A role for the human sperm glycine receptor/Cl- channel in the acrosome reaction initiated by recombinant ZP3
A role for the human sperm glycine receptor/Cl- channel in the acrosome reaction initiated by recombinant ZP3
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DOI：
10.1095/biolreprod66.1.91
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发表时间：
2002-01-01
影响因子：
3.6
通讯作者：
Meizel, S
中科院分区：
文献类型：
--
作者：
Bray, C;Son, JH;Meizel, S
在此之前，我们已经证明了神经元甘氨酸受体（GlyR）在小鼠和猪精子的顶体反应（AR）由卵透明带（ZP）启动的重要作用。在本研究中，我们已经证明了存在的GlyR在人类精子的免疫沉淀和蛋白质印迹分析，研究了潜在的重组人ZP 3（rhZP 3）制剂作为一种替代的研究工具，以溶解人ZP，并表明，人精子GlyR是必不可少的人AR由rhZP 3启动。此外，我们已经能够证明rhZP 3具有生物活性，因为它能够快速刺激获能人精子中的AR，并且其作用被添加百日咳毒素阻断。此外，荧光分光光度法研究使用fura-2加载的人精子已经表明，rhZP 3触发细胞内Ca 2+水平的峰值和平台升高，类似于溶解的哺乳动物ZP。这些结果表明，rhZP 3和溶解的ZP的作用是通过相同的信号转导途径引起的。此外，人精子与针对人脊髓GlyR的α 1亚基的抗体或与50 nM士的宁一起孵育在rhZP 3引发的AR中引起显著抑制。最后，使用载有fura-2的人精子的研究表明，50 nM士的宁也能够抑制与添加rhZP 3相关的Ca 2+内流。这些结果进一步支持了rhZP 3和ZP通过相同机制起作用的观点，表明GlyR参与rhZP 3启动的AR，并表明GlyR也可能在体内与ZP启动的AR相关的早期信号转导级联中起作用。
Previously, we have demonstrated an essential role for the neuronal glycine receptor (GlyR) in the acrosome reaction (AR) of mouse and porcine sperm initiated by the egg zona pellucida (ZP). In the present study, we have demonstrated presence of the GlyR in human sperm by immunoprecipitation and Western blot analysis, investigated the potential of a recombinant human ZP3 (rhZP3) preparation as an alternative research tool to solubilized human ZP, and shown that the human sperm GlyR is essential to the human AR initiated by rhZP3. Additionally, we have been able to demonstrate that rhZP3 possesses biological activity, because it is able to rapidly stimulate the AR in capacitated human sperm and its action is blocked by the addition of pertussis toxin. Moreover, spectrofluorometric studies using fura-2-loaded human sperm have shown that rhZP3 triggers a peak-and-plateau rise in intracellular Ca2+ levels similar to that seen with solubilized mammalian ZP These results suggest that the actions of rhZP3 and solubilized ZP are elicited via the same signal transduction pathways. Furthermore, incubation of human sperm with an antibody directed against the a, subunit of the human spinal cord GlyR or with 50 nM strychnine caused significant inhibition in the rhZP3-initated AR. Finally, studies using fura-2-loaded human sperm showed that 50 nM strychnine was also able to inhibit the Ca2+ influx associated with addition of rhZP3. These results further support the view that rhZP3 and the ZP work through the same mechanisms, show that the GlyR is involved in rhZP3-initiated AR, and suggest that the GlyR may also play a role in the early signal transduction cascades associated with ZP-initiated AR in vivo.