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

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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.