Down Regulating Long Noncoding RNA BDNF-AS Expression Protects Bupivacaine Induced Neurotoxicity In Mice Dorsal Root Ganglion Neurons

Objective:In the present research,we observed the expression pattern of BDNF-AS in neonatal mouse DRG neurons through bupivacaine induced neurotoxicity in vitro.Moreover,we used siRNA to specifically downregulate BDNF-AS expression to analyze the effects of neurite lengths and apoptotic neurons in neurotoxicity DRG.furthermore,we identify the relation between BDNF-AS downregulation and TrkB signaling pathway in local anesthetic injured DRG neurons.Considering the above-mentioned,we explore the possible mechanism of regulating BDNF-AS in protecting DRG neurons caused by bupivacaine induced neurotoxicity to provide valuable therapeutic targets for clinical local anesthetic-injured neurons.Methods:1.To establish the model of Neonatal mouse DRG neurons neurotoxicity induced by bupivacaine in vitro and detect Apoptosis of DRG cells with AnnexinV-FITC/PI double-staining flow cytometry;2.Measurement on mRNA levels of BDNF-AS extracted from DRG explant treated with various concentrations and times of bupivacaine was performed through quantitative real-time PCR(qRT-PCR);3.Local anesthetic neurotoxicity injury model in vitro DRG was randomly divided into two groups:siRNA-C group using 100 nm non-specific siRNA-C transfection for 24 hours;siRNA-BDNF-AS group using 100 nm specific siRNA-BDNF-AS transfection for 24 hours.qRT-PCR was used to detect the expression of BDNF-AS and BDNF;4.The specific siRNA against BDNF-AS(siRNA-BDNF-AS)was implanted,and then the effects of BDNF-AS downregulation on neurite regrowth and neuronal apoptosis in bupivacaine-injured DRG neurons were probed by immunocytochemistry assay and TUNEL assay respectively;5.BDNF-AS downregulation activating TrkB signaling pathway in local anesthetic injured DRG neurons was detected by western blot assay.Results:1.During the process of bupivacaine-induce neurotoxicity in DRG,BDNF-AS was significantly upregulated in both dosage and time dependent manners.2.In DRG neurons,specific siRNA-mediated BDNF-AS downregulation promoted neurite outgrowth and reduced neuronal apoptosis after bupivacaine-induce neurotoxicity.3.Suppressing BDNF-AS transfected with specific siRN A-BDNF-AS activated phosphorylated TrkB pathway after bupivacaine-induced neurotoxicity in DRG neurons.Conclusion:Downregulating BDNF-AS expression protected local anesthetic-induced neurotoxicity in DRG neurons,probably through the upregulation of BDNF and activation of neurotrophin TrkB signaling pathway.