Lawrence Y, Ozil J P, Swann K
Department of Anatomy and Developmental Biology, University College, Gower Street, London WC1E 6BT, UK.
Biochem J. 1998 Oct 15;335 ( Pt 2)(Pt 2):335-42. doi: 10.1042/bj3350335.
During fertilization in mouse eggs, the sperm triggers a series of intracellular Ca2+ oscillations that lead to egg activation, as indicated by pronuclear formation. We show that Ca2+ oscillations in fertilized mouse eggs can be inhibited by addition of either the Ca2+ chelator 1,2-bis-(o-aminophenoxy)ethane-N,N,N',N'-tetra-acetic acid acetoxymethyl ester (BAPTA-AM) or the heavy-metal-ion chelator N,N,N',N'-tetrakis-(2-pyridylmethyl)ethylenediamine (TPEN) plus dithiothreitol (DTT). Both treatments inhibited Ca2+ oscillations, but they had different effects upon egg activation. Blocking Ca2+ oscillations with BAPTA-AM after the occurrence of just two Ca2+ spikes resulted in most eggs forming pronuclei. However, we found that BAPTA-AM-treated fertilizing eggs showed a decreased rate of protein synthesis, which by itself can promote egg activation. In contrast, blocking Ca2+ oscillations with TPEN plus DTT was accompanied by the inhibition of egg activation with no significant effect on protein synthesis. In eggs that were fertilized and then treated with TPEN plus DTT, there was a correlation between the number of Ca2+ spikes and the proportion of eggs that formed pronuclei, as well as between the number of Ca2+ spikes and the time taken for pronuclear formation and the first mitosis to occur. The addition of TPEN plus DTT did not block the generation of Ca2+ spikes or pronuclear formation when eggs were artificially stimulated by electroporation pulses. These data suggest that TPEN plus DTT inhibits pronuclear formation in fertilizing eggs via the inhibition of Ca2+ oscillations and that the number of Ca2+ spikes may regulate egg activation.
在小鼠卵子受精过程中，精子触发一系列细胞内钙离子振荡，这些振荡会导致卵子激活，原核形成即表明了这一点。我们发现，添加钙离子螯合剂1,2-双（邻氨基苯氧基）乙烷-N,N,N',N'-四乙酸乙酰甲酯（BAPTA-AM）或重金属离子螯合剂N,N,N',N'-四（2-吡啶甲基）乙二胺（TPEN）加二硫苏糖醇（DTT）可抑制受精小鼠卵子中的钙离子振荡。两种处理均抑制了钙离子振荡，但它们对卵子激活有不同影响。在仅出现两个钙离子峰值后用BAPTA-AM阻断钙离子振荡，导致大多数卵子形成原核。然而，我们发现经BAPTA-AM处理的受精卵蛋白质合成速率降低，而蛋白质合成本身可促进卵子激活。相比之下，用TPEN加DTT阻断钙离子振荡伴随着卵子激活的抑制，对蛋白质合成无显著影响。在受精后用TPEN加DTT处理的卵子中，钙离子峰值数量与形成原核的卵子比例之间，以及钙离子峰值数量与原核形成和第一次有丝分裂发生所需时间之间存在相关性。当通过电穿孔脉冲人工刺激卵子时，添加TPEN加DTT不会阻断钙离子峰值的产生或原核形成。这些数据表明，TPEN加DTT通过抑制钙离子振荡抑制受精卵中的原核形成，并且钙离子峰值数量可能调节卵子激活。
