纹状体GDNF调控Glu兴奋性毒性影响PD运动症状

Striatal GDNF affects the motor symptoms of PD by regulating Glu excitotoxicity

目的探讨纹状体胶质细胞系源性神经营养因子(GDNF)含量降低,加重帕金森病(PD)运动症状的机制。方法雄性C57/BL小鼠(6~8周龄),纹状体立体定位注射(PBS、AAV-GDNF、AAV-shGDNF),联合亚急性PD模型,腹腔注射1-甲基-4-苯基-1,2,3,6-四氢吡啶(MPTP)30 mg/(kg·d),连续5 d,随机分为PBS组、阴性对照组(NC组)、AAV-shGDNF组、MPTP组、MPTP+AAV-GDNF组、MPTP+AAV-shGDNF组;采用行为学实验(转棒、爬杆和旷场)评估小鼠运动能力,ELISA试剂盒检测纹状体谷氨酸(Glu)含量,Western blot等技术检测GLAST、GLT-1、GluN2B等表达及分布,TUNEL染色观察纹状体神经元凋亡。结果与NC组比较,AAV-shGDNF组小鼠纹状体GDNF表达下调,小鼠运动能力下降,Glu含量升高、Glu转运体(GLAST、GLT-1)表达分布减少。与PBS组比较,MPTP组小鼠纹状体Glu含量升高、GluN2B表达降低。与MPTP组比较,MPTP+AAV-GDNF组小鼠运动能力提高、纹状体Glu含量降低、GluN2B表达升高、神经元凋亡减少;而MPTP+AAV-shGDNF组小鼠运动能力下降、纹状体Glu含量升高、GluN2B表达降低、神经元凋亡增多。结论PD病理进程中,纹状体GDNF表达降低可能通过下调Glu转运体的表达,促进Glu对神经元的兴奋性毒性损伤,加重运动症状。

Objective To explore the mechanism of motor symptoms in Parkinson′s disease(PD)aggravated by decreased glial cell line-derived neurotrophic factor(GDNF)in striatum.Methods Male C57/BL mouse(6-8 weeks),were administered of PBS,AAV-GDNF or AAV-shGDNF in striatum by brain stereoscopic injection,combinated with sub-acute PD model,1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP)30 mg/(kg·d)was administered through intraperitoneal injection for five consecutive days;subsequently,mice were randomly divided into PBS group,negative control(NC)group,AAV-shGDNF group,MPTP group,MPTP+AAV-GDNF group,MPTP+AAV-shGDNF group.Behavior tests(rotarod,pole and field)were applied for assessing the motor ability of mice;ELISA kit was used to detect striatal glutamic acid(Glu)content;Western blot and other techniques were carried out to detect the expression and distribution of GLAST,GLT-1 and GluN2B in striatum;TUNEL staining was applied for observing the apoptosis of neurons in striatum.Results Compared with the NC group,the mice of the AAV-shGDNF group,with down-regulation of GDNF in striatum,had poor motor ability,decreased Glu transporter(GLAST and GLT-1),and increased Glu content.Compared with the PBS group,the mice of the MPTP group had increased Glu content and decreased GluN2B in striatum.Compared with the MPTP group,the mice of the MPTP+AAV-GDNF group showed enhanced motor ability,along with decreased Glu content,increased GluN2B and less neurons apoptosis in striatum;while,the mice of the MPTP+AAV-GDNF group showed worse motor ability,along with augmented Glu content,reduced GluN2B and more neurons apoptosis in striatum.Conclusion In PD pathological process,decreased striatal GDNF may promote the neurons apoptosis by enhancing Glu excitotoxicity,thereby leading to the aggravation of motor symptoms.