In vitro growth and maturation of pig oocytes

Preantral follicles containing oocytes of 70-89.5 microns in diameter were isolated from pig ovaries and cultured in collagen gel for up to 16 days, in the presence of serum, FSH and oestradiol. Formation of follicular antra occurred as the culture proceeded. The oocytes had been enclosed by granulosa cells and contacts between the oocytes and processes of the enclosing cumulus cells were maintained over the culture period. After 16 days of culture, 30-40% of the oocytes were of normal appearance, and the diameters of about half of these oocytes were larger than 100 microns. When the oocytes grown in vitro were liberated from the follicles and cultured for a further 48 h in modified Krebs-Ringer bicarbonate solution, 6, 30 and 60% of the oocytes larger than 90, 100 and 110 microns underwent germinal vesicle breakdown, respectively. Progression to metaphase II was observed in 40% of oocytes that were over 110 microns in diameter, whereas no oocyte less than 90 microns in diameter resumed meiosis. The relationship between the size and meiotic competence of oocytes was similar for oocytes grown in vitro or in vivo. Oocytes grown and matured in vitro were penetrated by spermatozoa and formed a female pronucleus, but decondensation of the sperm head was incomplete. The results demonstrate for the first time that pig oocytes from preantral follicles can grow up to their final size, acquire meiotic competence, and be penetrated by spermatozoa in vitro.