Optogenetic Analysis of Neostriatal Circuits Engaged by Cholinergic Interneurons

描述（由申请人提供）：新纹状体胆碱能（ChAT）中间神经元通过编码外部事件的显著性和强化值，在选择和获得适应性行为动作中发挥重要作用。这一信息表示在精确的时间结构的同步多相人口反应的行为显着的刺激的介绍。一个重要的问题是要了解这些瞬态信号的ChAT interneurons检测和解码的neostriatal网络。我们的假设，制定的基础上的初步实验，是ChAT中间神经元控制纹状体电路的活动，使用烟碱受体介导的机制，激活多个平行的GABA能机制，引起动力学上不同的独立抑制反应的多刺投射神经元（SPN）和其他纹状体神经元。这些反应部分源于多种尚未鉴定类型的GABA能中间神经元的直接激活，所述GABA能中间神经元不同于含有快速尖峰（FS）和NPY表达的中间神经元的小清蛋白（PV），还可能涉及突触前烟碱促进GABA从轴突终末释放，并一起向SPN、ChAT和其他中间神经元提供选择性的细胞类型特异性输入。使用体外光遗传学抑制，我们还再现了在行为动物中最常见的ChAT中间神经元的群体响应，其由发射暂停随后是弱同步反弹兴奋组成，并证明了这种行为直接相关的活性模式也参与了上述抑制机制。由于这些反应足以抑制大量SPN中动作电位的产生，因此它们可能对基底神经节的功能产生显著影响并调节持续的行为。拟议的实验旨在了解这些GABA能电路和机制的功能组织，并定量表征ChAT中间神经元在体外和体内对SPN活性的强化相关群体反应的影响。光遗传学方法将用于控制双转基因动物中ChAT和其他中间神经元的活性，从而允许遗传靶向和鉴定特定细胞类型。ChAT中间神经元的双突触回路将通过鉴定由ChAT中间神经元激活的GABA能中间神经元，然后通过选择性光遗传学激活特定类别的中间神经元群体并通过检查各种中间神经元类型的光遗传学抑制对双突触引发的IPSC的影响来确定单个突触前细胞类型对SPN和其他神经元中各种GABA能反应组分的贡献来分析chat interneurons。最后，我们将在体内和体外的SPN的ChAT中间神经元的光遗传学再现暂停-兴奋反应的影响进行定量表征。这些实验将描述一个强大的新纹状体电路机制的功能组织，因此将显着推进对基底神经节功能的理解。 公共卫生相关性：许多重要的神经和精神障碍，包括帕金森病和亨廷顿病、精神分裂症、图雷特综合征、强迫症，都是由新纹状体的病理状况引起的。其中几个直接或间接地连接到一个主要的神经调节系统的胆碱能interneurons的这种结构，但至关重要的问题，这些interneurons的功能仍然没有答案。该提案将阐明一种新发现的回路机制的组织和功能作用，该机制负责介导这些神经元的强化相关群体反应，在病理条件下可能直接导致与这些疾病相关的特定行为异常。

DESCRIPTION (provided by applicant): Neostriatal cholinergic (ChAT) interneurons play an important role in the selection and acquisition of adaptive behavioral actions by encoding the salience and reinforcement value of external events. This information is represented in the precise temporal structure of synchronous multiphasic population responses given to the presentation of behaviorally significant stimuli. An important problem is to understand how these transient signals of ChAT interneurons are detected and decoded in the neostriatal network. Our hypothesis, formulated on the basis of preliminary experiments, is that ChAT interneurons control the activity of the striatal circuitry using nicotinic receptor mediated mechanisms to activate multiple parallel GABAergic mechanisms that elicit kinetically distinct independent inhibitory responses in the spiny projection neurons (SPNs) and in other striatal neurons. These responses originate in part from direct activation of multiple as yet unidentified types of GABAergic interneurons that are distinct from the parvalbumin (PV) containing fast spiking (FS) and NPY expressing interneurons, possibly also involve presynaptic nicotinic facilitation of GABA release from axon terminals and together provide selective, cell type specific inputs to SPNs, ChAT and other interneurons. Using in vitro optogenetic inhibition we also reproduced the population response of ChAT interneurons most commonly observed in behaving animals consisting of a pause of firing followed by weakly synchronous rebound-excitation and demonstrated that this behaviorally directly relevant pattern of activity also engages the inhibitory mechanisms described above. Since these responses are sufficient to inhibit action potential generation in large populations of SPNs they are likely to exert significant effects on the functioning of the basal ganglia and regulate ongoing behavior. The proposed experiments aim at understanding the functional organization of these GABAergic circuits and mechanisms and to quantitatively characterize the effects of reinforcement related population responses of ChAT interneurons on the activity of SPNs in vitro and in vivo. Optogenetic methods will be used to control the activity of ChAT and other interneurons in double transgenic animals allowing the genetic targeting and identification of specific cell types. The disynaptic circuits of ChAT interneurons will be analyzed by identifying the GABAergic interneurons activated by ChAT interneurons and then determining the contribution of individual presynaptic cell types to the various GABAergic response components in SPNs and other neurons by selective optogenetic activation of populations of specific classes of interneurons and by examining the effects of optogenetic inhibition of various interneuron types on IPSCs elicited disynaptically by ChAT interneurons. Finally, we will quantitatively characterize the effects of optogenetically reproduced pause-excitation responses of ChAT interneurons on SPNs in vivo and in vitro. These experiments will describe the functional organization of a powerful novel circuit mechanism of the neostriatum and therefore will significantly advance the understanding of the functioning of the basal ganglia. PUBLIC HEALTH RELEVANCE: Numerous important neurological and psychiatric disorders including Parkinson's and Huntington's diseases, schizophrenia, Tourette's syndrome, obsessive compulsive disorder arise from pathological conditions of the neostriatum. Several of these are directly or indirectly linked to a major neuromodultory system the cholinergic interneurons of this structure, yet critically important issues about the functional role of these interneurons remains unanswered. The proposal will elucidate the organization and functional role of a newly discovered circuit mechanism that is responsible for mediating the reinforcement related population responses of these neurons which under pathological conditions may directly contribute to specific behavioral abnormalities associated with these disorders.