Imaging astrocytic calcium dynamics in freely moving mice

描述（由申请人提供）：星形胶质细胞在控制哺乳动物行为中的作用仍然知之甚少，但除了在组织维持中的关键作用外，星形胶质细胞对神经发育和可塑性也很重要，并可调节神经元活性。事实上，星形胶质细胞和神经元很好地准备相互作用，星形胶质细胞能够调节神经网络的活动，并与一些形式的神经活动，促进动态的Ca 2+兴奋星形胶质细胞。这种相互作用对哺乳动物行为的意义仍然是个谜。有一种非常真实的--但在很大程度上还未被探索的--可能性， 星形胶质细胞及其Ca 2+动力学可能与认知和行为的计算过程密切相关。这个重要的问题仍然在很大程度上未经审查，由于普遍缺乏观察行为动物星形胶质细胞Ca 2+兴奋的工具。我们将通过创建用于在自由移动的小鼠中成像星形胶质细胞Ca 2+动态的工具来解决这一未满足的需求。 我们将结合联合收割机4最近开发的光学成像技术，这一起提供了第一次有机会跟踪astrocyticCa 2+动力学在数百个细胞在多个星期的活脑。我们将专注于海马的CA 1区，但在行为小鼠中长时间跟踪大量星形胶质细胞的能力将适用于各种各样的大脑区域，行为任务和认知形式。在开发我们的方法后，我们将通过检查CA 1星形胶质细胞动力学如何与海马空间认知和记忆形式相关的具体问题来验证和说明其效力。海马CA 1区对许多类型的空间认知至关重要，因此这个问题提供了第一个机会来研究星形胶质细胞如何有助于认知和行为的大脑计算。我们的四个目标是：目标1：建立成像的工具，在数百个独立的星形胶质细胞在自由行为的小鼠钙离子动力学。目的2：建立和验证CA 1星形胶质细胞Ca 2+动力学的延时成像，持续数周，并在行为小鼠的数百个细胞中进行。目标3：定量评估CA 1星形胶质细胞的Ca 2+动力学在60天内执行空间任务的动物中编码空间信息的程度。在小鼠反复访问2个不同的，熟悉的空间领域超过60天，我们将测试：假设1：CA星形胶质细胞活性编码空间信息;假设2：特定形式的CA 1星形胶质细胞Ca 2+活性与记忆回忆空间任务。假设3：CA 1星形胶质细胞活性的长期稳定性与长期空间记忆任务的表现相关。无论数据的结果如何，测试这些想法的能力都将是开创性的。目标4（资源共享）：在第4年和第5年，我们将每年为星形胶质细胞生物学家举办3次培训课程，让他们来到斯坦福大学学习我们的成像方法。如果成功，我们的工作可能会对多个领域产生显着影响，包括系统/认知神经科学和健康和疾病中的星形胶质细胞生物学研究。

DESCRIPTION (provided by applicant): Astrocytes' roles in the control of mammalian behavior remain poorly understood, but in addition to key roles in tissue maintenance astrocytes are important for neural development and plasticity and can modulate neuronal activity. Indeed, astrocytes and neurons are well poised to interact reciprocally, with astrocytes capable of modulating neural network activity, and with some forms of neural activity prompting dynamical Ca2+ excitation in astrocytes. The significance for mammalian behavior of such reciprocal interactions remains mysterious. There is the very real - but largely unexplored - possibility that astrocytes and their Ca2+ dynamics may be critically involved in the computational processes that underlie cognition and behavior. This vital issue remains largely unexamined due to a general lack of tools for observing astrocytic Ca2+ excitation in behaving animals. We will address this unmet need by creating tools for imaging astrocyte Ca2+ dynamics in freely moving mice. We will combine 4 recently developed optical imaging techniques, which together afford the first chance to track astroctytic Ca2+ dynamics in hundreds of cells over multiple weeks in the live brain. We will focus upon the CA1 area of hippocampus, but the capacity to track large populations of astrocytes over long time periods in behaving mice will be applicable to a wide variety of brain areas, behavioral tasks, and forms of cognition. After developing our approach, we will validate and illustrate its potency by examining the specific question of how do CA1 astrocytic dynamics relate to hippocampal forms of spatial cognition and memory? Hippocampal area CA1 is crucial for many types of spatial cognition, so this question poses a first opportunity to examine how astrocytes may contribute to brain computations underlying cognition and behavior. Our four aims are: Aim 1: Establish tools for imaging Ca2+ dynamics in hundreds of individual astrocytes in freely behaving mice. Aim 2: Establish and validate time-lapse imaging of CA1 astrocyte Ca2+ dynamics over weeks and across hundreds of individual cells in behaving mice. Aim 3: Quantitatively assess the degree to which CA1 astrocytes' Ca2+ dynamics encode spatial information in animals performing a spatial task over 60 days. In mice repeatedly visiting 2 distinct, familiar spatial arenas over 60 days we will test: Hypothesis 1: CA astrocyte activity encodes spatial information; Hypothesis 2: Specific forms of CA1 astrocyte Ca2+ activity are associated with memory recall in a spatial task. Hypothesis 3: The long-term stability of CA1 astrocyte activity correlates with performance on a long-term spatial memory task. Regardless of the data's outcome, the ability to test these ideas will be groundbreaking. Aim 4 (Resource Sharing): In years 4 and 5 we will hold 3 training sessions per year for astrocyte biologists to come to Stanford to learn our imaging methods firsthand. If successful, our work could have notable impact on multiple fields, including systems/cognitive neuroscience and the study of astroglial biology in both health and disease.