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目的中脑内向纹状体投射的mesostriatal多巴胺能细胞亚群是导致帕金森病的关键神经元,本实验拟寻找该亚群投射神经元的分布及有效分离方法。方法利用立体定位技术将绿色荧光逆行示踪剂注射到新生鼠纹状体内,24h后灌注取脑做冰冻切片;应用免疫荧光组织化学染色方法行酪氨酸羟化酶(TH)染色,采用激光显微切割技术将荧光示踪剂标记且呈TH染色阳性的神经元精确切割并收集。结果荧光示踪的神经元呈TH阳性者即属于mesostriatal亚群神经元,主要分布于中脑黑质,应用激光显微切割技术可精确分离这些逆行示踪且TH染色阳性的神经元。结论应用逆行荧光示踪和激光显微切割技术可精确分离mesostriatal亚群神经元。
The purpose of this study is to investigate the distribution of neurons in mesostriatal dopaminergic neurons and the effective methods for the isolation of neurons in Parkinson’s disease. Methods Green fluorescent retrograde tracer was injected into neonatal rat striatum by stereotaxic technique. Frozen sections were obtained 24 hours after perfusion. Tyrosine hydroxylase (TH) staining was performed by immunofluorescence histochemical staining. Microdissection excised and collected neurons stained with fluorescent tracer and stained positive for TH. Results The neurons stained positive for TH were neurons belonging to the mesostriatal subgroup, mainly distributed in the substantia nigra of the midbrain. These retrograde tracing and TH-positive neurons were accurately separated by laser microdissection. Conclusion The mesostriatal subpopulation neurons can be accurately separated by retrograde fluorescence tracing and laser microdissection.