Astrocyte-neuron interactions in visual cortex circuits

在初级视觉皮质(V1)，精确的时空神经元反应是视觉的基础 正在处理。虽然神经元在视觉处理中的作用已经被很好地研究了，但非神经元细胞的作用， 尤其是皮质突触和环路中的星形胶质细胞，目前仍知之甚少。皮质星形胶质细胞接触 并包裹几乎所有的兴奋性突触，创建由突触前输入组成的离散功能单位， 突触后棘突和星形胶质细胞突起。星形胶质细胞的一个重要功能是主动摄取谷氨酸。 从突触间隙通过转运蛋白，特别是GLT1。我们认为星形胶质细胞在根本上 通过GLT1激活V1通路，积极塑造突触和神经元的反应轮廓。焦点钙离子瞬变 最近在星形胶质细胞突起中被证明可能与突触谷氨酸摄取有关， 突触传递被证明主动地将星形细胞GLT1招募到突触活动的部位。新奇的高- 分辨率成像技术，以及细胞特异性标记、新的光学探针和遗传学 具有特定时间和空间控制蛋白质表达的工程小鼠，使我们能够分析 清醒小鼠体内史无前例分辨率的星形胶质细胞和神经元活动之间的串扰。我们的目标是 利用V1神经元视觉输入的精细组织来研究钙离子的相互作用 星形胶质细胞突起中微区、线粒体和谷氨酸转运体的功能贡献 视觉加工过程中星形胶质细胞转运蛋白对神经元突触和回路的影响 谷氨酸转运对V1环路发育和可塑性的影响。在目标1中，我们将检测星形胶质细胞 微域Ca~(2+)对视觉刺激的反应，包括定向特定的栅格和复杂的自然 图像，它们与线粒体的关系，以及GLT1的遗传或药物减少如何影响 星形胶质细胞和细胞特异性神经元反应的特异性和可靠性。在目标2中，我们将确定 单树突棘与邻近星形细胞突起功能关系的同步研究 视觉刺激过程中星形胶质细胞和神经元的双波长成像。我们还将确定GLT1如何 减少影响星形细胞的突起和神经元的脊椎反应和结构。在目标3中，我们将确定 GLT1在星形胶质细胞反应和视皮层回路的发育和可塑性中的作用。我们会 研究正常情况下GLT1的生殖系减少如何改变神经元和星形胶质细胞的微域反应 单眼剥夺后的发育和单眼剥夺，以及定向选择性和 定向偏好的双目匹配。我们的首要目标是批判性地检验这一假设 星形胶质细胞及其转运体是神经元在功能和发育过程中不可或缺的功能伙伴 大脑皮层环路。因此，对一系列神经发育的正常和异常功能的理解 神经退行性疾病将需要纳入星形胶质细胞的作用。

In primary visual cortex (V1), precise spatiotemporal neuronal responses are known to underlie visual processing. Though neuronal roles in visual processing have been well studied, the role of non-neuronal cells, particularly astrocytes, in cortical synapses and circuits remains poorly understood. Cortical astrocytes contact and ensheathe nearly all excitatory synapses, creating discrete functional units consisting of a presynaptic input, a postsynaptic spine and an astrocyte process. A crucial function of astrocytes is the active uptake of glutamate from the synaptic cleft via transporters, particularly GLT1. We propose that astrocytes contribute fundamentally to V1 circuits via GLT1 activity, actively shaping synaptic and neuronal response profiles. Focal Ca2+ transients potentially related to synaptic glutamate uptake have recently been demonstrated within astrocyte processes, and synaptic transmission shown to actively recruit astrocytic GLT1 to sites of synaptic activity. Novel high- resolution imaging techniques, together with cell-specific markers, new optical probes, and genetically engineered mice with specific temporal and spatial control of protein expression, enable us to analyze the crosstalk between astrocyte and neuronal activity at unprecedented resolution in awake mice in vivo. We aim to take advantage of the exquisite organization of visual inputs to V1 neurons to examine the interaction of Ca2+ microdomains, mitochondria and glutamate transporters in astrocyte processes, the functional contribution of astrocyte transporters to neuronal synapses and circuits during visual processing, and the impact of altered glutamate transport on the development and plasticity of V1 circuits. In Aim 1, we will examine astrocyte microdomain Ca2+ responses to visual stimuli, including orientation-specific gratings and complex natural images, their relationship to mitochondria, and how genetic or pharmacological reduction of GLT1 impacts the specificity and reliability of astrocyte and cell-specific neuronal responses. In Aim 2, we will determine the functional relationship between single dendritic spines and adjacent astrocytic processes using simultaneous dual-wavelength imaging of astrocytes and neurons during visual stimulation. We will also determine how GLT1 reduction affects astrocytic process and neuronal spine responses and structures. In Aim 3, we will determine the role of GLT1 in the development and plasticity of astrocyte responses and visual cortex circuits. We will examine how germline reduction of GLT1 alters neuronal and astrocyte microdomain responses during normal development and following monocular deprivation, along with the sharpening of orientation selectivity and the binocular matching of orientation preference. Our overarching goal is to critically examine the hypothesis that astrocytes and their transporters are integral functional partners with neurons in the function and development of cortical circuits. As such, an understanding of normal and abnormal function in a host of neurodevelopmental and neurodegenerative disorders will require incorporating the role of astrocytes.