Elucidating the role of the VTA-NAc medial shell projection in motivation, extinction, and reinstatement

项目总结 奖赏关联和动机是引导目标导向行为的关键过程。这些函数是 受中脑边缘多巴胺回路的调制，该回路包括从腹侧投射的多巴胺神经元 被盖区(VTA)至伏核(NAC)。VTA-NAC多巴胺投射广泛地促进了 学习动作-和提示-结果关联，并调节目标导向的消亡和恢复 行为。此外，一项新兴的研究表明，NAC壳的特定亚区整合在一起 中脑多巴胺能信号与感觉和边缘输入一起驱动参与维持的动作， 灭绝，并恢复追逐奖励的行为。然而，关于孤立性VTA是如何发生的，我们知之甚少 多巴胺对NAC内侧壳的输入参与了这些行为。这项提议的主要目标是 解剖动机行为中孤立性VTA投射到NAC内侧壳的神经回路。要定义 为了研究特定的VTA多巴胺投射到NAC内侧壳的作用，我们确定了一个遗传标记，小脑蛋白- 4(Cbln4)，在VTA多巴胺神经元亚群中显示限制性表达。使用病毒- 在Cbln4-IRES-Cre(Cbln4iCre)小鼠中，我发现VTACbln4神经元投射到NAC 内壳区，但不是侧壳区或核心区。此外，该电路的在体光学刺激是 足以促进在没有初级奖励的情况下维持寻求奖励的行为 增援。此外，对这一群体的刺激足以恢复已熄灭的寻求奖励行为。 行为反应。尽管这些研究信息量很大，但并没有研究内源性神经的作用。 VTACbln4-Nac内侧壳环的活动或多巴胺信号在调节兴奋中的作用 行为。利用Cbln4iCre小鼠品系和病毒介导的靶向策略，提出了两个特定的目标 解剖VTACbln4-NAC内侧壳环。目标1将从解剖和功能上描述VTACbln4- NAC内侧壳多巴胺投射的高分辨率定量图像分析和遗传学研究 编码的多巴胺传感器与电路特定切片成像相结合。AIM 2使用体内纤维光度法 记录VTACbln4神经元在获得、消退和线索诱导过程中的神经活动 恢复以食物为动力的操纵子反应。此外，这一目标考察了内生的必要性。 VTACbln4-Nac内侧壳环中神经活动和多巴胺的产生 这些行为使用化学发生抑制和细胞类型特异性CRISPR/SaCas9介导的基因敲除 接近了。这项研究将确定中脑边缘多巴胺回路中的独立投射如何调节不同的 激励行为的方方面面，并提供对这些回路中的中断如何促成实质的洞察 使用和强迫症。拟议的研究将有助于我在体内纤维方面的训练 光度学、体外切片成像和计算数据分析方法，以及为我从事 独立研究人员的职业生涯。

PROJECT SUMMARY Reward association and motivation are key processes guiding goal-directed behaviors. These functions are modulated by mesolimbic dopamine circuitry, which encompasses dopamine neurons projecting from the ventral tegmental area (VTA) to the nucleus accumbens (NAc). VTA-NAc dopamine projections broadly facilitate the learning of action- and cue-outcome associations and regulate the extinction and reinstatement of goal-directed behaviors. Further, an emerging body of research suggests that specific subregions of the NAc shell integrate midbrain dopaminergic signals with sensory and limbic inputs to drive actions involved in the maintenance, extinction, and reinstatement of reward-seeking behaviors. However, little is known about how isolated VTA dopamine inputs to the NAc medial shell contribute to these behaviors. The primary goal of this proposal is to dissect the neural circuitry of isolated VTA projections to the NAc medial shell in motivated behavior. To define the role of a specific VTA dopamine projection to the NAc medial shell, we identified a genetic marker, cerebellin- 4 (Cbln4), that displays restricted expression within a subpopulation of VTA dopamine neurons. Using viral- mediated circuit mapping in Cbln4-IRES-CRE (Cbln4iCre) mice, I found that VTACbln4 neurons project to the NAc medial shell region but not lateral shell or core regions. Additionally, in vivo optical stimulation of this circuit is sufficient to facilitate the maintenance of reward-seeking behaviors in the absence of primary reward reinforcement. Further, stimulation of this population is sufficient to reinstate an extinguished reward-seeking behavioral response. Although highly informative, these studies did not examine the role of endogenous neural activity or dopamine signaling in the VTACbln4-Nac medial shell circuit for its functions in modulating motivated behavior. Two specific aims are proposed using the Cbln4iCre mouse line with viral-mediated targeting strategies to dissect the VTACbln4-Nac medial shell circuit. Aim 1 will anatomically and functionally characterize the VTACbln4- Nac medial shell dopamine projection using high resolution quantitative image analysis and a genetically encoded dopamine sensor coupled with circuit-specific slice imaging. Aim 2 uses in vivo fiber photometry recording to determine the neural activity of VTACbln4 neurons during the acquisition, extinction, and cue-induced reinstatement of food-motivated operant responding. Further, this aim examines the necessity of endogenous neural activity and dopamine production in the VTACbln4-Nac medial shell circuit in determining the expression of these behaviors using chemogenetic inhibition and cell-type specific CRISPR/SaCas9-mediated gene knockout approaches. This study will define how isolated projections within mesolimbic dopamine circuitry regulate distinct aspects of motivated behavior and provide insight into how disruptions in these circuits contribute to substance use and obsessive-compulsive disorders. The proposed studies will facilitate my training in in vivo fiber photometry, ex vivo slice imaging, and computational data analysis methods, as well as prepare me to pursue a career as an independent researcher.