Investigation of Mechanical Allodynia Circuitry by the Nature of the Injury - Abstract Persistent pain continues to be a major clinical problem due to its high prevalence and lack of adequate treatment options. This proposal is focused on understanding the neuronal populations and central sensitization mechanisms that underlie mechanical allodynia in the dorsal horn. Mechanical allodynia is a common and debilitating condition in which normally innocuous touch or movement become painful after an injury. The dorsal horn is a major site for the integration of somatosensory information and for injury-induced circuitry changes that give rise to mechanical allodynia. In our previous studies to elucidate excitatory populations that convey mechanical allodynia, we identified a new concept for how the circuitry is organization. Specifically, we showed that rather than a single circuit, as it was previously modeled, the excitatory circuitry is composed of overlapping combinations of excitatory populations that differ depending on whether the injury is inflammatory or neuropathic in nature. Work here, will identify the excitatory circuitry for polyneuropathic injuries modeling type I and I diabetes and chemotherapeutic neuropathy. Preliminary data indicate that the previously identified excitatory populations are dispensable for this type of injury. Work here will also examine the inhibitory circuitry that gates mechanical allodynia induced by the three types of injuries. There are three major inhibitory populations that have been implicated in gating mechanical allodynia but their locations within the network and the mechanism of gating remain unclear. Preliminary data included here provides evidence for a fourth population that may specifically gate inflammatory injury induced mechanical allodynia. Aim 1 is focused on identifying the excitatory populations that convey mechanical allodynia as a function of injury type, the synaptic connectivity between them and mechanisms of sensitization. Experiments will utilize spatial transcriptomics, chemogenetics, rabies- and AAV1- mediated circuit tracing, patch-clamp electrophysiology with campari2 and cell targeted apex2 proximity labeling for proteomic analyses. Aim 2 is focused on identifying where in the network, the four inhibitory populations gate mechanical allodynia and in which type of injuries. What are the excitatory neurons that are directly gated and what are the gating mechanisms. Experiments will utilize chemogenetics, patch clamp electrophysiology and apex2 proximity labeling with proteomic analyses. Our work in the last grant period planted the seeds for the analysis that we will perform here. Work here will provide the essential framework of the dorsal horn mechanical allodynia network in the context of injury type, including the combinations of excitatory neurons that convey mechanical allodynia, the combinations of inhibitory neurons that gate mechanical allodynia. We will also gain insight into the synaptic connectivity and cell- and synapse- specific mechanisms of central sensitization for both the excitatory and inhibitory populations for different injury types. This framework can be used to best study how primary afferents and supraspinal inputs impact the induction and maintenance of the dorsal horn mechanical allodynia network. It will also reveal important new therapeutic targets.
