Leal C L, Liu L
Department of Development and Signaling, The Babraham Institute, Cambridge, UK.
Anim Reprod Sci. 1998 Jun 30;52(1):51-61. doi: 10.1016/s0378-4320(98)00084-0.
The 6-DMAP (6-dimethylaminopurine) has been reported to accelerate and enhance the formation of pronuclei (activation) in non-aged MII mouse and bovine eggs. In this study, effects of chemical activation (CA) were evaluated using ethanol + 6-DMAP and electrical activation (EA) on pig oocytes matured for 36 h (newly matured) and 48 h. After CA, the first pronuclei developed within 2 h, with maximal pronuclear formation at 8 h (> 90%) for oocytes matured for either 36 or 48 h. In addition, chemically activated oocytes did not extrude the second polar body. After EA treatment, pronuclear formation began at 4-6 h and was significantly lower at 8 h in newly matured than aged oocytes (34% vs. 85%). Cleavage rate at 24 h after treatment was lower for oocytes treated by CA (< 50%) than for EA (> 70%) and after 3 days of in vitro culture, fewer oocytes reached the four-cell stage in CA than in EA groups. No chemically activated oocytes developed beyond the four-cell stage; in contrast, EA resulted in development beyond this stage. Both CA and EA resulted in a drop of H1 kinase activity, but EA induced a more rapid reduction. With EA, H1 kinase activity rapidly declined but tended to increase again at 8 h in oocytes matured for 36 h, which was different from all other groups in which the kinase activity remained low at 8 h post-activation. We conclude that different mechanisms are involved in pronuclear formation with CA and EA and that 6-DMAP-activated oocytes may not be suitable for the induction of normal parthenogenetic development in pigs. It is also suggested that the ability of pig oocytes to become fully activated may be related to the modification of H1 kinase activity during the aging process. These modifications to H1 kinase may be necessary for parthenogenetic activation of in vitro matured oocytes and also for the completion of meiosis and cleavage.
据报道，6-二甲基氨基嘌呤（6-DMAP）可加速并增强未老化的MII期小鼠和牛卵原核的形成（激活）。在本研究中，使用乙醇+6-DMAP化学激活（CA）和电激活（EA）评估了对成熟36小时（新成熟）和48小时的猪卵母细胞的影响。CA处理后，对于成熟36小时或48小时的卵母细胞，第一个原核在2小时内形成，8小时时原核形成达到最大值（>90%）。此外，化学激活的卵母细胞未排出第二极体。EA处理后，原核形成在4-6小时开始，新成熟卵母细胞在8小时时的原核形成率显著低于老化卵母细胞（34%对85%）。处理后24小时，CA处理的卵母细胞的分裂率低于EA处理的卵母细胞（<50%对>70%），并且在体外培养3天后，CA组中达到四细胞期的卵母细胞比EA组少。没有化学激活的卵母细胞发育超过四细胞期；相反，EA导致卵母细胞发育超过此阶段。CA和EA均导致H1激酶活性下降，但EA诱导的下降更快。采用EA时，H1激酶活性迅速下降，但在成熟36小时的卵母细胞中8小时时又趋于再次升高，这与所有其他组不同，其他组在激活后8小时激酶活性仍保持较低水平。我们得出结论，CA和EA在原核形成过程中涉及不同机制，并且6-DMAP激活的卵母细胞可能不适用于诱导猪的正常孤雌生殖发育。还表明，猪卵母细胞完全激活的能力可能与老化过程中H1激酶活性的改变有关。这些对H1激酶的改变可能是体外成熟卵母细胞孤雌激活以及减数分裂和分裂完成所必需的。
