小胶质细胞CB2R在抑郁症认知损伤中的作用及机制研究

Classical activation (M1 phenotype) and alternative activation (M2 phenotype) are the two polars of microglial activation states. The inflammatory reaction mediated by it plays an important role in the pathogenesis of depression. CB2R is a membrane receptor on microglia. Our preliminary work found that the neuro inflammation was alleviated after applying CB2R agonist to animal model of depression. Since CB2R can alleviate neuro inflammation, it is wondered whether or not it has an effect on the impaired cognition of depressed animal model. We applied Morris water maze to evaluate the cognition of an LPS induced depression animal model after treatment of CB2R agonist or antagonist; RT-PCR and ELISA are applied to verify the effect of CB2R on the microglia activation phenotype, BDNF and 5-HT secretion in order to speculate the possible mechanism of how CB2R improves cognition; All of above indexes are reevaluated after blocking AMPK/CREB pathway in order to give further insight into signaling pathways CB2R works. The study can verify the hypothesis: the microglia M1 phenotype transform to M2 phenotype through AMPK/CREB path after CB2R is activated, and this may help alleviating neuro inflammation and accelerating 5-HT and BDNF secretion, ultimately the cognitive disorder is improved. The study provides theoretical basis for CB2R to become candidate target of improving cognitive abilities in depression patients.

激活的小胶质细胞具有M1和M2两种极化表型，其介导的神经炎症在抑郁症发病中发挥重要作用，CB2R是该细胞一种膜受体。我们的前期研究发现CB2R激动剂可减轻抑郁大鼠模型脑内炎症，据此本项目拟探索CB2R减轻炎症后能否改善抑郁动物认知损伤及可能机制。在LPS诱导的抑郁动物模型中使用CB2R激动剂/反向激动剂，借助水迷宫检测能否改善抑郁动物模型的认知障碍；用RT-PCR,ELISA等方法检测CB2R对小胶质细胞激活表型，BDNF及5-HT含量的影响，推测CB2R改善认知的可能机制；细胞学实验阻断AMPK/CREB通路后观察CB2R对小胶质细胞M1/M2极化的调控是否受阻。验证假说：CB2R经AMPK/CREB通路，促使小胶质细胞由M1向M2型转化，通过减轻炎症，促进5-HT和BDNF分泌，改善抑郁症认知障碍。本研究为CB2R成为改善抑郁症认知能力的候选药物靶标提供理论基础。

CB2R 和配体结合后调节激活的小胶质细胞极化表型，通过减轻神经炎症,达到改善抑郁动物模型的认知障碍的作用。本实验中慢性不可预知刺激抑郁动物模型中使用CB2R激动剂，借助 Morris水迷宫检测能否改善抑郁动物模型的认知行为，用旷场实验检测动物的焦虑和自主行为，用流式检测术,ELISA，免疫荧光等方法验证CB2R对小胶质细胞激活表型及神经炎症是否有影响。结果显示正常对照组相比，抑郁动物模型在Morris水迷宫实验中逃避潜伏期增加，穿台次数减少，这表明在抑郁心境下，大鼠的空间学习和记忆能力都受损；抑郁大鼠模型在旷场实验中的垂直得分，总运动距离，中央运动距离都较正常对照组下降，这表明抑郁大鼠在新环境中的自主和探索活动都减少。抑郁动物模型在接受CB2R激动剂后，空间认知能力较模型组有所改善，在新环境中自主行为和探究行为增多。根据流式实验结果和Elisa实验分析，和正常对照组相比，抑郁动物模型的小胶质细胞的比例都较正常对照组上升，且炎症因子IL-6、IL-1β量也都升高。但抑郁模型+CB2R激动剂（JWH-015）组里的炎症因子较抑郁模型组有所下降。综合以上实验结果，我们发现抑郁状态下使用CB2R激动剂确实能减轻神经炎症，改善实验动物模型的空间认知能力，缓解其焦虑心境，为CB2R 成为改善抑郁症患者认知能力的候选药物靶标提供详实的理论基础。