三叉神经脊束核尾侧亚核可塑性改变在神经根慢性压迫诱导三叉神经痛动物模型中的研究

Trigeminal neuralgia (TN) is one of the most painful clinical neuropathic pain，and the pathogenesis of TN is still unknown. As the specificity of the structure and function in trigeminal subnucleus caudalis (Vc),the study of the TN should be different from the theory of pain in the spinal cord.In this research, we will use the newly built animal model of TN by chronic compression trigeminal root previously, to study the following subjects: ① To observe the relationship of the synaptic plasticity dynamic changes and rat pain behaviors in different time points; ② Using nerve bundle tracking technology to study the trigeminal primary afferents sprouting in Vc; ③ To study the expression of neuron synaptic markers with PSD-95, synaptophysin and GAP-43 under light microscope, and analyze the ultrastructural changes of postsynaptic density and synaptic cleft width under electron microscope; ④ Using molecular biology technique, to semi-quantitative study the variation of the synaptic markers with PSD-95, synaptophysin and GAP-43 during time course; ⑤ Using whole-cell patch clamp and in vivo extracellular electrophysiological recording techniques, to study the neuron synaptic plasticity by the changes of long-term synaptic plasticity and short-term synaptic plasticity in lamina Ⅱ of Vc. Our purpose in this research is to systematically elucidate the mechanism of synaptic structure and functional plasticity changes in the animal model of TN induced by chronic compression trigeminal root, using behavioral, morphological, molecular biological and electrophysiological techniques. We believe that it would provide a new target for clinical treatment of trigeminal neuralgia, also provide a reliable theoretical basis and experimental evidence to exploiting new targeted drugs for treatment.

三叉神经痛（TN）是临床最痛苦的神经病理性痛之一，目前发病机制不明。三叉神经脊束核尾侧亚核（Vc）结构功能的特异性，决定了TN研究需独立于疼痛在脊髓的研究理论。探索突触可塑性在TN大鼠Vc中的作用，对寻找TN治疗突破口及开发新药具有重要的临床意义。本课题组在前期创立的大鼠三叉神经根慢性压迫TN模型上研究：①大鼠痛行为与突触可塑性动态改变的相关性；②神经束路追踪初级传入在Vc出芽（sprouting）现象；③Vc神经元突触标志物PSD-95、突触素、GAP-43表达变化及突触超微结构改变；④Western blot等半定量研究突触标志物表达与时程的相关性；⑤全细胞膜片钳和在体细胞外记录技术，观察VcⅡ层神经元长时程和短时程突触可塑性改变。本课题综合利用行为学、形态学、分子生物学和电生理学技术，系统阐明突触可塑性改变在TN压迫模型中的机制，为临床治疗TN及开发新的靶向药物提供理论和实验依据。

三叉神经痛（TN）是累及口面部的三大神经病理性痛之一，目前发病机制尚不明确。三叉神经根区慢性微血管压迫是临床TN常见病因，我们经大鼠眶下裂沿三叉神经逆行插管到达并压迫三叉神经根，首次建立模拟临床病因的TN慢性压迫动物模型。我们通过动物行为学检测了大鼠口面部自发理毛和Von Frey丝诱发的机械痛敏，发现TN模型组动物的自发痛表现和机械性诱发痛敏比对照组增强。免疫组化检测了三叉神经脊束核尾侧亚核（Vc）的突触素、GAP-43、PSD95等蛋白的表达，发现TN模型组相关蛋白表达上调; 三叉神经根区电镜观察结果，发现压迫损伤部位初级传入纤维周围脱髓鞘病变明显，包括神经根区出现明显的组织结构紊乱，髓鞘脱失现象，组织间隙散在分布大量的髓磷脂及肿胀的线粒体。为比较三叉神经与脊神经初级传入在痛觉传递中的不同影响，我们研究了脊神经节和脊髓背角Ⅱ层EM2/SP阳性的突触终末与GABA能中间神经元之间的相互作用，发现EM2阳性的初级传入与SP阳性的初级传入在DRG和脊髓背角都有双标，并且在脊髓背角与GABA也有三标，提示相互间的作用可影响周围神经痛觉传导和脊髓背角的痛觉调制。在体细胞外记录技术检测TN模型组和假手术组，发现TN模型组大鼠Vc中WDR神经元自发放电频率和诱发放电频率均显著高于对照组大鼠，提示三叉神经根慢性压迫损伤可诱导Vc内WDR神经元兴奋性增强。通过本项目的研究工作，完成课题相关的实用新型发明专利一项（标注基金号），推广了新型TN动物模型的方法，撰写发表了相关科研论文，培养了硕士研究生1名。本课题的部分实验结果，为进一步研究TN发病机制提供了一定的参考基础。