Genetic, Molecular and Anatomical Characterization of VTA Cell Types Involved in Pain and Addiction

摘要 VTA是两种常见病理疾病的中心枢纽--慢性疼痛和阿片成瘾。在……里面 在这两种情况下，VTA内神经元的生理特性都发生了显著的变化。近期 有证据表明，在阿片成瘾的情况下，并不是所有的VTA神经元都有相同的反应，在阿片成瘾的情况下 疼痛，不同的VTA神经元表现出互补的反应。VTA中细胞类型的清晰描述，其 同源投射和输入，以及这两种情况中哪一种可能涉及到这两种情况 阐明，并且对于理解这些病理情况是必不可少的。在这里，我们建议确定 VTA细胞类型的细胞、分子和解剖格局以及这些细胞在慢性疼痛和 上瘾模特。 VTA显示出巨大的细胞异质性，由DA神经元、GABA能神经元组成 和谷氨酸能神经元，以及非神经元细胞。此外，在DA神经元中还有额外的 异质层，至少有四种VTA DA亚型已被证实。这种巨大的异质性 提出了一个理解VTA电路及其在这些条件下的变化的问题。在这里，我们的目标是 是将小鼠的VTA分解成其组成的细胞成分，以了解这些 在疼痛和成瘾中，个体成分会发生变化。在特定的目标1中，我们将使用单细胞转录 分析慢性疼痛、吗啡注射或两者兼有的VTA细胞。我们希望首先细分 VTA分为其组成细胞类型，然后评估每种细胞类型内的分子变化，在每个细胞类型中 条件。这一目标将有助于发现治疗慢性疼痛的新分子靶点。在……里面 具体目标2，使用新开发的交叉遗传工具获得VTA细胞类型，我们将确定 几类VTA、DA和非DA神经元的投射。下一步，我们将开发一种横断式狂犬病 病毒标记法确定不同VTA细胞类型的输入。这一目标将提供一种神经解剖学 为了解参与这些病理情况的电路奠定了基础。最后，在具体目标3中，指导 通过来自目标1和目标2的数据，我们将识别和验证特定DA细胞类型的新靶点，目标是 逆转以慢性疼痛为特征的低多巴胺状态。 这些目标的结果将为理解提供一个分子、细胞和解剖学框架。 VTA细胞类型，这将与本P50中的所有项目相关。此外，发现和验证 不同VTA细胞类型的候选受体将为发育提供一个极好的切入点 阿片类药物在慢性疼痛治疗中的替代品。

Abstract The VTA is a central hub for two prevalent pathological conditions - chronic pain and opioid addiction. In both these conditions, the physiological properties of neurons in the VTA are significantly altered. Recent evidence suggests that in the case of opioid addiction, not all VTA neurons respond equally, and in the case of pain, different VTA neurons display complementary responses. A clear description of cell types in the VTA, their cognate projections and inputs, and which of these might be involved in these two conditions has not been well elucidated, and is essential to understand these pathological conditions. Here, we propose to determine the cellular, molecular and anatomical landscape of VTA cell types and how these are altered in chronic pain and addiction models. The VTA displays enormous cellular heterogeneity, being comprised of DA neurons, GABAergic neurons and Glutamatergic neurons, as well as non-neuronal cells. Further, among the DA neurons there is additional layer of heterogeneity, and at least four VTA DA subtypes have been demonstrated. This immense heterogeneity presents a problem for understanding VTA circuitry as well as its alterations in these conditions. Here, our goal is to disentangle the murine VTA into its constituent cellular components, towards understanding how these individual components are altered in pain and addiction. In Specific Aim 1, we will use single cell transcriptomics to analyze VTA cells in conditions of chronic pain, morphine administration, or both. We expect to first subdivide the VTA into its constituent cell types, and then evaluate the molecular changes within each cell type, in each condition. This aim will facilitate discovery of new molecular targets towards treatment of chronic pain. In Specific Aim 2, using newly developed intersectional genetic tools to access VTA cell types, we will determine the projections of several classes of VTA DA and non-DA neurons. Next, we will develop an intersectional rabies virus labeling approach to determine the inputs of distinct VTA cell types. This aim will provide a neuroanatomical foundation for understanding circuits involved in these pathological conditions. Finally, in Specific Aim 3, guided by data from Aim 1 and 2, we will identify and validate novel targets in specific DA cell types, with the goal of reversing the hypo-dopaminergic state that is characteristic of chronic pain. The results of these Aims will provide a molecular, cellular and anatomical framework for understanding VTA cell types, that will be relevant to all projects in this P50. Additionally, the discovery and validation of candidate receptors in distinct VTA cell types, will provide an excellent entry point towards developing alternatives to opioids in the management of chronic pain.