来源：神经科学通报
“突触”一词最早出自1897年由生理学家Michael Foster编撰的Textbook of Physiology（《生理学教科书》），源于希腊语Synapsis，意思是“连接”。现代神经科学将突触定义为神经元之间或神经元与效应细胞之间相互接触并实现功能联系的节点。当神经冲动到达上一级神经元（突触前神经元）的轴突终末时，触发神经递质的释放，这些神经递质穿过突触间隙作用到突触后膜受体，将神经信号传递给下一级神经元（突触后神经元）。因此，突触后膜受体在突触传递过程中发挥信号“接收器”的关键作用。通过构建特异的膜受体标记策略，结合荧光可视化技术，定位和实时追踪特定膜受体动态变化规律，是理解突触生理功能以及突触异常相关神经和精神疾病机制的首要任务。
Neuroscience Bulletin在2021年第2期以封面文章形式发表了上海交通大学医学院徐天乐教授团队的技术方法论文Postsynaptic Targeting and Mobility of Membrane Surface-Localized hASIC1a。封面图中，孙悟空的火眼金睛代表高效特异的神经元膜表面hASIC1a标记与可视化工具，识破白骨精的伪装代表对hASIC1a时空特异性的突触定位和动态膜转运规律的揭示。
突触，这一高度特化且复杂的亚细胞结构承载着神经细胞间的信息传递，是脑功能和整体行为的基础[1-3]。突触前、后膜遍布数量种类庞杂的受体和离子通道，这些膜蛋白的精确定位和动态调节是神经元执行信息输入、加工和传导的关键。而受体和离子通道的定位或数量调控异常是多种神经和精神性疾病的潜在机制[3, 4]。由此，神经元如何维持突触受体和离子通道的动态平衡是神经科学领域的重要科学问题。
图1 谷氨酸受体的膜转运与突触可塑性
（修改自Gambino F, Choquet D. Eyes Wide Open on AMPAR Trafficking during Motor Learning. Neuron. 2020）
膜受体或离子通道主要在细胞表面发挥功能。对于谷氨酸受体的突触定位和膜转运已经有比较系统的认识[5-7]。大量研究揭示谷氨酸受体突触定位和膜转运决定兴奋性突触传递、突触长时程增强（Long-term potentiation, LTP）与长时程抑制（Long-term depression, LTD）以及学习记忆等脑认知功能（图1）[8-11]。有趣的是，近年发现酸敏感离子通道1a（Acid-sensing ion channel 1a, ASIC1a）也参与突触传递和可塑性[12-17]。ASIC家族主要由4个基因（ACCN1，ACCN2，ACCN3，ACCN4）编码6种亚基（ASIC1a，ASIC1b，ASIC2a，ASIC2b，ASIC3，ASIC4），在脑中ASIC1a是通道功能的核心亚基。三聚体离子通道（ASIC4除外）感受胞外H+升高而开放，主要引起Na+和Ca2+内流，贡献学习记忆、慢性痛和负性情绪等生理和病理过程（图2）。但ASIC1a调节兴奋性突触功能的具体机制尚不清楚。通常认为突触小泡与突触前膜融合，H⁺和谷氨酸被一同释放到突触间隙，激活位于突触后膜的ASIC1a，从而贡献突触传递和可塑性，并调节学习记忆等相关行为（图2）[14, 18-21]。
图2 ASIC1a的结构功能
（修改自Jasti J et al. Structure of acidsensing ion channel 1 at 1.9 A resolution and low pH. Nature.2007和Kreple CJ et al. Drug abuse andthe simplest neurotransmitter. ACS Chem Neurosci. 2014）
然而，作为一种非经典突触后膜受体，神经元膜表面功能性ASIC1a的突触定位和膜转运规律仍不清楚，主要原因在于缺乏用于细胞和组织免疫染色的ASIC1a特异性抗体等标记工具。虽然小鼠（mASIC1a）和人源ASIC1a（hASIC1a）序列同源性高达98%，但两者在膜表面丰度和通道特性等方面存在差异，而对hASIC1a的研究更显不足[22-24]。徐天乐教授团队与合作者综合分子克隆、单细胞电生理、噬菌体展示、免疫染色和活细胞成像、单分子示踪等手段，筛选hASIC1a胞外域位点以插入标记序列（hASIC1a-298HA299或hASIC1a-298pHluorin299）和从噬菌体组合抗体库中筛选特异识别hASIC1a胞外域构象的抗体[25]（ASC06-IgG1或ASC06-IgG1-Alexa488），开发了细胞膜表面功能性hASIC1a的标记和可视化工具，用于固定状态下研究神经元表面hASIC1a的亚细胞定位和活细胞实时观察树突上hASIC1a的动态转运（图3）。hASIC1a分布于皮层神经元胞体和树突，尤其在突触后膜富集；活细胞及单分子水平观察到树突上hASIC1a的正向插膜和侧向迁移，外源给予脑源性营养因子（Brain-derived neurotrophic factor, BDNF）可以促进hASIC1a的膜表达和侧向运动以及靶向突触的膜转运（图4）。这项研究为ASICs领域提供了关键的标记工具和技术支持，揭示了hASIC1a在神经元细胞膜表面的分布和运动规律，提示hASIC1a时空转运在突触功能调节中的关键作用。
