益肾化浊方调控CREB/BDNF通路改善学习记忆及神经元突触可塑性的机制研究

Based on the CREB/BDNF pathway：Investigation of the effects of the kidney-benefiting and turbidity-resolving formula on learning，memory，and neuronal synaptic plasticity in senescence accelerated mouse-prone 8

关键词:  益肾化浊方  阿尔茨海默病  环磷腺苷效应元件结合蛋白  脑源性神经营养因子  突触可塑性

Key words:  Yishen Huazuo Formula  alzheimer’s disease  cAMP response element-binding protein  brain-derived neurotrophic factor  synaptic plasticity

摘要:

[目的] 观察益肾化浊方（简称YHD）早期干预对快速老化易感系8型小鼠（SAMP8）学习记忆能力、海马突触结构功能及环磷腺苷效应元件结合蛋白（CREB）/脑源性神经营养因子（BDNF）通路相关蛋白表达的影响。[方法] 24只雄性3月龄SAMP8小鼠随机分为模型组（SAMP8组）与YHD组，每组12只，同时以12只同月龄雄性抵抗快速老化R1系小鼠（SAMR1）为正常对照（SAMR1组）。YHD组予YHD中药颗粒溶液灌胃，其余两组予同剂量生理盐水灌胃，每日1次，共持续12周。灌胃后通过Morris水迷宫实验评价学习记忆能力；透射电子显微镜观察小鼠海马神经元结构；长时程增强实验（LTP）检测突触神经电生理功能；蛋白免疫印迹法（Western blot）检测各组小鼠海马组织中CREB/BDNF信号通路相关蛋白的表达情况。[结果] 与SAMP8组相比，YHD组在第4、5天逃避潜伏期时间缩短（P<0.05或P<0.01），穿越平台次数有增加趋势；经θ节律串刺激（TBS）刺激后，LTP的场兴奋性突触后电位（fEPSP）幅值和斜率提升幅度显著升高（P<0.05）；CREB蛋白表达显著增加（P<0.01），BDNF及甲基CpG结合蛋白２（MecP2）蛋白表达有升高趋势，三磷酸鸟苷酶活化蛋白（p250GAP）蛋白表达有降低趋势，无显著性差异。[结论] 采用YHD早期干预可以有效改善SAMP8小鼠的学习记忆能力，这可能与其调控CREB、BDNF蛋白表达，改善神经元超微结构，从而改善神经元突触功能相关。

Abstract:

[Objective] To investigate the effects of early intervention with the Yishen Huazuo Formula on learning and memory abilities，hippocampal synaptic structure and function，and the expression of proteins related to the cAMP response element-binding protein（CREB）/brain-derived neurotrophic factor（BDNF） pathway in SAMP8 mice，a model of accelerated aging. [Methods] Twenty-four 3-month-old male SAMP8 mice were randomly divided into a model（SAMP8） group and a Yishen Huazhuo Formula（YHD） group，with 12 mice in each group. Twelve male SAMR1 mice of the same age were selected as the control（SAMR1） group. The YHD group received oral administration of Yishen Huazhuo Formula granule solution，while the other two groups received the same dose of saline solution，administered once daily for 12 weeks. After administration，learning and memory abilities were evaluated using the Morris water maze test；Long-term potentiation（LTP） tests assessed synaptic neurophysiological function；Western blot analysis detected expression levels of proteins related to the “CREB/BDNF signaling pathway” in hippocampal tissue. [Results] Compared with the SAMP8 group，the YHD group showed significantly shorter escape latency times on days 4 and 5（P<0.05，P<0.01），with a trend toward increased platform crossings；The magnitude of LTP-induced fEPSP amplitude and slope enhancement was markedly elevated（P<0.05）. Following TBS stimulation，the slope enhancement of LTP-induced fEPSP showed a significant increase（P<0.05）. CREB protein expression was significantly increased（P<0.01），while BDNF and MecP2 protein expression showed an upward trend. p250GAP protein expression exhibited a downward trend without significant differences. [Conclusion] Early intervention with Yishen Huazuo Formula effectively improves learning and memory functions in SAMP8 mice. This may be related to its regulation of protein expression in the “CREB/BDNF signaling pathway，” improvement of neuronal ultrastructure，and subsequent enhancement of neuronal synaptic function.