Pharmacological targeting of native CatSper channels reveals a required role in maintenance of sperm hyperactivation.
Pharmacological targeting of native CatSper channels reveals a required role in maintenance of sperm hyperactivation.
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DOI：
10.1371/journal.pone.0006844
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发表时间：
2009-08-31
期刊：
影响因子：
3.7
通讯作者：
Babcock DF
中科院分区：
文献类型：
--
作者：
Carlson AE;Burnett LA;del Camino D;Quill TA;Hille B;Chong JA;Moran MM;Babcock DF
这四种精子特异性CatSper离子通道蛋白是超活化运动和男性生育力以及碱性去极化引起的Ca 2+进入所必需的。在没有外部Ca 2+的情况下，Na+通过电压钳位精子中的CatSper通道携带电流。在这里，我们表明，CatSper通道活动可以监测光学与[Na+] i-报告探针SBFI在群体中的完整精子。去除外部Ca 2+会增加野生型精子的SBFI信号，但不会增加CatSper 2缺失精子的SBFI信号。用NH 4Cl碱化的精子的[Na+]i的指示上升速率大于用丙酸酸化的精子，反映了CatSper电流的碱促进特征特性。相比之下，候选CatSper阻断剂HC-056456（IC 50 ≤ 3 µM）可减缓[Na+]i升高。HC-056456同样减缓了由碱性去极化诱发并由fura-2报告的[Ca 2 +]i升高。HC-056456还选择性和可逆地降低从膜片钳精子记录的CatSper电流。HC-056456不能阻止HCO 3-激活运动性，但可以通过获能孵育阻止超激活运动性的发展，从而产生CatSper无效精子的表型。当应用于超活化精子时，HC-056456导致鞭毛波形不对称性的快速、可逆损失，类似于通过去除外部Ca 2+终止Ca 2+通过CatSper通道进入时发生的损失。因此，开放的CatSper通道和外部Ca 2+通过它们的进入维持了过度激活的运动性。这些结果表明，CatSper通道的药理学靶向可能会对生育力施加选择性晚期阻滞，并且使用CatSper通道活性的光学报告子进行高通量筛选可能会识别出具有男性定向避孕潜力的其他选择性阻滞剂。
The four sperm-specific CatSper ion channel proteins are required for hyperactivated motility and male fertility, and for Ca2+ entry evoked by alkaline depolarization. In the absence of external Ca2+, Na+ carries current through CatSper channels in voltage-clamped sperm. Here we show that CatSper channel activity can be monitored optically with the [Na+]i-reporting probe SBFI in populations of intact sperm. Removal of external Ca2+ increases SBFI signals in wild-type but not CatSper2-null sperm. The rate of the indicated rise of [Na+]i is greater for sperm alkalinized with NH4Cl than for sperm acidified with propionic acid, reflecting the alkaline-promoted signature property of CatSper currents. In contrast, the [Na+]i rise is slowed by candidate CatSper blocker HC-056456 (IC50 ∼3 µM). HC-056456 similarly slows the rise of [Ca2+]i that is evoked by alkaline depolarization and reported by fura-2. HC-056456 also selectively and reversibly decreased CatSper currents recorded from patch-clamped sperm. HC-056456 does not prevent activation of motility by HCO3 − but does prevent the development of hyperactivated motility by capacitating incubations, thus producing a phenocopy of the CatSper-null sperm. When applied to hyperactivated sperm, HC-056456 causes a rapid, reversible loss of flagellar waveform asymmetry, similar to the loss that occurs when Ca2+ entry through the CatSper channel is terminated by removal of external Ca2+. Thus, open CatSper channels and entry of external Ca2+ through them sustains hyperactivated motility. These results indicate that pharmacological targeting of the CatSper channel may impose a selective late-stage block to fertility, and that high-throughput screening with an optical reporter of CatSper channel activity may identify additional selective blockers with potential for male-directed contraception.
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