图3 神经元细胞膜表面hASIC1a的标记与可视化。
hASIC1a的胞外域抗体ASC06-IgG1或HA标签用于固定状态下神经元膜表面hASIC1a的标记和定位分析（I）；胞外域pHluorin标签用于活细胞状态下实时观测突触表面的hASIC1a动态变化（II）；荧光基团修饰抗体ASC06-IgG1-Alexa 488用于单分子或单颗粒水平研究树突表面hASIC1a的侧向运动（III）。
未来利用该工具，借助人干细胞分化神经元和hASIC1a基因敲入小鼠，结合超高分辨和在体双光子成像，在更精细的水平研究内源hASIC1a的突触定位和膜转运规律，探究hASIC1a的动态调控和突触结构功能以及动物行为的关联，最终解析H+/ASICs参与学习记忆等脑功能的新机制，甚至在整体水平靶向ASIC1a膜转运实现对慢性痛、恐惧和焦虑的干预。
图4 BDNF调控hASIC1a的动态膜转运
荧光漂白恢复实验表明BDNF促进ASIC1a靶向树突棘的正向膜转运；单分子示踪实验提示BDNF加速树突表面hASIC1a沿着细胞膜的侧向迁移。
Song XL, Liu DS, Qiang M, Li Q, Liu MG, Li WG, et al. Postsynaptic targetingand mobility of membrane surface-localized hASIC1a. Neurosci Bull 2021, 37:145-165.
参考文献
[1] Hebb DO. The organization of behavior; a neuropsychological theory. New York,: Wiley, 1949.
[2] Takeuchi T, Duszkiewicz AJ, Morris RG. The synaptic plasticity and memory hypothesis: encoding, storage and persistence. Philos Trans R Soc Lond B Biol Sci 2014, 369: 20130288.
[3] Volk L, Chiu SL, Sharma K, Huganir RL. Glutamate synapses in human cognitive disorders. Annu Rev Neurosci 2015, 38: 127-149.
[4] Imbrici P, Liantonio A, Camerino GM, De Bellis M, Camerino C, Mele A, et al. Therapeutic Approaches to Genetic Ion Channelopathies and Perspectives in Drug Discovery. Front Pharmacol 2016, 7: 121.
[5] Compans B, Choquet D, Hosy E. Review on the role of AMPA receptor nano-organization and dynamic in the properties of synaptic transmission. Neurophotonics 2016, 3: 041811.
[6] Choquet D, Triller A. The dynamic synapse. Neuron 2013, 80: 691-703.
[7] Newpher TM, Ehlers MD. Glutamate receptor dynamics in dendritic microdomains. Neuron 2008, 58: 472-497.
[8] Gambino F, Choquet D. Eyes Wide Open on AMPAR Trafficking during Motor Learning. Neuron 2020, 105: 764-766.
[9] Kessels HW, Malinow R. Synaptic AMPA receptor plasticity and behavior. Neuron 2009, 61: 340-350.
