The role of glial cells in the development and function of retinal circuits

描述(申请人提供)：越来越多的证据表明，神经元和神经胶质细胞参与影响神经回路功能和发育的双向信号传递。神经胶质细胞可以通过控制突触外间隙中神经递质的数量来影响神经元的兴奋性，并可以直接调节突触传递。此外，神经胶质细胞是控制神经元代谢的神经血管相互作用的中心调节者。为了更好地了解神经胶质细胞的功能，有必要了解神经元和神经胶质细胞之间的密切相互作用是如何在发育过程中建立的，以及这对神经回路稳定的影响。这项建议描述了一种方法来阐明神经元-胶质细胞信号在小鼠视网膜回路发展中的作用。拟议的研究将为一项长期研究计划提供基础，该计划致力于探索神经元、胶质细胞和血管之间的准确信号是如何建立和维持的。胶质细胞是视网膜中最后一种增殖的细胞类型，在一段称为视网膜波的自发相关活动期间，它必须整合到现有的未成熟回路中。在睁开眼睛前不久，视网膜波由谷氨酸能信号介导，并被认为通过谷氨酸溢出传播到突触外空间。M？ler细胞在谷氨酸能波期间在形态和功能上成熟，并在波停止和视觉开始时稳定视网膜回路。在这项提议中，探索了两种形式的神经元-神经胶质细胞信号。在第一个目标中，我们将 确定神经胶质细胞转运蛋白功能的成熟如何影响神经元回路的发育。在第二个目标中，我们将研究自发和诱发的神经元活动如何影响神经胶质细胞钙信号。在目标1中，我们将使用光学和电生理方法来描述M？ler细胞转运蛋白功能的成熟。谷氨酸光学传感器可以直接观察到谷氨酸的水平，当谷氨酸结合时，这种传感器会增加荧光。我将使用这些传感器来监测从视网膜波到视觉的过渡过程中突触外的谷氨酸，并确定谷氨酸是否溢出到M？ler细胞。M？ler转运蛋白将被药物阻断，神经元群体的双光子钙成像将被用来评估对视网膜回路的影响。在目标2中，我们将使用在M？ller细胞中表达钙指示剂GCaMP3的转基因小鼠来研究神经活动是否在整个视网膜发育过程中驱动M？ler钙瞬变。了解神经胶质细胞如何在整合到成熟的回路中时改变神经元功能，将有助于对神经元-胶质细胞信号的细胞和分子机制有基本的了解。神经元-神经胶质细胞相互作用的维持对神经系统的健康至关重要，尤其是神经胶质转运蛋白的功能已被临床证明。 意义。

DESCRIPTION (provided by applicant): Mounting evidence suggests that neurons and glia engage in bi-directional signaling that affects the function and development of neural circuits. Glia can influence the excitability of neurons, by controlling the amount of neurotransmitters in the extrasynaptic space, and can directly modulate synaptic transmission. In addition, glia are the central regulators of the neurovascular interactions that control neuronal metabolism. To better understand glial function, it is essential to unravel how the close interaction between neurons and glia is established during development and the effect this has on the stabilization of neural circuits. This proposal describes an approach to elucidate the role of neuron-glia signaling in the development of murine retinal circuits. The proposed investigations will provide the basis for a long-term research program dedicated to exploring how the precise signaling between neurons, glia, and blood vessels is established and maintained. M¿ller glia are the last cell type to proliferate in the retina, and must integrate into an existing immature circuit duringa period of spontaneous correlated activity called retinal waves. Shortly before eye opening, retinal waves are mediated by glutamatergic signaling and are thought to propagate by glutamate spillover into the extrasynaptic space. M¿ller cells mature in morphology and function during glutamatergic waves, and may stabilize retinal circuits when waves cease and vision begins. Two forms of neuron-glia signaling are explored in this proposal. In the first aim, we will determine how the maturation of glial cell transporter function affects the development of neuronal circuits. In the second aim, we will investigate how spontaneous and evoked neuronal activity influences glial calcium signaling. In aim 1 we will use optical and electrophysiological methods to describe the maturation of M¿ller cell transporter function. Levels of glutamate can be directly observed with glutamate optical sensors that increase in fluorescence when glutamate binds. I will use these sensors to monitor extrasynaptic glutamate during the transition from retinal waves to vision, and to ascertain whether or not glutamate spillover reaches M¿ller cells. M¿ller transporters will be blocked pharmacologically and two-photon calcium imaging of neuronal populations will be used to assess the effect on retinal circuits. In aim 2 we will use transgenic mice expressing the calcium indicator GCaMP3 in M¿ller cells to investigate whether neural activity drives M¿ller calcium transients throughout retinal development. An understanding of how glial cells change neuronal function as they integrate into maturing circuits will lead to fundamental insights about the cellular and molecular mechanisms of neuron-glia signaling. The maintenance of neuron-glia interactions is vital for the health of the nervous system, and in particular, the glial transporter function has proven clinical significance.