Cholinergic receptor interactions in the… (F32DA065348) – Project Grant to The University Of Chicago

Abstract: Chronic pain affects over 20% of adults in the united states, and 22.1% of those afflicted individuals have used a prescription opioid in the past 3 months. While opioids provide effective pain relief, they carry adverse side effects including misuse and tolerance. The opioid overdose caused nearly 300,000 deaths between 1999 and 2022. These facts highlight the urgent need to better understand the pain control mechanisms and develop safer, non-opioid pain therapies. The descending pain control pathway, including the ventrolateral periaqueductal grey (vlpag) and its projections to the rostral ventromedial medulla (RVM), is responsible for modulating the ascending pain pathway that carries pain signals from the periphery to the central nervous system. While the role of endogenous opioids in this descending pathway has been extensively studied, the impact of cholinergic system in this pathway under pain conditions is poorly understood. Our previous work showed that pain conditions suppress acetylcholine (ACH) levels in the vlpag in mice. M2 muscarinic acetylcholine receptors (machrs) and Α7 nicotinic acetylcholine receptors (nachrs) are strongly expressed on gabaergic neurons of the vlpag (vlpaggaba), and both acute painful stimuli and chronic pain state increase the activity of these neurons. Additionally, our preliminary data suggested that activating either Α7 or M2 receptors using agonists induces analgesic effects in mice by inhibiting these vlpaggaba neurons. Thus, painful conditions suppress ACH levels in the vlpag, resulting in decreased Α7 and M2 receptor activity, which disinhibits vlpaggaba neurons. However, the extent to which Α7 nachrs and M2 machrs are differentially distributed across gabaergic neuron subtypes remains unknown. Aim 1 will focus on studying the expression of Α7 and M2 receptors on parvalbumin (PV) and somatostatin (SST) neurons in the vlpag using high-resolution fluorescence in situ hybridization and immunohistochemistry. Our previous data showed significant overlap in expression of Α7 and M2 receptors in a subset of vlpag neurons, and we found that endogenous acetylcholine release will activate both receptor classes. Therefore, in aim 2, we will test whether Α7 and M2 receptors in the vlpag interact to modulate pain signaling through additive or synergistic mechanisms using isobologram analysis. We will use a fluorescent calcium sensor, GCAMP8S, with in vivo fiber photometry to monitor SST and PV neuronal activity in the vlpag under acute and chronic pain. Acute pain will be assessed using radiant heat source and VON frey filament test, while chronic neuropathic pain will be induced using the spared nerve injury model. Whole-cell and cell-attached electrophysiological recordings will be conducted to examine pain-induced changes in intrinsic excitability and outward potassium currents of vlpaggaba neurons. Together, these data will advance our understanding of cholinergic modulation in the descending pain pathway and help identify novel targets for chronic pain treatments.

Cholinergic receptor interactions in the modulation of descending pain control pathway

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