Miro1/PINK-1调控线粒体自噬逆转七氟烷麻醉下老年性记忆减退的作用机制

Neurogenesis in the dentate gyrus (DG) of the adult hippocampus has become one of important pathogenesis mechanisms of age-related memory deficits induced by sevoflurane. In our previous study, mitochondrial movement was involved in the process of neurogenesis. but its underlying mechenisms are still unknown. It has been identified that autophagy can alleviate the impairment of learning and memory after anesthesia in elderly rats. Recent studies have indicated that mitophagy disorder is attributed to the imbalance of distribution of motor protein for transport of mitochondria (Miro1/PINK-1).We hypothesize that Miro1/PINK-1-mediated mitophagy enhances neurogenesis through driving mitochondrial movements, which is one of the key mechanisms of persistent memory deficits caused by sevoflurane. To test this hypothesis, we will use specific knockout mice and cellular aging model in vivo and in vitro, generate AdMiro1 and Adshmiro1, AdPINK-1 and AdshPINK-1, and infect with postnatal neural stem/progenitor cells to investigate the effects of Miro1/PINK-1 regulated mitophagy in adult neurogenesis using molecular biology, immunofluorescence and confocal microscopy, and to observe its effects on reversing age-related memory deficits induced by sevoflurane. It will also provide new ideas and strategy for the memory recovery after general anesthesia in senile patients.

成年海马神经发生障碍是七氟烷麻醉下老年性记忆减退的重要机制。我们前期研究发现，线粒体动力学参与了老年麻醉的神经发生过程，但其调控机制未明。文献报道，诱导自噬可改善七氟烷对老龄大鼠的记忆损害；线粒体驱动蛋白Miro1/PINK-1分布失衡导致线粒体自噬障碍。因此我们假设Miro1/PINK-1通过驱动线粒体运动，促进线粒体自噬，调控神经发生持续稳定的更新，是七氟烷麻醉下老年性记忆减退恢复的关键机制之一。本项目将拟通过选择性基因敲除小鼠和神经干细胞/前体细胞衰老模型为研究对象，构建腺病毒载体干预Miro1和PINK-1表达，采用分子生物学、免疫荧光、共聚焦显微等技术探讨Miro1/PINK-1介导线粒体自噬在神经发生中的调控机制，阐明其在七氟烷麻醉下老年性记忆减退恢复中的作用。为老年患者全麻下记忆减退的早期防治提供新靶点和新策略。

麻醉药物诱发的记忆和认知减退,是老年患者围术期管理及术后快速康复面临的主要临床问题。线粒体自噬调控参与神经元能量代谢，然而线粒体自噬在麻醉诱导的老年认知减低中，如何消除功能失调的线粒体，维持衰老神经元细胞稳定的具体机制仍不清楚。. 本项目（1）采用PET-CT和核磁波谱明确七氟烷干预下脑代谢率降低，海马神经递质（谷氨酸和γ-氨基丁酸）代谢动力学降低；发现老龄小鼠在七氟烷干预下海马CA1神经元的树突远端的树突棘密度降低，七氟烷干预新生小鼠的海马DG区神经发生降低，与线粒体自噬水平密切相关。（2）证实自噬激动剂可提高七氟烷麻醉下幼龄小鼠海马DG区神经发生，组蛋白去乙酰化酶抑制剂丁酸钠(NaB)可改善七氟烷对新生神经元增殖的抑制作用，恢复七氟醚降低的BDNF和TrkB表达；证实二甲双胍可调控AMP活化蛋白激酶和ULK1磷酸化，改善七氟烷麻醉后老龄小鼠海马神经元自噬水平，促进自噬受体p62/SQSTM1通过线粒体和蛋白质聚集体的自噬作用靶向降解受损细胞溶质成分；阐明组蛋白乙酰化调控七氟醚麻醉下自噬和NLRP3炎症小体之间的相互作用，SAHA通过增加小鼠海马和原代神经元组蛋白H3和H4乙酰化从而改善自噬，并降低神经炎症。（3）证实七氟烷降低了老年小鼠海马神经元糖代谢和氨基酸代谢，采用AAV-PINK-1、AAV-shPINK-1和CaMKⅡα-Cre小鼠、Vglut2-cre小鼠阐明七氟烷对线粒体动力学(包括线粒体融合、分裂和自噬)过程的影响，证实七氟烷抑制海马CA1区神经元线粒体Parkin募集，导致线粒体自噬降低。七氟烷诱导老龄小鼠线粒体融合蛋白Mfn1和Mfn2减少，由于细胞外膜水平介导膜融合减少，导致兴奋性神经元神经递质谷氨酸转运和释放减少，突触后膜对兴奋性递质的敏感性降低。. 本研究阐明线粒体动力学在七氟烷麻醉下老年认知减退中的作用机制。为老年患者全麻下记忆减退的早期防治提供新靶点和新策略。