Optogenetic resource for studying cerebral cortex network function

描述（由申请人提供）：大脑皮层是由神经化学定义的细胞亚群组成的复杂网络。了解不同的神经群如何相互作用以产生动态电生理振荡，这些振荡是清醒和睡眠期间大脑皮层网络的特征，是神经科学领域的一个紧迫挑战。在这种情况下，对新的实验资源来解决假设的需求是一个关键的未满足的需求。我们开发了一种新的转基因小鼠系（NPY-ChR2- eYFP），其中神经肽Y （NPY）启动子驱动光敏阳离子通道Channelrhodopsin2 （ChR2）和标记蛋白增强黄色荧光蛋白（eYFP）的表达。大脑皮层中的npy阳性细胞亚群（睡眠活跃神经元；SANs）被认为是大脑皮层网络慢波活动、可塑性和血流的睡眠依赖性变化的调节剂。我们的初步数据表明，转基因在大脑中表达，并且光遗传学操作大脑皮层触发转基因表达小鼠脑电图中缓慢活动的增加。这些数据表明，光遗传学策略来操纵这些动物的大脑皮层神经元网络特性是可行的。这项工作的总体目标是开发一套方案，其中NPY-ChR2-eYFP小鼠可以用来描述大脑皮层表达npy的中间神经元群体的功能，特别是SAN群体，在产生大脑皮层电生理事件。我们将从两个方面来实现这一目标。在Aim 1中，我们将使用免疫组织化学来验证NPY-ChR2-eYFP构建体将转基因表达靶向于大脑皮层的npy阳性细胞和SANs。在目标2中，我们将优化光遗传刺激方案，以操纵体内靶细胞群的活性。总的来说，这些研究将产生一套方案，以推进我们对npy阳性人群在调节皮层电生理振荡中的功能的认识。此外，转基因小鼠系及其实验方案将成为其他研究领域的公共资源，如神经血管耦合、应激反应和喂养的神经调节以及中风的病理生理学。

DESCRIPTION (provided by applicant): The cerebral cortex is a complex network of neurochemically-defined subpopulations of cells. Understanding how distinct neural populations interact to generate the dynamic electrophysiological oscillations that characterize the cerebral cortical network during wake and sleep is a pressing challenge to the field of neuroscience. There is a critical unmet need for novel experimental resources to address hypotheses in this context. We have developed a novel transgenic mouse line (NPY-ChR2- eYFP) in which the neuropeptide Y (NPY) promoter drives expression of the light-sensitive cation channel Channelrhodopsin2 (ChR2) and the marker protein enhanced yellow fluorescent protein (eYFP). A subpopulation of Npy-positive cells in the cerebral cortex (sleep-active neurons; SANs) is a putative regulator of sleep-dependent changes in cerebral cortex network slow wave activity, plasticity and blood flow. Our preliminary data demonstrate that the transgene is expressed in the brain, and that optogenetic manipulation of the cerebral cortex triggers an increase in slow activity in the electroencephalogram of transgene-expressing mice. These data demonstrate that an optogenetic strategy to manipulate cerebral cortical neuronal network properties in these animals is feasible. The overarching goal of this work is to develop a set of protocols in which NPY-ChR2-eYFP mice can be used to delineate the functions of the cerebral cortical NPY-expressing interneuron population generally, and the SAN population specifically, in generating cerebral cortical electrophysiological events. We will achieve this goal by pursuing two aims. In Aim 1, we will use immunohistochemistry to verify that the NPY-ChR2-eYFP construct targets transgene expression to NPY-positive cells and SANs in the cerebral cortex. In Aim 2, we will optimize optogenetic stimulation protocols for manipulating the activity of the target cell population in vivo. Collectively, these studies will yield a set of protocols to advance our knowledge of the function of the NPY-positive population in regulating cortical electrophysiological oscillations. Additionally, the transgenic mouse line and protocols for experimentation on this line will become a public resource applicable in other areas of research such as neurovascular coupling, neural regulation of stress responses and feeding, and the pathophysiology of stroke.