Effects of follicular cells on oocyte maturation .1. Effects of follicular hemisections on bovine oocyte maturation in vitro

This study was designed to evaluate the role of follicular cells in the maintenance of meiotic arrest (germinal vesicle [GV] stage). Bovine ovaries were obtained from a slaughterhouse. Ten oocytes were selected and cocultured for 24 h with two follicular hemisections in a 50-mu l drop of TCM-199 supplemented with 10% fetal calf serum. Follicular hemisections were separated into cellular components-granulosa, theca interna, and theca externa. When the oocytes were cocultured in physical contact with granulosa cells, 36% were maintained in the GV stage. However, the percentage of oocytes in the GV stage was significantly increased (p < 0.05) when the theca interna layer associated with (86%) or not with (78%) granulosa cells was used rather than the hemisection consisting of all three follicular cell layers (57%). When we cocultured oocytes without direct contact, using theca interna associated with granulosa cells, only 35% of the oocytes were maintained in the GV stage. Conditioned medium obtained after culture with two follicular hemisections also maintained a high percentage of oocytes in the GV stage when it was replenished every 4 h. An additional culture of 12 h with fresh unconditioned medium resulted in 95% meiotic resumption. In conclusion, these results suggest that the maintenance of bovine oocytes in the GV stage is influenced by theca cells, independent of contact, and that the oocytes can be maintained by conditioned medium from follicular hemisections.