Astrocytic Regulation of Neuronal Excitability in Vivo

描述（由申请人提供）：虽然很明显，星形胶质细胞表现出能够调节体内神经元活动的特性，但我们认为有必要开发模型系统，以便研究星形胶质细胞在神经生理学和行为中的作用。为了实现这一目标，我们开发了条件基因敲除系统，使我们能够检查特定星形细胞基因产物在发育和成熟小鼠中的作用。我们计划使用星形胶质细胞特异性诱导敲除（i-cKO）小鼠来验证假设，即在神经元活动期间，星形胶质细胞通过Kir4.1通道摄取K+，并通过基于connexin43的间隙连接产生的星形胶质细胞合胞体将该离子分散。我们制备了一种可诱导的Cre-loxP系统，使我们能够灭活95%以上的发育和发育后星形胶质细胞中的“floxed”基因。使用海马脑切片进行实验，以确定星形胶质细胞特异性，Kir4.1或Cx43的i-cKO是否影响K+稳态，和/或原位神经元兴奋性，以及动物行为。我们还制备了非诱导型Kir4.1和Cx43 cKO小鼠，它们表现出惊人的细胞和行为变化。非诱导型Kir4.1 cKO小鼠在整个大脑和脊髓中表现出异常的髓鞘形成。我们将验证在非诱导型Kir4.1 cKO小鼠中观察到的异常髓鞘形成的假设，该假设是由于少突胶质细胞发育缺陷或成熟少突胶质细胞维持髓鞘的能力缺陷引起的。在过去的几年里，我们已经开发了许多分子工具，使我们能够进行星形细胞特异性诱导基因敲除。这些分子工具，结合在本实验室和其他实验室中产生的floxed小鼠，将在研究星形胶质细胞在从细胞水平的突触可塑性到行为水平的学习和记忆等过程中的功能方面非常有用。我们建议建立一个网站，描述我们实验室开发的遗传模型，使其他人能够充分利用这些试剂来揭示星形胶质细胞在神经生理学和行为中的作用。

DESCRIPTION (provided by applicant): While it is clear that astrocytes exhibit properties that could enable them to modulate neuronal activity in vivo, we believe that it is essential to develop model systems whereby the role of astrocytes in neurophysiology and behavior can be examined. To accomplish this goal, we have developed conditional gene knockout systems that enable us to examine the role of specific astrocytic gene products in developing and mature mice. We plan to use astrocyte-specific, inducible knockout (i-cKO) mice to test the hypothesis that during neuronal activity, astrocytes take up K+ through Kir4.1 channels and disperse this ion through an astrocytic syncytium created by connexin43-based gap junctions. We have prepared an inducible Cre-loxP system that enables us to inactivate "floxed" genes in greater than 95% of astrocytes during development and post-developmentally. Experiments using hippocampal brain slices will be carried out to determine if an astrocyte-specific, i-cKO of Kir4.1 or Cx43 affects K+ homeostasis, and/or neuronal excitability in situ, and animal behavior. We have also prepared non-inducible Kir4.1 and Cx43 cKO mice that exhibit striking cellular and behavioral changes. Non-inducible Kir4.1 cKO mice exhibit aberrant myelination throughout the brain and spinal cord. We will test the hypothesis that the abnormal myelination observed in non-inducible Kir4.1 cKO mice arises from a defect in either the development of oligodendrocytes or the ability of mature oligodendrocytes to maintain myelin. Over the past several years we have developed a number of molecular tools that enable us to carry out astrocyte-specific inducible gene knockouts. These molecular tools, combined with floxed mice being generated in this and other laboratories, will be extremely useful in studying the function of astrocytes in processes ranging from synaptic plasticity at the cellular level to learning and memory at the behavioral level. We propose to develop a website describing the genetic models developed in our laboratory to enable others to take full advantage of these reagents in unraveling the role of astrocytes in neurophysiology and behavior.