examined the effects of 17β-estradiol (E2) on in vitro maturation and subsequent in vitro fertilization of pig oocytes matured with or without cumulus cells. When E2 (10 ng/ml) was added to the protein-free maturation medium, the proportions of cumulus-enclosed oocytes that underwent germinal vesicle breakdown and reached metaphase II were significantly reduced (P<0.05), and cumulus expansion was also significantly inhibited (P<0.05) compared with the control (no E2 added). Although oocytes matured in the presence of E2 were penetrated by sperm in vitro at the same level as the control, the incidences of male pronuclear (MPN) formation and activated oocytes were significantly lower (P<0.05) than the control. These inhibitory effects of E2 were prevented when the medium was supplemented with E2 together with its antagonist, ICI 182,780 (1 µg/ml), although the presence of the antagonist alone in the medium had no effect on the maturation and fertilization in vitro of oocytes. In cumulus-free oocytes, E2 had no effect on nuclear maturation and penetration in vitro, but low MPN formation was observed in oocytes matured in the presence and absence of E2. When cumulus-enclosed oocytes were cultured in the presence of progesterone (P4; 600 ng/ml) alone or together with E2, no significant differences in nuclear maturation, cumulus expansion or penetration in vitro were observed compared with control oocytes. The concentration of P4 in maturation medium was significantly (P<0.01) lower when cumulus-enclosed oocytes were cultured for 44 h in the medium with E2 than in medium without E2. These results indicate that E2 inhibits both nuclear and cytoplasmic maturation of cumulus-enclosed pig oocytes, and that this inhibition can be prevented by an E2 antagonist or P4. This E2 inhibition may occur indirectly via the cumulus cells and inhibition of P4 synthesis. Key words: 17β-estradiol, In vitro fertilization, In vitro maturation, Oocyte, Pig (J. Reprod. Dev. 50: [305][306][307][308][309][310][311][312][313] 2004) n mammalian follicles, oocytes are surrounded by follicular cells in antral follicles and grow and mature in an environment of follicular fluid. During follicular development, the concentration of estrogen in follicular fluid gradually increases due to stimulation by follicle stimulating hormone (FSH) [1][2][3][4][5]. However, after the ovulatory lutenizing hormone (LH) surge or treatment with human chorionic gonadotrophin (hCG), the concentration of estrogen decreases drastically, and an increase in the concentration of progesterone follows, shortly before ovulation [1][2][3][4][5]. In addition to these changes in the concentrations of estrogen and progesterone in follicular fluid, the LH surge also stimulates meiotic resumption in oocytes. Therefore, it is possible that these steroids play an important role(s) not only for nuclear maturation