Physiological significance of hypotonicity-induced regulatory volume decrease:: reduction in intracellular Cl- concentration acting as an intracellular signaling
Physiological significance of hypotonicity-induced regulatory volume decrease:: reduction in intracellular Cl- concentration acting as an intracellular signaling
复制标题
DOI：
10.1152/ajprenal.00244.2006
复制
发表时间：
2007-05-01
影响因子：
4.2
通讯作者：
Marunaka, Yoshinori
中科院分区：
文献类型：
--
作者：
Miyazaki, Hiroaki;Shiozaki, Atsushi;Marunaka, Yoshinori
低渗引起的细胞肿胀后会发生调节体积减少（RVD）。 RVD 是由离子通道和转运蛋白激活引起的，导致 K+、Cl- 和 H2O 外流，导致细胞萎缩。最近，我们发现低渗性通过肾上皮 A6 细胞中上皮 Na+ 通道 (α-ENaC) 表达的升高以 RVD 依赖性方式刺激跨上皮 Na+ 重吸收，而细胞内 Cl- 浓度 ([Cl-](i)) 的降低刺激了 Na+ 重吸收。这些表明 RVD 通过还原 [Cl-](i) 来揭示其对 Na+ 重吸收的刺激作用。然而，由于卤化物敏感荧光染料的技术困难，RVD 期间 [Cl-](i) 的还原尚未得到明确分析。在本研究中，我们开发了一种使用高分辨率流式细胞仪和卤化物特异性荧光染料 N-(6-甲氧基喹啉基)乙酰乙酯测量 RVD 期间 [Cl+](i) 变化的新方法。等渗培养基中 A6 细胞中的 [Cl+] i 为 43.6 +/- 3.1 mM。低渗休克（268至134 mosmol/ kgH(2)O）后，细胞体积迅速增加，随后发生RVD。 RVD 导致 [Cl-] i 从 43.6 mM 急剧减少至 10.8 mM。在用 NPPB（Cl- 通道阻断剂）或奎宁（K+ 通道阻断剂）阻断 RVD 的条件下，我们没有检测到 [Cl-] i 的减少。基于这些观察，我们得出结论，RVD 的生理意义之一是 [Cl-] i 的减少，并且 RVD 通过减少 [Cl-] i 作为调节细胞生理功能的细胞内信号来显示其作用。
Regulatory volume decrease (RVD) occurs after hypotonicity-caused cell swelling. RVD is caused by activation of ion channels and transporters, which cause effluxes of K+, Cl-, and H2O, leading to cell shrinkage. Recently, we showed that hypotonicity stimulated transepithelial Na+ reabsorption via elevation of epithelial Na+ channel (alpha-ENaC) expression in renal epithelia A6 cells in an RVD-dependent manner and that reduction of intracellular Cl- concentration ([Cl-](i)) stimulated the Na+ reabsorption. These suggest that RVD would reveal its stimulatory action on the Na+ reabsorption by reducing [Cl-](i). However, the reduction of [Cl-](i) during RVD has not been definitely analyzed due to technical difficulties involved in halide-sensitive fluorescent dyes. In the present study, we developed a new method for the measurement of [Cl+](i) change during RVD by using a high-resolution flow cytometer with a halide-specific fluorescent dye, N-(6-methoxyquinolyl) acetoethyl ester. The [Cl+] i in A6 cells in an isotonic medium was 43.6 +/- 3.1 mM. After hypotonic shock (268 to 134 mosmol/ kgH(2)O), a rapid increase of cell volume followed by RVD occurred. The RVD caused drastic diminution of [Cl-] i from 43.6 to 10.8 mM. Under an RVD- blocked condition with NPPB (Cl- channel blocker) or quinine (K+ channel blocker), we did not detect the reduction of [Cl-] i. Based on these observations, we conclude that one of the physiological significances of RVD is the reduction of [Cl-] i and that RVD shows its action via reduction of [Cl-] i acting as an intracellular signal regulating cellular physiological functions.