Effect of intracellular pH on depolarization-evoked calcium influx in human sperm
Effect of intracellular pH on depolarization-evoked calcium influx in human sperm
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DOI：
10.1152/ajpcell.00141.2004
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发表时间：
2004-12-01
影响因子：
5.5
通讯作者：
González-Martínez, MT
中科院分区：
文献类型：
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作者：
Fraire-Zamora, JJ;González-Martínez, MT
人类精子被赋予了假定的电压依赖性钙通道（VDCC），其产生可测量的细胞内钙浓度（[Ca2+](i)）的增加，以响应钾的膜去极化。这些通道被镍阻断，在无钙培养基中1-2分钟失活，并被NH4Cl显著刺激，这表明细胞内pH值（pH(i)）的作用。在之前的研究中，我们发现在体外“获能”过程中，通过这些通道的钙通透性增加了大约一倍，这是精子使卵子受精所需的钙依赖过程。在这项工作中，我们确定了精子VDCC的pH(i)依赖性。同时用NH4Cl去极化和pH(i)碱化诱导[Ca2+](i)增加，这取决于NH4Cl的添加量。VDCC刺激随pH值(i)的变化呈s型曲线，pH值在6.6 ~ 7.2范围内，pH值接近7.00时，VDCC刺激为半峰值。在较高的pH值(i)下（大于或等于7.3），会发生进一步的刺激。钙从内部储存的释放不有助于pH(i)的刺激作用，因为[Ca2+](i)增加由黄体酮诱导，打开钙渗透途径，不涉及门控VDCC，不受铵的影响。在不同的测试去极化值下，pH(i)刺激与非刺激钙内流的比例几乎是恒定的。同样，在pH(i)刺激和非刺激细胞中，去极化诱导的钙内流同样被镍阻断。在我们的获能条件下，pH(i)增加了0.11个pH单位，表明在精子获能过程中观察到的钙内流刺激可能部分是由pH(i)碱化引起的。此外，还检测到仅由pH(i)碱化触发的钙通透性途径。
Human sperm are endowed with putative voltage-dependent calcium channels (VDCC) that produce measurable increases in intracellular calcium concentration ([Ca2+](i)) in response to membrane depolarization with potassium. These channels are blocked by nickel, inactivate in 1-2 min in calcium-deprived medium, and are remarkably stimulated by NH4Cl, suggesting a role for intracellular pH (pH(i)). In a previous work, we showed that calcium permeability through these channels increases approximately onefold during in vitro "capacitation," a calcium-dependent process that sperm require to fertilize eggs. In this work, we have determined the pH(i) dependence of sperm VDCC. Simultaneous depolarization and pH(i) alkalinization with NH4Cl induced an [Ca2+](i) increase that depended on the amount of NH4Cl added. VDCC stimulation as a function of pH(i) showed a sigmoid curve in the 6.6-7.2 pH(i) range, with a half-maximum stimulation at pH similar to7.00. At higher pH(i) (greater than or equal to7.3), a further stimulation occurred. Calcium release from internal stores did not contribute to the stimulating effect of pH(i) because the [Ca2+](i) increase induced by progesterone, which opens a calcium permeability pathway that does not involve gating of VDCC, was unaffected by ammonium. The ratio of pH(i)-stimulated-to-nonstimulated calcium influx was nearly constant at different test depolarization values. Likewise, depolarization-induced calcium influx in pH(i)-stimulated and nonstimulated cells was equally blocked by nickel. In our capacitating conditions pH(i) increased 0.11 pH units, suggesting that the calcium influx stimulation observed during sperm capacitation might be partially caused by pH(i) alkalinization. Additionally, a calcium permeability pathway triggered exclusively by pH(i) alkalinization was detected.