RESUMO
Parameters of electrofusion of 2-cell mouse embryos were optimized for application as a model for nuclear transplantation. There was considerable lysis of embryos with M(2) as the medium for fusion; however, 100% fusion (n = 58) was obtained with a single 0.31-kv / cm, 1280-musec pulse. With mannitol and sucrose solutions as the medium, a wide range of field strengths (0.31-1.41 kv / cm for 0.26 M sucrose solution and 0.31 to 2.04 kv / cm for 0.3 M mannitol solution) and durations of the electrical pulse (10-1280 musec) resulted in high rates of fusion (often 100%). Likewise, osmolarity of sucrose and mannitol solutions did not affect the rate of fusion using a 0.47-kv / cm pulse. With a field strength of 2.04 kv / cm, the proportion of embryos that fused in mannitol solution increased (P<0.05) and the proportion that were lysed decreased (P<0.05) as osmolarity increased. Both fused (162 642 , 25%) and control embryos (32 72 , 44%) continued to develop in culture for 48 h, after which they began to compact. Fused embryos were only at the 4-cell stage by this time, while control embryos were at the 8-cell stage. Optimal pulse durations are plotted for field strengths between 0.31 and 1.41 kv / cm with 0.26 M sucrose as fusion medium.
RESUMO
The objective of this study was to examine the effects of different culture systems on the development of early bovine embryos in vitro. A total of 1089 oocytes were aspirated from 2 to 5 mm follicles of ovaries collected at a local abattoir; a high proportion of the oocytes matured in vitro were fertilised by spermatozoa capacitated with caffeine and heparin. Seven to eight hours after insemination, the oocytes were transferred into three in vitro systems: A, TCM 199 + 10 per cent fetal calf serum culture medium, B, coculture with a monolayer of granulosa cells and C, coculture with bovine oviductal epithelial cells. The results showed that the proportion of the early bovine embryos which overcame the block at eight to 16 cells and developed to the morula and blastocyst stages in system C was significantly higher than in systems A or B.