Voltage imaging of astrocyte-neuron interactions

星形胶质细胞-神经元相互作用的电压成像

• 批准号: 10192852
• 负责人: Chris G Dulla
• 金额: $ 61.24万
• 依托单位: TUFTS UNIVERSITY BOSTON
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-15 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10192852
• 关键词: Acute; Address; Adult; Affect; Amino Acid Transporter; Area; Astrocytes; Ataxia; Binding; Brain; Buffers; Cerebral cortex; Characteristics; Data; Disease; Distal; Due Process; Electrophysiology (science); Ensure; Epilepsy; Excitatory Amino Acids; Extracellular Space; Frequencies; Genetic; Glutamate Transporter; Glutamates; Housekeeping; Image; Kinetics; Knowledge; Lead; Measures; Membrane; Membrane Potentials; Migraine; Monitor; Morphology; Mutation; N-Methyl-D-Aspartate Receptors; N-Methylaspartate; Neurons; Neurotransmitters; Optics; Permeability; Pharmacology; Potassium; Process; Property; Prosencephalon; Receptor Activation; Resistance; Resistance Process; Rodent; Shapes; Signal Transduction; Site; Slice; Sodium; Specificity; Synapses; Synaptic Transmission; Synaptic plasticity; Testing; base; designer receptors exclusively activated by designer drugs; extracellular; glutamatergic signaling; in vivo; insight; millisecond; neuronal cell body; neuronal patterning; neurotransmission; novel; optogenetics; response; sensor; spatiotemporal; tool; uptake; voltage

Project summary Astrocytes remove the excitatory neurotransmitter glutamate from the extracellular space following neuronal activity via sodium-driven, voltage-dependent excitatory amino acid transporters (EAATs). Robust glutamate uptake by EAATs ensures the temporal and spatial fidelity of glutamate signaling. Interestingly, we recently found that neuronal activity rapidly (within milliseconds) and reversibly slows glutamate uptake in the adult cerebral cortex. This slowing prolongs neuronal NMDA responses, consistent with prolonged extracellular glutamate dynamics, and is highly dependent on the frequency and duration of stimulation. Additionally, glutamate clearance can be modulated by neuronal activity with synapse specificity, even within a single astrocyte. We believe this may have important consequences on neurotransmission, extrasynaptic receptor activation, and synaptic plasticity. Based on this finding, we hypothesized that neuronal activity induces microdomain-level changes in astrocyte membrane potential (Vm) that locally modulate EAAT function. GLT1 is the predominant astrocytic EAAT in the adult forebrain, is abundantly expressed, and ensures that glutamate in the extracellular space is rapidly sequestered by EAAT binding. Once bound to EAATs, the transport of glutamate into the astrocyte is both sodium-driven and voltage-dependent. Under normal conditions, astrocytes are hyperpolarized (-80 mV) due to their high permeability to potassium. However, neuronal activity increases extracellular potassium, [K+]e, and astrocyte Vm is especially sensitive to [K+]e changes. Therefore, it stands to reason that neuronal activity can alter EAAT function by depolarizing astrocytes. Changes in astrocytic Vm may be especially relevant in fine astrocytic processes, where EAATs are concentrated, and where small intracellular volumes may amplify changes in Vm, as compared to soma. We will also explore alternative mechanisms including voltage-independent modulation of EAATs by increases in [K+]e. A major challenge to testing our hypothesis, however, is an inability to monitor astrocyte Vm at distal processes due to low membrane resistance and process morphology. Overcoming this challenge is important because astrocyte distal processes are the site of synaptic interaction and EAATs localization. In order to detect distal changes in astrocyte Vm, we developed an approach to image Vm in astrocyte processes using genetically-encoded voltage indicator (GEVI) imaging. Utilizing astrocyte and neuron electrophysiological recording, optogenetic manipulation of astrocyte Vm, and GEVI imaging of astrocyte membrane potential we have generated preliminary data that supports our hypothesis that EAAT function can be modulated by activity-induced changes in astrocyte Vm.

项目摘要 在神经元之后，星形胶质细胞从细胞外空间中去除兴奋性神经递质谷氨酸 通过钠驱动的电压依赖性兴奋性氨基酸转运蛋白（EAATS）的活性。坚固的谷氨酸 Eaats的吸收可确保谷氨酸信号传导的时间和空间保真度。有趣的是，我们最近 发现神经元活性迅速（在毫秒内），并且可逆地减慢成人的谷氨酸摄取 大脑皮层。这种减慢的延长神经元NMDA反应，与细胞外延长一致 谷氨酸动力学，高度依赖于刺激的频率和持续时间。此外， 谷氨酸清除率可以通过神经元活性具有突触特异性来调节，即使在单个 星形胶质细胞。我们认为，这可能会对神经传递，外鼻间受体产生重要影响 激活和突触可塑性。基于这一发现，我们假设神经元活性会诱导 微域级变化星形胶质膜电位（VM），该电位（VM）局部调节EAAT功能。 Glt1是 成人前脑中主要的星形胶质细胞EAT，大量表达，并确保谷氨酸在 细胞外空间被EAAT结合迅速隔离。一旦绑定到Eaats， 谷氨酸进入星形胶质细胞既是钠驱动的，又依赖电压。在正常情况下，星形胶质细胞 由于对钾的渗透性高，因此被超极化（-80 mV）。但是，神经元活动增加 细胞外钾，[K+] E和星形胶质细胞VM对[K+] E变化特别敏感。因此，它是 神经元活性可以通过去极化星形胶质细胞来改变EAAT功能的原因。星形细胞VM的变化可能 在EAAT浓缩的精细星形胶质细胞过程中特别相关，细胞内小 与SOMA相比，体积可能会放大VM的变化。我们还将探索替代机制 包括通过[K+] e的增加对EAAT的电压调制。测试我们的主要挑战 然而，假设由于低膜电阻而无法在远端过程下监测星形胶质细胞VM 和过程形态。克服这一挑战很重要，因为星形胶质细胞远端过程是 突触相互作用和EAATS定位的位点。为了检测星形胶质细胞VM的远端变化，我们 使用遗传编码的电压指示器（GEVI）开发了一种在星形胶质细胞过程中图像VM的方法 成像。利用星形胶质细胞和神经元电生理记录，星形胶质细胞VM的光学遗传操作， 和星形胶质细胞膜电位的GEVI成像，我们产生了支持我们的初步数据 假设EAAT功能可以通过活性引起的星形胶质细胞VM的变化来调节。