Is autocrine ATP release required for activation of volume-sensitive chloride channels?
Is autocrine ATP release required for activation of volume-sensitive chloride channels?
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体积敏感氯离子通道的激活是否需要自分泌 ATP 释放？
DOI：
10.1152/jn.00615.2003
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发表时间：
2003
影响因子：
2.5
通讯作者：
Kimelberg,HaroldK
中科院分区：
文献类型：
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作者：
Mongin,AlexanderA;Kimelberg,HaroldK
致编辑：我们怀着极大的兴趣阅读了达比等人发表在《神经生理学杂志》2003年4月刊上的论文（达比等人，2003）。作者证明，在培养的星形胶质细胞中，低渗细胞肿胀通过与多药耐药转运蛋白相似的药理学特征的途径导致ATP释放。然后，这种ATP刺激P2 Y1样受体，并通过适当的细胞内信号传导机制激活体积调节阴离子通道（VRAC），测量为全细胞Cl电流。他们还表明，在非肿胀的星形胶质细胞中，外源性ATP激活Cl电流，类似于VRAC携带的电流。这些数据符合先前提出的自分泌ATP释放模型，该模型对于肝癌细胞中VRAC的活化是必需的（Wang et al. 1996）。值得注意的是，星形胶质细胞中这些发现的意义超出了体积敏感性氯离子通道如何响应细胞体积变化而被激活的“学术”问题。星形胶质细胞VRAC及其在其他细胞中的对应物可渗透各种小的有机阴离子和不带电分子，包括兴奋性氨基酸谷氨酸和天冬氨酸（Kimelberg et al. 1990）。因此，在脑中，在突触传递或星形胶质细胞-星形胶质细胞通信期间释放的ATP可能导致VRAC介导的谷氨酸从星形胶质细胞释放。这种星形胶质细胞谷氨酸释放现在被认为是神经元星形胶质细胞通信中的关键反馈信号（Haydon 2001），ATP可能充当主要的神经元到星形胶质细胞和星形胶质细胞到星形胶质细胞的细胞间信使（Fields和Stevens 2000）。我们还研究了ATP在星形胶质细胞VRACs活化中的作用，测量为预加载的[3 H]-标记的兴奋性氨基酸D-天冬氨酸的释放。我们发现，在培养的星形胶质细胞中，外源性10 μM ATP激活非肿胀细胞中的短暂兴奋性氨基酸释放，并强烈增强低渗肿胀细胞中的氨基酸释放（Mongin and Kimelberg 2002;图1）。这种ATP诱导的D-[3 H]天冬氨酸外排可被VRAC阻断剂5-硝基-2-（3-苯丙氨基）苯甲酸、DIDS和根皮素有效抑制。中等渗透压摩尔浓度增加10%也可抑制该作用（Mongin and Kimelberg 2002）。在这些数据的基础上，我们提出ATP不能直接激活VRAC，而是积极地调节一小部分在非肿胀细胞中活跃的VRAC。ATP诱导的D-[3 H]天冬氨酸释放在我们的实验涉及非肿胀细胞不超过十分之一的释放诱导的介质渗透压降低100 mOsm。相比之下，达比及其同事报告了ATP在非肿胀星形胶质细胞中对Cl电流的激活显著更高，达到低渗Cl电流值的一半（达比等人，2003）。如果转化为ATP诱导的VRAC介导的谷氨酸释放，这种作用应该对正常的脑生理产生很大影响。达比等人观察到的ATP的作用是浓度依赖性的，在1和5 mM时具有最大值。然而，细胞外的毫摩尔浓度的ATP阻断星形胶质细胞中VRAC介导的兴奋性氨基酸释放（Haskew等人，2002年; Mongin和Kimelberg 2002），以及几种细胞系中VRAC介导的Cl电流，IC 50在0.2和5 mM ATP之间变化（Okada 1997）。如图1所示，在培养的星形胶质细胞中，10 μM、100 μM和1 mM ATP以相同的效力激活瞬时D-[3 H]天冬氨酸释放。然而，在5和10 mM ATP时，D-[3 H]天冬氨酸释放的第一个瞬时相不存在，并且可能被抑制。
To the Editor: We read with great interest the paper by Darby et al. in the April 2003 issue of the Journal of Neurophysiology (Darby et al. 2003). The authors demonstrated that, in cultured astrocytes, hyposmotic cell swelling causes release of ATP via a pathway with pharmacological profile similar to the multidrug resistance transporter. This ATP then stimulates P2Y1-like receptors and activates, via appropriate intracellular signaling mechanisms, volume-regulated anion channels (VRACs), measured as whole cell Cl currents. They also show that in nonswollen astrocytes, exogenous ATP activates Cl currents resembling currents carried by VRACs. These data fit the previously proposed model of autocrine ATP release that is obligatory for the activation of VRACs in hepatoma cells (Wang et al. 1996). It is important to note that the significance of such findings in astrocytes extends beyond the “academic” question of how volume-sensitive chloride channels are activated in response to cell volume changes. Astrocytic VRACs, as well as their counterparts in other cells, are permeable toward a variety of small organic anions and uncharged molecules, including the excitatory amino acids glutamate and aspartate (Kimelberg et al. 1990). Therefore in the brain, ATP released during synaptic transmission or astrocyteto-astrocyte communication may cause VRAC-mediated glutamate release from astrocytes. Such astrocytic glutamate release is now considered a key feedback signal in neuronastrocyte communication (Haydon 2001), with ATP likely acting as the main neuron-to-astrocyte and astrocyte-to-astrocyte intercellular messenger (Fields and Stevens 2000). We have also studied the role of ATP in activation of astrocytic VRACs, measured as release of preloaded [3H]-labeled excitatory amino acid D-aspartate. We found that, in cultured astrocytes, exogenous 10 μM ATP activates a transient excitatory amino acid release in nonswollen cells and strongly potentiates amino acid release in hyposmotically swollen cells (Mongin and Kimelberg 2002; Fig. 1). This ATP-induced D-[3H] aspartate efflux was potently inhibited by the VRAC blockers 5-nitro-2-(3-phenylpropylamino) benzoic acid, DIDS, and phloretin. It was also suppressed by a 10% increase in medium osmolarity (Mongin and Kimelberg 2002). On the basis of these data, we propose that ATP is incapable of direct VRAC activation, but instead, positively modulates a small fraction of VRACs that are active in nonswollen cells. The ATP-induced D-[3H] aspartate release in our experiments involving nonswollen cells did not exceed one-tenth of the release induced by a 100 mOsm reduction in medium osmolarity. In contrast, Darby and colleagues report a substantially higher activation of the Cl currents by ATP in nonswollen astrocytes, reaching one-half of the value of the hyposmotic Cl currents (Darby et al. 2003). If translated to ATP-induced VRAC-mediated glutamate release, such effect should have a large impact on normal brain physiology. The effect of ATP seen by Darby et al. was concentrationdependent, with the maximum values at 1 and 5 mM. However, millimolar concentrations of extracellular ATP block VRAC-mediated excitatory amino acid release in astrocytes (Haskew et al. 2002; Mongin and Kimelberg 2002), as well as VRAC-mediated Cl currents in several cell lines, with the IC50 varying between 0.2 and 5 mM ATP (Okada 1997). As seen in Fig. 1, in cultured astrocytes, 10 μM, 100 μM, and 1 mM ATP activated the transient D-[3H] aspartate release with equal potency. However, at 5 and 10 mM ATP, the first transient phase of D-[3H] aspartate release is not present and is presumably inhibited …