迷走神经磁刺激对创伤性脑损伤后大鼠α7烟碱乙酰胆碱受体表达变化的影响

关键词：创伤性脑损伤  迷走神经磁刺激  空间学习记忆能力  α7烟碱乙酰胆碱受体  小胶质细胞

Keywords:traumatic brain injury  vagus nerve magnetic stimulation  spatial learning and memory ability  α7 nicotinic acetylcholine receptor  microglia

摘要：

      摘要 目的：探讨迷走神经磁刺激（VNMS）对创伤性脑损伤（TBI）大鼠行为学及α7烟碱乙酰胆碱受体（α7nAChR）表达及小胶质细胞活化的影响。 方法：24只雄性Wistar大鼠按随机数字表法分为sham TBI组、TBI组、VNMS组、VNMS+MLA组，每组6只。采用自由落体撞击法制备中度TBI模型，sham TBI组仅暴露硬脑膜不施加撞击。VNMS组造模后给予VNMS干预（5Hz，刺激时间10s，间歇时间60s，重复25次，总脉冲数1250，每天1次，连续10天），VNMS+MLA组腹腔注射α7nAChR拮抗剂MLA 5mg/kg，45min后行VNMS治疗（参数与VNMS组一致）。造模后第10天，采用水迷宫实验评估各组大鼠的空间学习记忆能力，取各组大鼠的海马组织进行Western Blot检测α7nAChR蛋白、小胶质细胞不同表型标记物CD11c、Arg-1及炎性因子IL-1β、IL-10的表达水平，通过免疫荧光组织染色观察海马小胶质细胞α7nAChR的表达。 结果：与sham TBI组大鼠相比，TBI组的潜伏期延长（P<0.05），在目标象限上停留时间缩短（P<0.05），穿越平台次数减少（P<0.01），TBI组海马区的α7nAChR蛋白表达显著降低（P＜0.05），M1型标记物CD11c的表达显著增加（P<0.001），IL-1β的表达显著上调（P<0.05），IL-10的表达显著减少（P<0.001）；与TBI组比较，VNMS组大鼠潜伏期缩短（P<0.05），在目标象限上停留的时间延长（P<0.01），穿越平台的次数增加（P<0.05），VNMS组的海马区及小胶质细胞的α7nAChR表达水平显著增多（P＜0.01），Arg-1表达显著增加（P<0.05），CD11c的表达显著降低（P<0.01），IL-1β显著降低（P<0.05），IL-10显著增加（P<0.01）；与VNMS组相比，使用MLA抑制剂后，VNMS对大鼠的空间学习记忆能力的改善作用及其对海马区及小胶质细胞的α7nAChR蛋白表达，Arg-1、CD11c的表达，IL-1β、IL-10的调节作用受到抑制。 结论：VNMS能够改善TBI大鼠的空间学习记忆能力，其机制与上调海马区及小胶质细胞α7nAChR表达调节小胶质细胞活化有关。

Abstract:

      Abstract Objective： To Investigate the effects of vagus nerve magnetic stimulation （VNMS） on behavioral changes， α7 nicotinic acetylcholine receptor （α7nAChR） expression， and microglia activation in rats with traumatic brain injury （TBI）. Method： Twenty-four male Wistar rats were randomly divided into sham TBI， TBI， VNMS， and VNMS+MLA， with 6 rats in each group. A moderate TBI model was prepared using Feeney's Free-fall Impact Model Device. The rats in sham TBI group were exposed to the dura mater without impact. The VNMS group received VNMS intervention （5Hz， 10s stimulation time， 60s rest time， repeated 25 times， a total of 1250 pulses， once a day for 10 consecutive days）. The VNMS+MLA group received intraperitoneal injection of the（α7nAChR） antagonist MLA at a dose of 5mg/kg， followed by VNMS treatment 45 minutes later （with the same parameters as the VNMS group）. On the 10th day， the spatial learning and memory ability of rats in each group was evaluated using a water maze experiment. The hippocampal tissue of rats was collected for Western Blot to detect the expression levels of α7nAChR protein， different phenotypic markers of microglia （CD11c and Arg-1）， and inflammatory factors （IL-1β and IL-10）. The expression of α7nAChR in hippocampal microglia cells was observed through immunofluorescence tissue staining. Result： Compared to the sham TBI group， the TBI group showed an extended latency period （P<0.05）， a reduced time spent in the target quadrant （P<0.05）， and a decreased number of platform crossings （P<0.01）. The TBI group also exhibited significantly decreased expression of α7nAChR protein in the hippocampal region （P<0.05）， increased expression of M1 microglia marker CD11c （P<0.001）， up-regulated expression of IL-1β （P<0.05）， and decreased expression of IL-10 （P<0.001）. Compared to the TBI group， the VNMS group showed a shortened latency period （56.75±6.41 vs 30.10±10.7， P<0.05）， an increased time spent in the target quadrant （P<0.01）， and an increased number of platform crossings （P<0.05）. The VNMS group exhibited significantly increased expression of α7nAChR in the hippocampal region and microglia cells （P<0.01）， increased expression of Arg-1 （P<0.05）， decreased expression of CD11c （P<0.01）， decreased expression of IL-1β （P<0.05）， and increased expression of IL-10 （P<0.01）. Compared to the VNMS group， the administration of MLA inhibitor after VNMS treatment inhibited the improvement of spatial learning and memory abilities in rats， as well as the expression of α7nAChR protein in the hippocampal region and microglia cells， and the expression of Arg-1， CD11c， IL-1β， and IL-10. Conclusion： VNMS can improve the spatial learning and memory abilities in TBI rats， and its mechanism is related to the upregulation of α7nAChR expression in the hippocampal region and microglia cells， which regulates the activation of microglia cells.