Cholinergic modulation of Descending Pain Control Pathways

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Chronic pain conditions plague more than 20% of adults in the United States, emphasizing the need for a more comprehensive understanding of pain control mechanisms and better pain therapies. This need is amplified further by the current opioid crisis, which killed 47,600 Americans in 2018. While opioids are remarkably effective treatments for acute and chronic pain, adverse side effects, including abuse liability and tolerance to the analgesic effects with repeated use, highlight the need for novel non-opioid pain therapies. This need motivates our investigations of pain modulation by acetylcholine (ACh) and its receptors (AChRs) in the ventrolateral periaqueductal gray (vlPAG). Ascending pain signaling from periphery to central nervous system is modulated by the descending pain pathway, including the vlPAG and its projections to rostral ventromedial medulla (RVM) and locus ceruleus (LC). This descending pain pathway is a key site of action of opioids and endogenous pain control. While much is known about this circuitry, the pain modulatory effects of cholinergic inputs to vlPAG are understudied. Our preliminary data show that optogenetic stimulation of cholinergic projections from the pedunculopontine tegmental nucleus (PPTg) to vlPAG is antinociceptive in acute pain. Using cell- and circuit-specific optogenetic approaches, we will test the effect of stimulating cholinergic projections to vlPAG on nociception in a chronic neuropathic pain model, identify the cholinergic receptors mediating these antinociceptive effects, and explore possible interactions between cholinergic signaling and the opioid system. In Aim 1, we will use optogenetic approaches to test antinociceptive effects of stimulating cholinergic PPTg neurons projecting to vlPAG in mice experiencing chronic neuropathic pain. We will also assay affective pain relief of stimulating these projections using a real-time place preference assay. In Aim 2, we will identify cholinergic receptor(s) mediating synaptic communication between PPTg and vlPAG neurons, using high- throughput fluorescence in situ hybridization assay to visualize mRNA expression of muscarinic and nicotinic AChRs on vlPAG neurons. After identifying candidate AChR(s) based on mRNA expression, we will test functional role of these receptors in synaptic communication, using slice electrophysiology and ex vivo optogenetics. Finally, we will test the causal role of identified AChRs in antinociception in vivo using optogenetics and pharmacology in neuropathic pain assays. In Aim 3, we will explore possible interactions between these cholinergic mechanisms and the opioid system, by testing the efficacy of cholinergic analgesia in morphine-tolerant mice and during naloxone precipitated withdrawal from chronic morphine treatment. These studies will employ histological, electrophysiological, optogenetic and pharmacological approaches to advance our knowledge of cholinergic modulation of descending pain pathways, to ultimately help identify novel opioid-independent targets for treating chronic pain.