Voltage imaging of astrocyte-neuron interactions

星形胶质细胞-神经元相互作用的电压成像

基本信息

批准号：
10630180
负责人：
Chris G Dulla
金额：
$ 59.45万
依托单位：
TUFTS UNIVERSITY BOSTON
依托单位国家：
美国
项目类别：
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/10630180
关键词：
Acute Address Adult Affect Amino Acid Transporter Area Astrocytes Ataxia Binding Brain Buffers Cerebral cortex Characteristics Data Disease Distal Electrophysiology (science)Ensure Epilepsy Excitatory Amino Acid Transporter 1 Excitatory Amino Acid Transporter 2 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 Rodent Shapes Signal Transduction Site Slice Sodium Specificity Synapses Synaptic Receptors Synaptic Transmission Synaptic plasticity Testing designer receptors exclusively activated by designer drugs extracellular glutamatergic signaling in vivo insight millisecond neuronal cell body neuronal patterning neurotransmission novel optogenetics pharmacologic response sensor spatiotemporal tool uptake voltage

项目摘要

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.

项目概要星形胶质细胞在神经元释放后从细胞外间隙去除兴奋性神经递质谷氨酸通过钠驱动、电压依赖性兴奋性氨基酸转运蛋白（EAAT）进行活性。强健的谷氨酸盐 EAAT 的摄取确保了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动迅速（在几毫秒内）并可逆地减慢成人对谷氨酸的吸收大脑皮层。这种减慢延长了神经元 NMDA 反应，与延长的细胞外反应一致谷氨酸动力学，并且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具有突触特异性的神经元活动来调节，甚至在单个星形胶质细胞。我们相信这可能对神经传递、突触外受体产生重要影响激活和突触可塑性。基于这一发现，我们假设神经元活动诱导星形胶质细胞膜电位 (Vm) 的微域水平变化可局部调节 EAAT 功能。 GLT1 是成人前脑中主要的星形胶质细胞 EAAT 大量表达，并确保谷氨酸细胞外空间被 EAAT 结合迅速隔离。一旦绑定到 EAAT，运输谷氨酸进入星形胶质细胞既是钠驱动的又是电压依赖性的。正常情况下，星形胶质细胞由于其对钾的高渗透性而超极化（-80 mV）。然而，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，可以说原因是神经元活动可以通过去极化星形胶质细胞来改变 EAAT 功能。星形胶质细胞 Vm 的变化可能与精细星形胶质细胞过程尤其相关，其中 EAAT 集中，并且小的细胞内与体细胞相比，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 进行与电压无关的调节。测试我们的一个重大挑战然而，假设是由于膜阻力低而无法监测远端星形胶质细胞的 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突是突触相互作用和 EAAT 定位的位点。为了检测星形胶质细胞Vm的远端变化，我们开发了一种使用基因编码电压指示器（GEVI）对星形胶质细胞过程中的 Vm 进行成像的方法成像。利用星形胶质细胞和神经元电生理记录、星形胶质细胞Vm的光遗传学操作，和星形胶质细胞膜电位的 GEVI 成像，我们已经生成了支持我们的初步数据假设 EAAT 功能可以通过活动诱导的星形胶质细胞 Vm 变化来调节。