The Mechanism Of 5-HT₃ Receptor In Trigeminal Neuralgia

Objectives:To investigate the expression of 5-HT3 receptor（5-hydroxytryptamine-3 receptor,5-HT3R）in the trigeminal ganglion（TG）of trigeminal neuralgia（TN）and the mechanism of its pain regulation.Methods:（1）We were going to establish a chronic constrictive injury to theinfraobital nerve（CCI-ION）model of rats.Western blot and real-time quantitative PCR（RT-qPCR）were used to detect the expression of 5-HT3R in the TG of CCI-ION rats.Immunofluorescence staining was used to determine the distribution of 5-HT3R in TG.（2）Electrophysiological techniques were used to record the changes of 5-HT current of CCI-ION rats.（3）Targetscan software,RT-qPCR,fluorescence in situ hybridization（FISH）and dual luciferase experiments were used to validate the role of miR-193a-5p in regulating 5-HT3R expression.（4）Chemical synthesis reagents were injected into rat TG to change the content of miR-193a-5p.Von Frey,western blot and electrophysiology were used to detect the expression of 5-HT3R in the TG.（5）Whole transcriptome sequencing and small RNA sequencing and dual luciferase reporter gene experiments were used to determine the binding of lncRNA with miRNA-193a-5p.（6）RT-qPCR、western blot、electrophysiological techniques and von Frey were used to estimate whether lncRNA actually regulated 5-HT3R.The regulation of lncRNA in 5-HT3R in vivo was detected by overexpression of lncRNA.Results:（1）The mechanical pain threshold was decreased in the 14d after CCI-ION,and the expression of 5-HT3R protein and mRNA was significantly increased.Immunofluorescence showed that 5-HT3R was mainly distributed in the medium-small neurons of IB4 and CGRP markers,and less with NF-200.（2）Electrophysiological results showed that 5-HT current density was increased in the TG of CCI-ION rats.（3）The dual luciferase reporter gene experiment verified that miR-193a-5p can regulate 5-HT3R expression.（4）The expression of miR-193a-5p was down-regulated in the TG of CCI-ION rats.Over-expression of miR-193a-5p down-regulated the 5-HT3R expression as well as the current density of 5-HT,and alleviate the mechanical hyperalgesia.Injection of antagomir-193a-5p into TG increased the 5-HT3R expression as well as the current density of 5-HT,and induced the mechanical hyperalgesia.（5）Dual luciferase experiments showed that lncRNA XR 595534.2 binds to miR-193a-5p.（6）The expression of lncRNA XR₅95534.2 was increased in the TG of CCI-ION rats.Over-expression of lncRNA XR 595534.2 up-regulated the 5-HT3R expression as well as the current density of 5-HT,and induced the mechanical hyperalgesia.Conclusions:CCI-ION induced a sustainable incressing in the lncRNA XR 595534.2 expression and lncRNA XR₅95534.2 combined with miR-193a-5p is involved in trigeminal neuralgia by targeting 5-HT3R.