Wake-promoting effects of vagus nerve stimulation after traumatic brain injury: upregulation of orexin-A and orexin receptor type 1 expression in the prefrontal cortex

Orexins, produced in the lateral hypothalamus, are important neuropeptides that participate in the sleep/wake cycle, and their expres- sion coincides with the projection area of the vagus nerve in the brain. Vagus nerve stimulation has been shown to decrease the amounts of daytime sleep and rapid eye movement in epilepsy patients with traumatic brain injury. In the present study, we investigated whether vagus nerve stimulation promotes wakefulness and affects orexin expression. A rat model of traumatic brain injury was established using the free fall drop method. In the stimulated group, rats with traumatic brain injury received vagus nerve stimulation （frequency, 30 Hz, current, 1.0 mA; pulse width, 0.5 ms; total stimulation time, 15 minutes）. In the antagonist group, rats with traumatic brain injury were intracerebroventricularly injected with the orexin receptor type 1 （OXIR） antagonist SB334867 and received vagus nerve stimulation. Changes in consciousness were observed after stimulation in each group. Enzyme-linked immunosorbent assay, western blot assay and immunohistochemistry were used to assess the levels of orexin-A and OX1R expression in the prefrontal cortex. In the stimulated group, consciousness was substantially improved, orexin-A protein expression gradually increased within 24 hours after injury and OX1R expres- sion reached a peak at 12 hours, compared with rats subjected to traumatic brain injury only. In the antagonist group, the wake-promoting effect of vagus nerve stimulation was diminished, and orexin-A and OX1R expression were decreased, compared with that of the stim- ulated group. Taken together, our findings suggest that vagus nerve stimulation promotes the recovery of consciousness in comatose rats after traumatic brain injury. The upregulation of orexin-A and OXIR expression in the prefrontal cortex might be involved in the wake-promoting effects of vagus nerve stimulation.

Hub genes and key pathways of traumatic brain injury: bioinformatics analysis and in vivo validation

The exact mechanisms associated with secondary brain damage following traumatic brain injury(TBI)remain unclear;therefore,identifying the critical molecular mechanisms involved in TBI is essential.The m RNA expression microarray GSE2871 was downloaded from the Gene Expression Omnibus(GEO)repository.GSE2871 comprises a total of 31 cerebral cortex samples,including two post-TBI time points.The microarray features eight control and seven TBI samples,from 4 hours post-TBI,and eight control and eight TBI samples from 24 hours post-TBI.In this bioinformatics-based study,109 and 66 differentially expressed genes(DEGs)were identified in a Sprague-Dawley(SD)rat TBI model,4 and 24 hours post-TBI,respectively.Functional enrichment analysis showed that the identified DEGs were significantly enriched in several terms,such as positive regulation of nuclear factor-κB transcription factor activity,mitogen-activated protein kinase signaling pathway,negative regulation of apoptotic process,and tumor necrosis factor signaling pathway.Moreover,the hub genes with high connectivity degrees were primarily related to inflammatory mediators.To validate the top five hub genes,a rat model of TBI was established using the weight-drop method,and real-time quantitative polymerase chain reaction analysis of the cerebral cortex was performed.The results showed that compared with control rats,Tnf-α,c-Myc,Spp1,Cxcl10,Ptprc,Egf,Mmp9,and Lcn2 were upregulated,and Fn1 was downregulated in TBI rats.Among these hub genes,Fn1,c-Myc,and Ptprc may represent novel biomarkers or therapeutic targets for TBI.These identified pathways and key genes may provide insights into the molecular mechanisms of TBI and provide potential treatment targets for patients with TBI.This study was approved by the Experimental Animal Ethics Committee of the First Affiliated Hospital of Nanchang University,China(approval No.003)in January 2016.