AFFERENT REGULATION OF CHOLINERGIC FOREBRAIN NEURONS

描述（由申请人提供）：本研究计划的主要目标是了解基底前脑（BF）胆碱能（BFC）和非胆碱能神经元是如何组织调节特定皮质区域的。尽管它参与皮层激活、注意力和记忆，但由于该区域的解剖学复杂性，其功能细节尚未得到很好的理解。阿尔茨海默病及相关痴呆患者皮质乙酰胆碱明显减少，BF胆碱能神经元出现病理改变。因此，对其功能组织的全面了解是必要的。本应用的中心假设是胆碱能神经元在BF（“细胞簇”）中构成局部集合，通过局部分支和/或共同输入及其对皮质区域的投影，为分布式功能网络提供神经基础，以选择性地调节认知过程。我们将在4个相互关联的特定目标中验证这一假设，使用传统和单突触病毒追踪，大规模网络的计算分析，BF神经元的体外膜片clamp记录，以及通过光遗传刺激确定的BF胆碱能神经元对自由运动大鼠皮质网络活动的高分辨率监测。在Aim 1中，我们将使用传统的逆行追踪技术建立一个相对完整的数据库，该数据库具有200 μ m分辨率的BF胆碱能和非胆碱能神经元。胆碱能簇将在由此产生的“数据库”中定义，并在簇体积中确定投射细胞种群的显著关联。在目标2中，我们将验证清醒行为大鼠BFC系统特定组织的功能意义。利用新开发的多阵列硅探针植入ChAT-Cre大鼠的两个特定皮质区域，光辅助摄动各种胆碱能细胞群，

DESCRIPTION (provided by applicant): The broad objective of this research program is to understand how basal forebrain (BF) cholinergic (BFC) and non-cholinergic neurons are organized to modulate specific cortical regions. Despite its involvement in cortical activation, attention, and memory, the functional details of the BF are not well understood due to the anatomical complexity of the region. Patients with Alzheimer's disease and related dementias have a significant decrease of acetylcholine in the cortex and show pathological changes in cholinergic neurons in the BF. Thus, a complete understanding of its functional organization is warranted. The central hypothesis of this application is that cholinergic neurons constitute local ensembles in the BF ('cell clusters') that via local collaterals and/or common inputs with their projections to cortical areas provide the neural basis of a distributed functional network to selectively modulate cognitive processes. We will test this hypothesis in 4 interrelated Specific Aims using traditional and monosynaptic viral tracing, computational analysis of large-scale networks, in vitro patch-clamp recording of BF neurons, and high-resolution monitoring of cortical network activity in freely-moving rats with optogenetic stimulation of defined BF cholinergic neurons. In Aim 1 we will build up a relatively complete database with 200 �m resolution of mapped BF cholinergic and non-cholinergic neurons using conventional retrograde tracing techniques. Cholinergic clusters will be defined in the resulting 'database' and in the cluster volume significant association of projection cell populations will be determined. In Aim 2 we will validate of the functional significance of the specific organization of the BFC system in wake-behaving rats. With newly developed multi-array silicon probes implanted into two specific cortical areas and light-assisted perturbation of various cholinergic cell groups in ChAT-Cre rats, in which cholinergic cells were transfected to express channelrhodopsin (ChR2), we will determine the emerging cholinergic ensembles in the BF and their effect on the functional connectivity of various large-scale cortical networks during various brain states. In Aim 3 we will define the input to cholinergic neurons in various subdivisions of the BF using commercially available Cre-dependent AAV "helper viruses" (AAV-EF1a-FLEX- TVAmCherry and AAV-CA-FLEX-RG) and a replication deficient rabies vector (RV: EnvA G-deleted Rabies- eGFP). In Aim 4 we will determine, using in vitro patch clamp recording and retrograde tracing in ChAT-cre X ChAT-eGFP crossbred mice, how does the system of early (EF) and late firing (LF) cholinergic neurons and local cholinergic axon arborizations fit into the global organization of the BFC system. The in vivo large-scale, high-density recording design that is built on a realistic forebran model will lead to substantially improved animal models for addressing function in behavioral studies. Concomitantly, it will facilitate the understanding of the aberrant processing in basalo-cortical networks and may help the development of new treatment strategies to ameliorate the cognitive symptoms in Alzheimer's and related disorders.