Kline D, Kline J T
Department of Biological Sciences, Kent State University, Ohio 44242.
Dev Biol. 1992 Jan;149(1):80-9. doi: 10.1016/0012-1606(92)90265-i.
The role of calcium in cortical granule exocytosis and activation of the cell cycle at fertilization was examined in the mouse egg using the calcium chelator BAPTA (1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid) and the fluorescent calcium indicator fluo-3. BAPTA and fluo-3 were introduced into zona-free mouse eggs by a 30-min incubation with 0.01-50 microM BAPTA acetoxymethyl ester (AM) and/or 1-20 microM fluo-3 AM prior to in vitro fertilization. Incubation of eggs in greater than or equal to 5.0 microM BAPTA AM inhibited cortical granule exocytosis in all cases. Introduction of the calcium chelator into the egg blocked second polar body formation at greater than or equal to 1.0 microM BAPTA AM. Sperm entry occurred in all eggs regardless of the BAPTA AM concentration. Sperm induce a large transient increase in calcium lasting 2.3 +/- 0.6 min, followed by repetitive transients lasting 0.5 +/- 0.1 min and occurring at 3.4 +/- 1.4-min intervals. Incubation with greater than or equal to 5.0 microM BAPTA AM inhibited all calcium transients. Introduction of BAPTA also inhibited calcium transients, exocytosis, and the resumption of meiosis following application of the calcium ionophore A23187 or SrCl2, which activate eggs. These results demonstrate that the calcium increase at fertilization is required for cortical granule exocytosis and resumption of the cell cycle in a mammalian egg.
利用钙螯合剂BAPTA（1,2 - 双（邻氨基苯氧基）乙烷 - N，N，N'，N' - 四乙酸）和荧光钙指示剂fluo - 3，在小鼠卵中研究了钙在受精时皮质颗粒胞吐作用和细胞周期激活中的作用。在体外受精前，将无透明带的小鼠卵与0.01 - 50微摩尔/升的BAPTA乙酰氧基甲酯（AM）和/或1 - 20微摩尔/升的fluo - 3 AM孵育30分钟，将BAPTA和fluo - 3引入卵中。在所有情况下，将卵在大于或等于5.0微摩尔/升的BAPTA AM中孵育均会抑制皮质颗粒胞吐作用。当BAPTA AM浓度大于或等于1.0微摩尔/升时，将钙螯合剂引入卵中会阻止第二极体形成。无论BAPTA AM浓度如何，所有卵中都会发生精子进入。精子诱导钙大幅瞬时增加，持续2.3±0.6分钟，随后是重复的瞬时增加，持续0.5±0.1分钟，间隔为3.4±1.4分钟。与大于或等于5.0微摩尔/升的BAPTA AM孵育会抑制所有钙瞬变。引入BAPTA也会抑制钙瞬变、胞吐作用以及在应用钙离子载体A23187或SrCl2（可激活卵）后减数分裂的恢复。这些结果表明，受精时的钙增加是哺乳动物卵中皮质颗粒胞吐作用和细胞周期恢复所必需的。
