扶土抑木法对TS大鼠纹状体Glu、GABA及其受体表达的影响

Tourette syndrome is a common chronic neuropsychiatric disease at the onset of childhood. One of the main pathogenesis is thought to be an imbalance in neurotransmitters located in CSTC loop. Clinical practice found that supporting Spleen and suppressing Liver principle could inhibit tics of children. With a view to research advances of Glu reduction and GABA increment participating in TS pathogenesis at home and abroad, we Put forward the hypothesis that supporting Spleen and suppressing Liver principle involves in regulating imbalance of glutamate and γ-aminobutyric acid and its receptor expression in striatum of TS model rats. In this project, our research group will build up TS rats model by intraperitoneal injecting IDPN, and detecte Glu and GABA content and the ratio change in striatum of model rat by the means of high - performance liquid chromatogram -electrochemical detection. And detect the expression of Glu receptors NMDA and GABA receptor GABA-A in striatum of TS rat by Western blot technique. And detect the expression of NMDAmRNA and GABAAmRNA in TS rat striatum by RT-PCR technique. Explore the mechanisms of supporting Spleen and suppressing Liver principle regulating imbalance of amino acid neurotransmitters from protein and nucleic acid level. And reveal the scientific connotation of supporting Spleen and supressing Liver principle, meantime provide experimental and theoretical supports for further applications in clinic.

多发性抽动症（TS）是一种常见的儿童时期发病的慢性神经精神障碍性疾病，CSTC回路中神经递质失衡是TS发病的主要机制。临床实践表明，扶土抑木法可有效抑制患儿抽动行为，结合Glu降低，GABA升高参与TS发病的国内外研究进展，提出扶土抑木法参与调节纹状体Glu、GABA失衡及其受体表达的研究假说。本项目拟以亚氨基二丙腈（IDPN）腹腔注射法建立TS大鼠模型，采用高效液相色谱电化学技术检测TS大鼠纹状体内Glu、GABA含量及其比值变化；采用Western blot技术检测TS大鼠纹状体Glu受体NMDA、GABA受体GABA-A的表达；采用RT-PCR技术检测TS大鼠纹状体Glu受体NMDAmRNA、GABA受体GABAAmRNA的表达。从蛋白、核酸水平探索扶土抑木法对TS模型大鼠纹状体氨基酸类神经递质失衡后的调节机制，揭示扶土抑木法的科学内涵，为其在临床进一步推广应用提供实验依据和理论支持

CSTC回路中神经递质失衡是TS发病的主要机制，扶土抑木法可抑制患儿抽动，本项目通过IDPN腹腔注射建立TS大鼠模型，采用HPLC法、Western blot及RT-PCR技术，从纹状体Glu、GABA失衡及受体表达角度探索健脾止动汤的作用靶点，揭示扶土抑木法科学内涵。.1.筛选TS模型 采用IDPN腹腔注射，每日1次，连续7天，建立不同剂量TS大鼠模型。自200mg/kg组始，大鼠出现刻板运动。250mg/kg组评分高于200mg/kg组（P＜0.05），200mg/kg组纹状体Glu含量低于空白组（P＜0.05），250 mg/kg组纹状体GABA含量高于空白组（P＜0.05）。.2.健脾止动汤对TS纹状体氨基酸类递质的影响 造模后，随机分为空白组、模型组、泰必利组、健脾止动汤组。灌胃6周后，各组间差异无统计学意义（P＞0.05）。.3.健脾止动汤对TS大鼠GABA ARα4mRNA表达的影响 Western Blot：模型组GABA ARα4mRNA较空白组升高（P<0.01），健脾止动汤组较模型组降低（P<0.01）。RT-PCR：GABA ARα4mRNA表达各组间无差异（P>0.05），但健脾止动汤组较模型组趋于降低。.4.健脾止动汤对TS大鼠纹状体谷氨酸受体NMDA亚基NR1、NR2B表达的影响.Western Blot：模型组大鼠纹状体GluN1、GluN2B蛋白表达均低于空白组，但无显著差异（P>0.05）；健脾止动汤组NR2B蛋白表达低于模型组（P<0.05）；健脾止动汤组NR1蛋白表达低于空白组和泰必利组（P<0.05）。RT-PCR：模型组大鼠纹状体GluN1mRNA、GluN2BmRNA表达均高于空白组（P<0.05）；健脾止动汤组NR1mRNA表达高于模型组，但无显著差异（P>0.05）；泰必利组、健脾止动汤组NR2BmRNA表达均低于模型组（P<0.05）；健脾止动汤组NR2BmRNA表达低于泰必利组（P<0.05）。. 结论 250 mg/kg IDPN腹腔注射，每天1次，连续7天建立的TS大鼠模型表面效度和结构效度比较理想。健脾止动汤可降低TS大鼠中枢神经兴奋性，疗效随时间延长而增强。健脾止动汤通过降低纹状体Glu含量，升高GABA含量，下调GABA ARα4mRNA表达，下调GluN2B基因和蛋白表达，拮抗抽动。