Functional and genetic characterization of human DRG and spinal cord at single cell resolution

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Specific Aims: Tens of millions of Americans suffer from chronic pain. Unfortunately for these patients, there is very little we can offer in the way of treatments. Opioids represent the main tool for treating pain, but their use in chronic pain conditions suffers from a poor evidence base and the inherent risk of addiction. The current crisis of opioid-related deaths highlights the risk associated with widespread opioid use. The parallel crises of chronic pain and opioid-related deaths has led the NIH to launch the HEAL Initiative, with one of the major goals being the identification of novel analgesics for the treatment of pain, with a focus on drugs that are non-addicting. The PRECISION Human Pain Network seeks to provide foundational data on the diversity among cell types that comprise the pain neuraxis. Goals for our proposed PRECISION Human Pain Network U19 center include the development of optimized experimental protocols and functional assays utilizing primary human cells and tissue (including dorsal root ganglion neurons and spinal cord slices), and the elucidation of comprehensive molecular/cellular phenotypes that underlie human pain transduction, transmission, and processing under different pain conditions. This project seeks to increase our understanding of the physiological properties of human neurons in the pain neuraxis. The three aims here will begin to catalog the functional profiles and transcriptinoal signatures of DRG and spinal neurons from donors with and without a history of pain or opioid use. We will continue our work to optimize protocols for cutting edge functional and genetic studies in human nervous system tissue. We will work with other centers to optimize protocols to assess physiological, transcriptional, and morphological features of human DRG and spinal cord neurons at cellular resolution, and how these properties are impacted in tissues recovered from patients with a history of pain. Because our donor population also includes patients with substance use and misuse, and in some cases this overlaps with chronic pain conditions, we will also characterize similar properties of DRG and spinal cord neurons in patients with and without a recent history of opioid use. These studies will provide foundational knowledge of human neurons in the pain neuraxis by mapping electrophysiological features of neurons in the human DRG and spinal cord onto genetically-defined subpopulations of human DRG or morphological classes of spinal neurons. The study will also profile tissues derived from donors with and without a history of chronic pain or opioid use/misuse, providing opportunity for new understanding of the impact of pain and substance use on these neurons in the human pain neuraxis. Another major deliverable from the present study will be the development of optimized protocols for key aspects of the proposed analyses, including protocols for RNA sequencing from patch clamped neurons (Patch-seq) and for the preparation, maintenance, and recording form human spinal cord slices.