Cell Type Specific Tracing of Neocortical Circuits Using Viral Vectors

使用病毒载体对新皮质回路进行细胞类型特异性追踪

基本信息

批准号：
8469103
负责人：
David C Lyon
金额：
$ 19.23万
依托单位：
UNIVERSITY OF CALIFORNIA-IRVINE
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-05-01 至 2014-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8469103
关键词：
Brain Cells Characteristics Code Complement Computer software Dependovirus Development Effectiveness Equilibrium Felis catus Gene Targeting Genes Glycoproteins Goals Histology Immunohistochemistry Individual Infection Injection of therapeutic agent Label Lateral Location Maps Mediating Methods Microscopy Neocortex Neurons Neurotropism Pattern Population Process Proteins Rabies Rabies virus Radial Rattus Reporter Genes Role Signal Transduction Specificity Stimulus Subfamily lentivirinae Techniques Testing Time Tracer Viral Vector Virus Virus Diseases Visual Cortex Visual system structure Work area striata cell type excitatory neuron gamma-Aminobutyric Acid inhibitory neuron innovation insight neocortical neural circuit novel optical imaging orientation selectivity preference promoter rabies virus glycoprotein G receptor recombinant viral vector recombinant virus selective expression tool vector

项目摘要

DESCRIPTION (provided by applicant): The neocortex is comprised of a dense population of excitatory neurons that are balanced by a diverse and more sparsely distributed array of inhibitory neurons. As such, cortical inhibitory neurons provide a gain control that works under a myriad of conditions. In primary visual cortex (V1), for example, inhibition is essential for severl basic functional characteristics of individual neurons, including preferences for stimulus contrast, size, and orientation. While inhibitory neurons serve to control excitation it is not knon whether inhibitory and excitatory cell-types are mediated through the same set of cortical circuits, nor whether the functional selectivity of these circuits differ. Determining the origins f the neural circuits to these two major cell-types will provide tremendous insight into the basic mechanisms of cortical processing. However, in order to do so it is necessary to develop a technique for cell-type specific neuroanatomical tracing. The proposal here is to develop such a technique by taking advantage of the neurotropism of recombinant viral vectors (adeno associated virus and lentivirus) and selectivity of cell-type specific promoters (GAD-67 and ¿-CamKII) to deliver and allow for expression of two key genes (TVA and RabG). These genes will thereby be selectively expressed in either inhibitory or excitatory neocortical cells, but not both. The TVA and RabG proteins expressed in these specific cell types will allow for targeted viral infection and trans-complementation of a novel genetically modified and pseudo-typed rabies virus (EnvA-¿RabG) that acts as a monosynaptic retrograde tracer (Wickersham, Lyon et al., 2007, Neuron). This highly innovative combination of cell-type specific expression of these genes and the targeted selectivity of the EnvA-¿RabG rabies virus will enable independent tracing of the connections of local clusters of inhibitory or excitatory cells. Moreover, once developed it will be used to trace the long-range connectivity of inhibitory and excitatory neurons in cat V1 with respect to the orientation map; maps that will be derived through intrinsic signal optical imaging. In this way, the functional preference of inputs to inhibitory neurons in neocortex can be determined for the first time and compared to the preference of inputs to excitatory cell populations. Finally, while the goal of this proposal is to ultimately implement th new technique to understand intrinsic circuits in cat V1, this technique will be available to other for use in any mammalian species and any region of neocortex.

描述（由申请人提供）：新皮质由密集的兴奋性神经元群组成，这些兴奋性神经元群由多样化且分布更稀疏的抑制性神经元阵列组成，因此，皮质抑制性神经元提供了在多种条件下工作的增益控制。例如，在初级视觉皮层 (V1) 中，抑制对于单个神经元的几个基本功能特征至关重要，包括对刺激对比度、大小和方向的偏好，而抑制性神经元则用于控制兴奋。了解抑制性细胞类型和兴奋性细胞类型是否通过同一组皮质回路介导，以及这些回路的功能选择性是否不同，确定这两种主要细胞类型的神经回路的起源将为了解基本机制提供深刻的见解。然而，为了做到这一点，有必要开发一种用于细胞类型特异性神经解剖学追踪的技术，这里的建议是利用重组病毒的向神经性来开发这种技术。载体（腺相关病毒和慢病毒）和细胞类型特异性启动子（GAD-67 和 ¿ -CamKII) 传递并允许两个关键基因（TVA 和 RabG）的表达，从而这些基因将在抑制性或兴奋性新皮质细胞中选择性表达，但不表达。在这些特定细胞类型中表达的 TVA 和 RabG 蛋白将允许靶向病毒感染和新型转基因假型狂犬病病毒 (EnvA-¿RabG) 的反式互补，该病毒充当单突触逆行示踪剂 (Wickersham, Lyon 等人，2007，Neuron）。这些基因的细胞类型特异性表达和 EnvA-¿ 的靶向选择性的高度创新组合。 RabG 狂犬病病毒将能够独立追踪抑制性或兴奋性细胞的局部簇的连接，一旦开发出来，它将用于追踪抑制性和兴奋性神经元的远程连接。在cat V1中相对于方向图；通过内在信号光学成像导出的图，可以首次确定新皮质中抑制性神经元的输入的功能偏好，并将其与对新皮质的输入的偏好进行比较。最后，虽然该提案的目标是最终实施新技术来了解 cat V1 的内在回路，但该技术将可供其他人用于任何哺乳动物物种和新皮质的任何区域。