[10] Diering GH, Huganir RL. The AMPA Receptor Code of Synaptic Plasticity. Neuron 2018, 100: 314-329.
[11] Choquet D. Linking Nanoscale Dynamics of AMPA Receptor Organization to Plasticity of Excitatory Synapses and Learning. J Neurosci 2018, 38: 9318-9329.
[12] Wemmie JA, Chen J, Askwith CC, Hruska-Hageman AM, Price MP, Nolan BC, et al. The acid-activated ion channel ASIC contributes to synaptic plasticity, learning, and memory. Neuron 2002, 34: 463-477.
[13] Wemmie JA, Coryell MW, Askwith CC, Lamani E, Leonard AS, Sigmund CD, et al. Overexpression of acid-sensing ion channel 1a in transgenic mice increases acquired fear-related behavior. Proc Natl Acad Sci U S A 2004, 101: 3621-3626.
[14] Du J, Reznikov LR, Price MP, Zha XM, Lu Y, Moninger TO, et al. Protons are a neurotransmitter that regulates synaptic plasticity in the lateral amygdala. Proc Natl Acad Sci U S A 2014, 111: 8961-8966.
[15] Kreple CJ, Lu Y, Taugher RJ, Schwager-Gutman AL, Du J, Stump M, et al. Acid-sensing ion channels contribute to synaptic transmission and inhibit cocaine-evoked plasticity. Nat Neurosci 2014, 17: 1083-1091.
[16] Li WG, Liu MG, Deng S, Liu YM, Shang L, Ding J, et al. ASIC1a regulates insular long-term depression and is required for the extinction of conditioned taste aversion. Nat Commun 2016, 7: 13770.
[17] Li HS, Su XY, Song XL, Qi X, Li Y, Wang RQ, et al. Protein Kinase C Lambda Mediates Acid-Sensing Ion Channel 1a-Dependent Cortical Synaptic Plasticity and Pain Hypersensitivity. J Neurosci 2019, 39: 5773-5793.
[18] Jasti J, Furukawa H, Gonzales EB, Gouaux E. Structure of acid-sensing ion channel 1 at 1.9 A resolution and low pH. Nature 2007, 449: 316-323.
[19] Beg AA, Ernstrom GG, Nix P, Davis MW, Jorgensen EM. Protons act as a transmitter for muscle contraction in C. elegans. Cell 2008, 132: 149-160.
[20] Highstein SM, Holstein GR, Mann MA, Rabbitt RD. Evidence that protons act as neurotransmitters at vestibular hair cell-calyx afferent synapses. Proc Natl Acad Sci U S A 2014, 111: 5421-5426.
[21] Kreple CJ, Lu Y, LaLumiere RT, Wemmie JA. Drug abuse and the simplest neurotransmitter. ACS Chem Neurosci 2014, 5: 746-748.
[22] Xu Y, Jiang YQ, Li C, He M, Rusyniak WG, Annamdevula N, et al. Human ASIC1a mediates stronger acid-induced responses as compared with mouse ASIC1a. FASEB J 2018, 32: 3832-3843.
[23] Jing L, Chu XP, Jiang YQ, Collier DM, Wang B, Jiang Q, et al. N-glycosylation of acid-sensing ion channel 1a regulates its trafficking and acidosis-induced spine remodeling. J Neurosci 2012, 32: 4080-4091.
[24] Li M, Inoue K, Branigan D, Kratzer E, Hansen JC, Chen JW, et al. Acid-sensing ion channels in acidosis-induced injury of human brain neurons. J Cereb Blood Flow Metab 2010, 30: 1247-1260.
[25] Qiang M, Dong X, Zha Z, Zuo XK, Song XL, Zhao L, et al. Selection of an ASIC1a-blocking combinatorial antibody that protects cells from ischemic death. Proc Natl Acad Sci U S A 2018, 115: E7469-E7477.
特别声明：以上内容(如有图片或视频亦包括在内)为自媒体平台“网易号”用户上传并发布，本平台仅提供信息存储服务。
Notice: The content above (including the pictures and videos if any) is uploaded and posted by a user of NetEase Hao, which is a social media platform and only provides information storage services.
