Investigation of Nociceptive Endogenous Opioid Dynamics in the Periaqueductal Gray

Opioid analgesics mimic endogenous opioid peptide function in nociceptive neural circuits by engaging mu opioid receptor (MOR) signaling, resulting in antinociception and pain relief. The ventrolateral periaqueductal gray (vlPAG) is a hub of nociceptive and endogenous opioid signaling in the brain. However, much remains unknown regarding opioid peptide release properties and the modulation of functionally distinct vlPAG MOR-expressing neurons during acute and chronic pain. Thus, the purpose of this study is to dissect the spatial and temporal activity dynamics of this opioid peptide and MOR signaling system in the vlPAG. In our initial approach to this objective, we employed a complementary set of novel adeno-associated viruses that drive expression of the genetically-encoded calcium indicator GCaMP6f selectively in MOR-positive cells, or expression of a novel fluorescent enkephalin sensor, DOR-Light. First, in freely behaving mice, our fiber photometry data demonstrated that vlPAG MOR-expressing neurons are responsive to noxious stimuli, displaying heightened nociceptive calcium transients corresponding with behavioral hyperalgesia following hindpaw injection of the inflammatory agent Complete Freund's Adjuvant (CFA). Second, viral retrograde labeling revealed several local and forebrain circuits expressing enkephalins, which innervate the vlPAG and may modulate nociception. Indeed, optogenetic activation of vlPAG enkephalinergic neurons produced antinociception in Penk-Cre mice during a ramped-temperature hotplate protocol. Finally, DOR-Light photometry recordings revealed CFA-related enhancement of enkephalin release in vlPAG during noxious hotplate exposure. We are continuing to explore our hypothesis that acute noxious and persistent inflammation conditions engage a similar nociceptive subset of vlPAG MOR neurons, which promote top-down release of enkephalins to regulate ongoing midbrain nociceptive signaling. Through precise optical excitation of different enkephalinergic circuits across the brain, we will determine neuropeptide release mechanics and subsequently control endogenous pain resolution, thereby providing foundational support for future targeted therapies that safely recruit the endogenous opioid system for chronic pain treatment. Grant support from 1F32DA055458-01 (awarded to Blake Kimmey).