CRMP-2介导的早年应激对成年后应激抵抗的影响及其神经可塑性机制研究

Individual subjected to stress may express two different phenotypes: Stress-sensitive and stress-resistance. Early stresses in the form of childhood stresses may affect individual reactivity to stress and coping in adult stage. Numerous functions for Collapsing Response Mediator Protein-2 (CRMP-2) have been identified related to various functions in neuroplasticity. Function important to our investigation is the ability of CRMP-2 to regulate axon and dendrite outgrowth via transport and stabilization of microtubule dimers, as well as enhancement of tubulin’s intrinsic GTPase activity for microtubule depolymerization. . Several studies, including our previously study, have shown that CRMP-2 is involved in stress and neuronal plasticity of hippocampus. These studies suggest that CRMP-2 may involve in neuronal plasticity in depression via microtubular regulation. However, the role of CRPM-2 in neuronal plasticity of stress resistance phenotype and stress sensitive and the mechanism of CRMP-2 involving in microtubular system associated neuroplasticity remain unclear and need more researches. . The primary aim of the present study is to investigate possible alteration of CRMP-2 protein involved in the effects of early stress on adult stress resilience and the neuroplasticity mechanisms. Early stress model is established by exposing the rat to stress in early stage. After the rats reach adult stage, they will be exposed to chronic unpredictable mild stress (CUMS) in order to explore the effect of early stress on the reaction to the adult CUMS. Furthermore, by silencing CRMP-2 and increasing or decreasing CRMP-2 phosphorylation in rat pup, we may discuss how the change in CRMP-2 expression and activation may affect stress resistance, microtubular system and neuroplasticity in adult stage on cellular level after the rats are exposed to adult CUMS. We hope to open up a new angle to look into neuronal plasticity of stress resistance in depression via CRMP-2 and subordinate cytoskeletal microtubular system. Our study may provide helpful exploration for new antidepressant drug target.

个体应对应激时会出现应激敏感和应激抵抗两种不同状态。早年应激会影响成年后个体对应激的反应。脑衰反应调节蛋白（CRMP-2）影响细胞支架微管系统微管二聚体的聚合或解聚，从而调节轴突和树突的生长，参与神经可塑性过程。其他学者及我们的前期研究显示CRMP-2和应激及海马神经元可塑性有关，提示CRMP-2可能通过调节微管系统参与抑郁症神经可塑性过程。但应激抵抗模型CRMP-2介导的细胞支架微管相关的神经可塑性机制尚未明了。本项目拟建立早年母爱剥夺应激模型，成年后给予CUMS，探讨CRMP-2介导的早年应激对成年后应激反应的影响。进一步地，幼年沉默CRMP-2，检测改变的CRMP-2对成年后应激抵抗的影响及细胞支架微管系统、神经可塑性改变；并从细胞水平进一步验证。由此从CRMP-2及所属的细胞支架微管系统角度，探索抑郁症应激抵抗神经可塑性损伤的新视角，为药物的新靶点提供有益的研究基础。

背景：应激可导致抑郁症的发病和复发，虽绝大多数个体对应激敏感，但仍有部分个体对应激产生特殊抵抗状态。这是值得重点关注的领域，有助于抑郁症的防治。.主要研究内容：本项目建立不同程度的早年应激模型，待动物成年后再次给予CUMS,检测CRMP2介导的早年应激对成年后应激反应的影响及其神经可塑性机制。进一步地，沉默 CRMP2或给药后，分别改变CRMP2的表达和活性，检测CRMP2表达及活性的改变对成年动物的行为学影响，以及上述生化指标的变化。并进行PC12细胞的培养，在体实验进一步佐证。.重要结果及关键数据：长期母婴分离联合CUMS组雄性大鼠行为学表现最差（P<0.05），其次是CUMS组。短期母婴分离联合CUMS组与正常对照组行为学表现最好，且两者之间无显著差异。免疫共沉淀结果显示大鼠海马CRMP2与微管蛋白tubulin之间存在相互作用。免疫荧光显示海马CA1区、CA3区、DG区，CRMP2与tubulin及actin均存在共定位表达。长期母婴分离使P-CRMP2的水平明显升高（P<0.05），降低了Tyr-tubulin水平，使Acet-tubulin水平升高（P<0.05），而短期母婴分离对其水平无影响。长期母婴分离联合CUMS应激使P-CRMP2水平升高（P<0.05），使CRMP2水平下降（P<0.05）。海马注射LY294002后影响CRMP2活性明显降低了大鼠OFT中的中心移动距离以及中心持续时间（P<0.05），注射CRMP2干扰慢病毒后，siCRMP2组大鼠的不动时间显著升高（P<0.05）。细胞学研究发现SB216763、wortmannin可明显改变细胞活性，影响CRMP2、tubulin表达水平（P<0.05）。.科学意义：早年短期母婴分离应激可促进大鼠应激抵抗的产生，与CRMP2相关的细胞支架微管介导的神经可塑性机制密切相关，调节CRMP2的表达和活性可影响细胞支架微管系统，促进大鼠应激抵抗行为的发生。本研究从抑郁症应激抵抗神经可塑性损伤的视角，为药物的新靶点提供新的研究基础。