3月6日及4月18日，国际著名《神经科学杂志》（The Journal of Neuroscience）相继发表上海交通大学基础医学院徐天乐教授课题组系列研究成果，揭示了酸敏感离子通道（ASIC）的内源性新配体和ASIC内吞分子机制及相关病理生理意义。

　　成果一：Serotonin facilitates peripheral pain sensitivity in a manner that depends on the nonproton ligand sensing domain of ASIC3 channel

　　组织酸化和炎症因子堆积是导致慢性痛的常见原因之一。徐天乐课题组通过电生理筛选，发现经典炎症因子五羟色胺（5-HT）能够特异增强ASIC3通道的持续激活。在之后的实验中通过综合运用分子细胞生物学、电生理学、配体结合和化学修饰等实验手段，发现5-HT的增强作用是通过与ASIC3通道上非质子配体感受域直接相互作用而实现的，该过程并不依赖于经典5-HT受体。不仅如此，5-HT能够显著易化酸化诱导的小鼠疼痛样行为，而该行为在ASIC3基因敲除小鼠明显减弱，表明了5-HT通过作用于ASIC3通道，协同易化炎性痛行为。这项成果揭示了ASIC3通道是5-HT作用的靶分子之一，为炎性慢性痛治疗提供了新途径，并因此被推荐为亮点论文。

   成果二：Molecular mechanism of constitutive endocytosis of acid-sensing ion channel 1a and its protective function in acidosis-induced neuronal death

   离子通道作为细胞接受外界信号的感受器，其功能的实现依赖于在细胞膜表面的正确定位。这一定位在上膜和内吞机制的共同调控下达到平衡。徐天乐课题组发现ASIC1a通道通过网格蛋白和动力蛋白依赖的途径内吞；位于ASIC1a胞内段羧基端的一段非经典内吞信号序列465LCRRG469，对ASIC1a与内吞分子机器之间的相互作用至关重要；阻断ASIC1a的内吞能够显著加重酸诱导的神经元死亡。此项研究成果在揭示ASIC1a细胞膜表达调控新机制的基础上，进一步提出了靶向ASIC1a膜转运分子机制的神经保护新策略。

　　徐天乐教授2011年5月受聘担任上海交通大学基础医学院院长、神经细胞信号调控调控研究组PI（http://yky.shsmu.edu.cn/LNS/index.html），在离子通道生物学功能和病理生理学意义研究方面取得了一系列创新性进展，曾多次在《Journal of Neuroscience》、《Neuron》等神经科学高水平期刊上发表研究成果。此次2篇论文第一作者王相、曾维政为其博士研究生，该成果在研期间受到国家自然科学基金重点项目资助。

原文摘要：

Serotonin Facilitates Peripheral Pain Sensitivity in a Manner That Depends on the Nonproton Ligand Sensing Domain of ASIC3 Channel

Tissue acidosis and inflammatory mediators play critical roles in inflammatory pain. Extracellular acidosis activates acid-sensing ion channels (ASICs), which have emerged as key sensors for extracellular protons in the central and peripheral nervous systems and play key roles in pain sensation and transmission. Additionally, inflammatory mediators, such as serotonin (5-HT), are known to enhance pain sensation. However, functional interactions among protons, inflammatory mediators, and ASICs in pain sensation are poorly understood. In the present study, we show that 5-HT, a classical pro-inflammatory mediator, specifically enhances the proton-evoked sustained, but not transient, currents mediated by homomeric ASIC3 channels and heteromeric ASIC3/1a and ASIC3/1b channels. Unexpectedly, the effect of 5-HT on ASIC3 channels does not involve activation of 5-HT receptors, but is mediated via a functional interaction between 5-HT and ASIC3 channels. We further show that the effect of 5-HT on ASIC3 channels depends on the newly identified nonproton ligand sensing domain. Finally, coapplication of 5-HT and acid significantly increased pain-related behaviors as assayed by the paw-licking test in mice, which was largely attenuated in ASIC3 knock-out mice, and inhibited by the nonselective ASIC inhibitor amiloride. Together, these data identify ASIC3 channels as an unexpected molecular target for acute actions of 5-HT in inflammatory pain sensation and reveal an important role of ASIC3 channels in regulating inflammatory pain via coincident detection of extracellular protons and inflammatory mediators.

Molecular Mechanism of Constitutive Endocytosis of Acid-Sensing Ion Channel 1a and Its Protective Function in Acidosis-Induced Neuronal Death

Acid-sensing ion channels (ASICs) are proton-gated cation channels widely expressed in the peripheral and CNSs, which critically contribute to a variety of pathophysiological conditions that involve tissue acidosis, such as ischemic stroke and epileptic seizures. However, the trafficking mechanisms of ASICs and the related proteins remain largely unknown. Here, we demonstrate that ASIC1a, the main ASIC subunit in the brain, undergoes constitutive endocytosis in a clathrin- and dynamin-dependent manner in both mouse cortical neurons and heterologous cell cultures. The endocytosis of ASIC1a was inhibited by either the small molecular inhibitor tyrphostin A23 or knockdown of the core subunit of adaptor protein 2 (AP2) μ2 using RNA interference, supporting a clathrin-dependent endocytosis of ASIC1a. In addition, the internalization of ASIC1a was blocked by dominant-negative dynamin1 mutation K44A and the small molecular inhibitor dynasore, suggesting that it is also dynamin-dependent. We show that the membrane-proximal residues 465LCRRG469 at the cytoplasmic C terminus of ASIC1a are critical for interaction with the endogenous adaptor protein complex and inhibition of ASIC1a internalization strongly exacerbated acidosis-induced death of cortical neurons from wild-type but not ASIC1a knock-out mice. Together, these results reveal the molecular mechanism of ASIC1a internalization and suggest the importance of endocytic pathway in functional regulation of ASIC1a channels as well as neuronal damages mediated by these channels during neurodegeneration.

作者简介：

徐天乐

徐天乐研究员现任上海交通大学医学院基础医学院，生化与分子细胞生物学系“神经细胞信号调控”研究组PI，研究员和博导，同时担任基础医学院院长和医学科学研究院院长。长期致力于研究神经细胞信号调控和重大脑疾病的关系，重点关注膜表面受体（如甘氨酸受体）和离子通道（如酸敏感离子通道）的生物学功能和作用机制。担任中国神经科学会常务理事和中国生物物理学会常务理事；国际学术期刊《ACS Chemical Neuroscience》、《Molecular Pain》、《Frontiers in Pharmacology of Ion Channel and Channelopathies》、《Neuroscience Bulletin》以及《Journal of Neurogenetics》编委。

实验室简介
神经细胞信号调控研究组以阐述离子通道与神经信号调控的关系为目标，致力于研究神经细胞膜离子通道的生物学功能和作用机制，探索重要疾病中离子通道和神经信号调控异常机制，在此基础上发现疾病相关靶标和新药先导化合物，指导新药发现。