MPP^+对原代培养SAMP8小鼠中脑神经元的毒性作用

Neurotoxicity effect of MPP^+ on primary cultured mesencephalon neurons of SAMP8 mouse

目的:建立MPP+(1-甲基-4-苯基吡啶离子)干预SAMP8(快速老化小鼠P8)小鼠中脑神经元的体外细胞模型。方法:SAMP8新生1 d小鼠的中脑神经元原代混合培养6 d,加入100μmol/L浓度的MPP+,再培养6、9、12 h和24 h后分别对各时间点的中脑原代培养神经元免疫荧光染色或提取蛋白测定TH水平。结果:MPP+导致原代培养的SAMP8小鼠中脑神经元形态学改变。正常对照组神经元形态完整,免疫反应性强,胞体大呈椭圆形,突起多而长且粗壮;MPP+组神经元随时间逐渐出现形态变化,MPP+后6 h即可见突起不完整断续,9 h突起数目减少,缩短;12 h神经元胞体明显变小,24 h突起多消失,神经元胞体小且免疫反应性弱。MPP+导致原代培养的SAMP8小鼠中脑神经元数量在MPP+后9 h开始有显著减少,同时TH蛋白的表达也开始有显著减少,24 h最低。结论:MPP+对原代培养的SAMP8小鼠中脑神经元具有毒性作用。MPP+导致SAMP8小鼠中脑神经元原代混合培养体系的神经元数量下降,同时TH蛋白表达降低,提示MPP+导致其中脑DA能神经元损伤死亡。

AIM：To investigate the effect of MPP＋ on the midbrain neurons of SAMP8 mouse in vitro,which provide the cell model for PD study.METHODS：SAMP8 mouse primary midbrain cell cultures were obtained from mice within 1 day after birth.On the sixth day after culturing,the cultures were treated with 100 μmol/L of MPP＋ for 6 hours,9 hours,12 hours and 24 hours,then the cells were fixed and followed by immunofluorescence or Western blot analysis for the expressed location and the level of TH respectively.RESULTS：MPP＋ led to the morphological changes of primary cultured midbrain neurons of SAMP8 mouse.The neurons had intact morphous with strong immunoreactivity,many and long dendrites in control group.Compared with the control group,the neurons had fewer and thinness dendrites with weak immunoreactivity in MPP＋ group.MPP＋ significantly decreased midbrain neurons numbers with marked decrease of TH protein levels after its treatment for 9 h.CONCLUSION：MPP＋ had neurotoxicity effect on primary cultured midbrain neurons of SAMP8 mouse.Since MPP＋ led to marked decrease of neuronal numbers and TH protein levels,it is suggested that MPP＋ can cause the DA neuronal degeneration in primary midbrain cell cultures.