Effects of follicular cells on oocyte maturation .2. Theca cell inhibition of bovine oocyte maturation in vitro

This study was undertaken to assess the role of follicular cells in the maintenance of meiotic arrest (germinal vesicle [GV] stage). Bovine cumulus-oocyte complexes (COG) were obtained by puncture of ovaries collected at a slaughterhouse. Different monolayers of follicular cells-granulosa cells, theca interna, theca externa, or both types of theca cells together-were cultured in 24-well plates with 1 ml of TCM-199 supplemented with 10% fetal calf serum. Theca cells were obtained by digesting theca layers with collagenase. The medium was renewed 48 h before coculturing selected COC with confluent follicular cell monolayers. Oocytes were maintained in GV stage when cocultured for 12 h directly on monolayers of theca cells. The percentage of oocytes in GV stage was not significantly different between treatments using different types of theca cells (51-66%). Whether COC were cocultured in contact or not in contact with theca cells, the percentage of GV stage was similar (61%). The reversibility of this inhibition was high (85%). However, granulosa cells did not exert meiotic arrest (10%). When oocytes were denuded of their cumulus cells and cocultured with theca cells, only 3% were maintained in GV stage; 59% were maintained in GV stage when COC were not removed before culture. This data provided evidence of the essential role of cumulus cells in maintaining GV stage. In conclusion, we have demonstrated that theca cells, in vitro, maintained bovine oocytes in meiotic arrest. The inhibitory factor(s) produced by theca cells is soluble in the medium and acts through the cumulus cells.