Spinal Neurons Ascending Projection To On-cells In Medulla: A Study On A Neural Circuit Of Chronic Pain Generation

Background: Rostral ventromedial medulla(RVM)is one of the key nuclei of the descending modulation system and is thought to be the gate for pain modulation.Neuronal discharges in RVM has been found to be either increased(ON-cells),decreased(OFFcells)or unchanged during noxious stimulation.But it is still unsolved what role ONcells play in pain generation and modulation.Unraveling it helps clarify pain mechanisms and find targets for pain treatment.Purpose: To explore the effect of specific regulation of ON-cells on acute and chronic pain and to explore the neural circuits that ON-cells receive from spinal cord.Methods: 1)CPISPR-Cas9 was used to construct Gper1-Cre mice and homozygous mice were identified by short segment primer PCR and agarose gel electrophoresis.Adenoassociated virus(AAV)containing diphtheria toxin A-chain(DTA)or channelrhodopsin 2(ChR2)were injected in RVM of Gper1-Cre mice to specifically modulate ON-cells.Pain threshold changes after modulation of ON-cells under pathological conditions which were induced by complete Freund's adjuvant(CFA)or dextran sulfate sodium(DSS)were evaluated by hot plate test,formalin test and colorectal distention test.2)Anterograde and retrograde AAV were used to explore the projection between spinal neurons and ON-cells in RVM.The property of this specific type of spinal neurons was investigated by immunofluorescence.These neurons are regulated by chemical genetics or DTA virus to explore the role in pain modulation.3)Calcium signal changes of ON-cells were observed using two-photon and calcium imaging during pain stimulation or optogenetic activation of spinal neurons to explore whether spinal neurons can activate ON-cells directly.Results: 1)Gper1-Cre homozygous mice were successfully constructed and stably propagated.Chronic pain including inflammation pain,thermal pain and visceral pain could be alleviated by destroying ON-cells in RVM,while it could be facilitated after activation of ON-cells.2)There is a type of excitatory neurons ascending projection to RVM(SNAPR).Mechanical hyperalgesia induced by CFA could be reduced by inhibition of SNAPR.3)Firing activity of ON-cells in RVM can be increased during pain stimulation and optogenetic activation of SNAPR.Conclusion: SNAPR can directly project to ON-cells in RVM.The hyperalgesia of chronic peripheral and visceral pain can be reduced by destroying ON-cells and inhibition of descending facilitation system.The “SC-RVM-SC” neural circuit may be one of the reasons for pain chronicity and complexity of chronic pain.