Maneb-induced dopaminergic neuronal death is not affected by loss of mitochondrial complex I activity: results from primary mesencephalic dopaminergic neurons cultured from individual Ndufs4+/+ and Ndufs4-/- mouse embryos.
Maneb-induced dopaminergic neuronal death is not affected by loss of mitochondrial complex I activity: results from primary mesencephalic dopaminergic neurons cultured from individual Ndufs4+/+ and Ndufs4-/- mouse embryos.
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DOI：
10.1097/wnr.0000000000000271
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发表时间：
2014-12-03
期刊：
影响因子：
1.7
通讯作者：
Xia Z
中科院分区：
文献类型：
--
作者：
Choi WS;Xia Z
胚胎小鼠腹侧中脑的原代培养被广泛用于研究帕金森病模型中多巴胺能神经元死亡的机制。具体而言，当在基因分型之前准备神经元培养物时，来自同窝出生小鼠的单个小鼠或胚胎培养物对于涉及转基因小鼠的比较研究非常有用。然而，制备单个小鼠胚胎培养物在技术上具有挑战性，因为每个胚胎的中脑中存在少量细胞（150，000 - 300，000），其中仅0.5-5%是酪氨酸羟化酶（TH）阳性的多巴胺能神经元。在这项研究中，我们优化的程序，从个别小鼠胚胎制备原代中脑神经元培养。将中脑神经元精细地解离，铺在Aclar膜盖玻片上，并在补充有FBS的DMEM中孵育5天，然后补充N2 1天，导致来自每个胚胎的多巴胺能神经元的最佳存活。使用这种优化的方法，我们制备了中脑神经元培养单Ndufs 4 +/+或Ndufs 4-/-胚胎，并研究了线粒体复合物I在maneb诱导的多巴胺神经元死亡的作用。我们的研究结果表明，Maneb毒性多巴胺神经元的线粒体复合物I的活性在Ndufs 4-/-文化的损失不受影响。
Primary cultures from embryonic mouse ventral mesencephalon are widely used for investigating the mechanisms of dopaminergic neuronal death in Parkinson's disease models. Specifically, single mouse or embryo cultures from littermates can be very useful for comparative studies involving transgenic mice when the neuron cultures are to be prepared before genotyping. However, preparing single mouse embryo culture is technically challenging because of the small number of cells present in the mesencephalon of each embryo (150,000-300,000), of which only 0.5-5% are tyrosine hydroxylase (TH) -positive, dopaminergic neurons. In this study, we optimized the procedure for preparing primary mesencephalic neuron cultures from individual mouse embryos. Mesencephalic neurons that are dissociated delicately, plated on Aclar film coverslips, and incubated in DMEM supplemented with FBS for 5 days and then N2 supplement for 1 day resulted in the best survival of dopaminergic neurons from each embryo. Using this optimized method, we prepared mesencephalic neuron cultures from single Ndufs4+/+ or Ndufs4-/- embryos, and investigated the role of mitochondrial complex I in maneb-induced dopamine neuron death. Our results suggest that maneb toxicity to dopamine neurons is not affected by loss of mitochondrial complex I activity in Ndufs4-/- cultures.