| Popis: |
The incidence of polyspermy in lamb oocytes matured and fertilized in vitro is very high and this results in a reduced developmental potential of embryos arising from them. We have attempted to produce oocytes more resistant to this fertilization anomaly. The oocytes from prepubertal lambs 7-12 weeks old were matured in a medium supplemented with various blood sera and oviductal fluid and fertilized in vitro. Significantly higher monospermic penetration was found in a medium supplemented with BSA--3 mg/ml (63.9%) and OF--20% concentration (55.8%). Lower monospermy was recorded in the presence of 10% LS (44.6%) or 10% SS (40.8%), and particularly in a medium with 10% FCS (26.9%). In contrast, high monospermy (78.7%) was observed in oocytes from adult donors matured and fertilized in an identical system. In another set of experiments we estimated whether polyspermy can be reduced by improvement of the cytoplasmic maturation of prepubertal oocytes using a two-step maturation protocol. After artificial arrest of the maturation for 24 h with a specific cdk inhibitor--BL-I, 50 miocroM--more than 80% oocytes from prepubertal and adult donors did not resume meiosis. When incubated thereafter in a drug-free medium for another 24 h, the oocytes of both categories progressed to MII in the rate comparable with control (80% to 90% MII). However, after fertilization no significant differences in the level of monospermic penetration was recorded between the arrested group (59.8%) and control (58.8%), both matured in the presence BSA, and 46.6% and 52.3% after treatment with OF. Also, no significant difference was observed between the arrested and control oocytes from adult donors (72.6% and 84.8%, respectively). These results suggest that high polyspermy in prepubertal oocytes is caused by developmental imperfection and can't be fully eliminated either by modifying the composition of culture media or by prolongation of the culture interval. |
