Abbott A L, Fissore R A, Ducibella T
Sackler School of Biomedical Sciences, Tufts University School of Medicine and New England Medical Center, Boston, Massachusetts, 02111, USA.
Dev Biol. 1999 Mar 1;207(1):38-48. doi: 10.1006/dbio.1998.9159.
Immature oocytes of many species are incompetent to undergo cortical granule (CG) exocytosis upon fertilization. In mouse eggs, CG exocytosis is dependent primarily on an inositol 1,4,5-trisphosphate (IP3)-mediated elevation of intracellular calcium ([Ca2+]i). While deficiencies upstream of [Ca2+]i release are known, this study examined whether downstream deficiencies also contribute to the incompetence of preovulatory mouse oocytes to release CGs. The experimental strategy was to bypass upstream deficiencies by inducing normal, fertilization-like [Ca2+]i oscillations in fully grown, germinal vesicle (GV) stage oocytes and determine if the extent of CG exocytosis was restored to levels observed in mature, metaphase II (MII)-stage eggs. Because IP3 does not stimulate a normal Ca2+ response in GV-stage oocytes, three alternate methods were used to induce oscillations: thimerosal treatment, electroporation, and sperm factor injection. Long-lasting oscillations from thimerosal treatment resulted in 64 and 10% mean CG release at the MII and GV stages, respectively (P < 0.001). Three electrical pulses induced mean [Ca2+]i elevations of approximately 730 and 650 nM in MII- and GV-stage oocytes, respectively, and 31% CG release in MII-stage eggs and 9% in GV-stage oocytes (P < 0.001). Sperm factor microinjection resulted in 86% CG release in MII-stage eggs, while similarly treated GV-stage oocytes exhibited < 1% CG release (P < 0.001). Taken together, these results demonstrate a deficiency downstream of [Ca2+]i release which is developmentally regulated in the 12 h prior to ovulation.
许多物种的未成熟卵母细胞在受精时无法进行皮质颗粒（CG）胞吐作用。在小鼠卵中，CG胞吐作用主要依赖于肌醇1,4,5-三磷酸（IP3）介导的细胞内钙（[Ca2+]i）升高。虽然已知[Ca2+]i释放上游存在缺陷，但本研究探讨了下游缺陷是否也导致排卵前小鼠卵母细胞释放CG的能力不足。实验策略是通过在完全成熟的生发泡（GV）期卵母细胞中诱导正常的、类似受精的[Ca2+]i振荡来绕过上游缺陷，并确定CG胞吐作用的程度是否恢复到在成熟的中期II（MII）期卵中观察到的水平。由于IP3不会在GV期卵母细胞中刺激正常的Ca2+反应，因此使用了三种替代方法来诱导振荡：硫柳汞处理、电穿孔和精子因子注射。硫柳汞处理引起的持久振荡分别导致MII期和GV期的平均CG释放率为64%和10%（P < 0.001）。三个电脉冲分别在MII期和GV期卵母细胞中诱导平均[Ca2+]i升高约730和650 nM，MII期卵中的CG释放率为31%，GV期卵母细胞中的释放率为9%（P < 0.001）。精子因子显微注射导致MII期卵中的CG释放率为86%，而同样处理的GV期卵母细胞的CG释放率<1%（P < 0.001）。综上所述，这些结果表明[Ca2+]i释放下游存在缺陷，该缺陷在排卵前12小时受到发育调控。
