Voltage Imaging of Astrocyte-Neuron Interactions

抽象的。 星形胶质细胞控制神经传递，有助于建立强大的血脑屏障 (BBB)，并提供代谢支持 神经元活动。在阿尔茨海默病 (AD) 中，星形胶质细胞在 A 斑块附近变得活跃，有助于 神经炎症，并导致突触和回路异常。星形胶质细胞的作用 通过从细胞外空间去除谷氨酸来控制突触兴奋在 AD 中尤其令人感兴趣。 星形胶质细胞表达高水平的兴奋性氨基酸转运蛋白 (EAAT)，包括 GLT1 和 GLAST， 它们的外周过程结合并去除细胞外谷氨酸以限制突触兴奋。 EAAT，如 众所周知，星形胶质细胞蛋白以及其他星形胶质细胞蛋白在 AD 进展后期会减少。谷氨酸信号异常 谷氨酸受体美金刚的临床使用证明，被认为会导致 AD 进展 拮抗剂作为 AD 疗法，与 EAAT 密切相关，与 AD 的进展密切相关。最后，可溶性 A 抑制 EAAT 在神经元活动增强期间发挥作用。有趣的是，我们最近表明神经元活动 引起星形胶质细胞去极化，从而驱动 EAAT 功能的电压依赖性抑制，以增强 神经元激活。在母体 R01 中，我们使用星形胶质细胞表达的基因编码电压指标 （GEVI）光学量化 Vm 并表明神经元活动使外周星形胶质细胞过程去极化 （PAP）具有突触特异性。在此补充请求中，我们提供了正常老化过程中的初步数据 星形胶质细胞偶尔会失去 GLT1、GLAST 和 Kir4.1 的表达，并呈现出称为 非典型星形胶质细胞（AtAs）。重要的是，AtAs 不是反应性星形细胞增多症的一种形式。初步数据表明 AtAs 在 AD 的 APPNL-G-F 小鼠模型中更丰富且出现更早，并且在老年人中也可见到 大脑。在这里，我们将检验他们的假设，因为缺乏 Kir4.1 和 GLT-1 的 AtAs 在 APPNL-G-F 小鼠海马中，星形胶质细胞经历活动诱导的去极化增加，导致 过度抑制谷氨酸的吸收。这造成了一种情况，即增加的神经元活动可以 AD 中驱动协同电压依赖性和 A 介导的 EAAT 抑制。我们选择使用 APPNL-G-F 该模型由于其发病较早，与AtAs的正常发育重叠，没有APP过度表达。 使用 APPNL-G-F 小鼠，我们将确定：（SA1）“APPNL-G-F 小鼠的海马星形胶质细胞是否表现出增加” Vm 对神经元活动的反应？” (SA2)“谷氨酸摄取的活性依赖性抑制是否在 APPNL-G-F 小鼠的海马？”。活动诱导的 PAP 去极化增加和 A 升高都可以 改变 APPNL-G-F 小鼠中活动诱导的 EAAT 抑制。如果是这样，这将定位星形胶质细胞-神经元相互作用， K 和谷氨酸稳态是 AD 早期进展的关键因素，并会促进谷氨酸的增强 和钾的吸收以减少 AD 相关病理。完成后，我们将知道是否有新发现的变化 PAP Vm 中的值被夸大并导致 APPNL-G-F 小鼠中谷氨酸的异常兴奋。我们将蓄势待发 利用这些发现来增强钾和谷氨酸稳态，以防止 AD 中的病理兴奋。

ABSTRACT. Astrocytes control neurotransmission, contribute to a robust blood-brain-barrier (BBB), and metabolically support neuronal activity. In Alzheimer’s Disease (AD), astrocytes become reactive near A plaques, contribute to neuroinflammation, and contribute to synaptic and circuit abnormalities. The role that astrocytes play in controlling synaptic excitation by removing glutamate from the extracellular space is particularly intriguing in AD. Astrocytes express high levels of excitatory amino acid transporters (EAATs), including GLT1 and GLAST, in their peripheral processes that bind and remove extracellular glutamate to limit synaptic excitation. EAATs, as well as other astrocyte proteins, are known to be decreased late in AD progression. Aberrant glutamate signaling is thought to contribute to AD progression, as evidenced by the clinical use of memantine, a glutamate receptor antagonist, as an AD therapy, strongly implicating EAATs in the progression of AD. Finally, soluble A inhibits EAAT function during heightened neuronal activity. Interestingly, we recently showed that neuronal activity causes astrocyte depolarization, which drives voltage-dependent inhibition of EAAT function to enhance neuronal activation. In the parent R01, we use astrocyte-expressed genetically encoded voltage indicators (GEVIs) to optically quantify Vm and showed that neuronal activity depolarizes peripheral astrocyte processes (PAPs) with synapse-specificity. In this supplement request, we provide preliminary data that during normal aging astrocytes sporadically lose the expression of GLT1, GLAST, and Kir4.1 and take on a phenotype known as atypical astrocytes (AtAs). Importantly, AtAs are not a form of reactive astrocytosis. Preliminary data suggests AtAs are more abundant and occur earlier in the APPNL-G-F mouse model of AD and are seen in the aged human brain. Here we will test they hypothesis that because AtAs, which lack Kir4.1 and GLT-1, are abundant in the hippocampus of APPNL-G-F mice, astrocytes experience increased activity-induced depolarization, leading to exaggerated inhibition of glutamate uptake. This creates a situation in which increased neuronal activity could drive synergistic voltage-dependent and A-mediated EAAT inhibition in AD. We chose to use the APPNL-G-F model due to its early onset, overlapping with the normal development of AtAs, without APP overexpression. Using the APPNL-G-F mice, we will determine: (SA1) “Do hippocampal astrocytes in APPNL-G-F mice show increased Vm responses to neuronal activity?” and (SA2) “Is activity-dependent inhibition of glutamate uptake enhanced in the hippocampus of APPNL-G-F mice?“. Both increased activity-induced PAP depolarization and elevated A could alter activity-induced EAAT inhibition in APPNL-G-F mice. If so, this would position astrocyte-neuron interactions, K+ and glutamate homeostasis as key players in early AD progression and would motivate enhancing glutamate and K+ uptake to reduce AD-related pathology. When complete, we will know whether newly discovered changes in PAP Vm are exaggerated and contribute to aberrant glutamate excitation in APPNL-G-F mice. We will be poised to leverage these findings to enhance K+ and glutamate homeostasis to prevent pathological excitation in AD.