Studying Axonal Outgrowth and Regeneration of the Corticospinal Tract in Organotypic Slice Cultures.
Studying Axonal Outgrowth and Regeneration of the Corticospinal Tract in Organotypic Slice Cultures.
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DOI：
10.1089/neu.2014.3467
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发表时间：
2015-09
影响因子：
4.2
通讯作者：
M. Pohland;R. Glumm;L. Stoenica;Markus Höltje;Jürgen Kiwit;Gudrun Ahnert-Hilger;Ulf Strauss;Anja U Bräuer;Friedemann Paul;Jana Glumm
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作者：
M. Pohland;R. Glumm;L. Stoenica;Markus Höltje;Jürgen Kiwit;Gudrun Ahnert-Hilger;Ulf Strauss;Anja U Bräuer;Friedemann Paul;Jana Glumm
脊髓损伤后轴突生长和再生的研究因所涉及事件的复杂性而受到阻碍。在这里，我们提出了一种简单且改进的体外方法来研究皮质脊髓束的生长、再生和内在实质反应。我们使用来自普遍表达绿色荧光蛋白的产后供体小鼠运动皮层的外植体和来自同龄野生型幼鼠的颈脊髓来制备器官型共培养物。我们的数据表明：a) 运动皮质生长在培养 1 天后就已经可检测到，并且具有来源特异性； b) 用来自肉毒梭菌的神经营养蛋白-3和C3转移酶处理可显着增强培养过程中轴突的生长； c) 长出的轴突形成突触连接，如免疫组织化学和钙成像所证明的； d) 运动皮质起源的迁移细胞无需事先追踪即可可靠地识别，并且大部分是在脊髓实质中存活和成熟的神经前体细胞。因此，我们的模型适用于筛选增强皮质脊髓束生长和再生的候选物质，并适用于研究损伤诱导后内源性神经前体的作用。
Studies of axonal outgrowth and regeneration after spinal cord injury are hampered by the complexity of the events involved. Here, we present a simple and improved in vitro approach to investigate outgrowth, regeneration of the corticospinal tract, and intrinsic parenchymal responses. We prepared organotypic co-cultures using explants from the motor cortex of postnatal donor mice ubiquitously expressing green fluorescent protein and cervical spinal cord from wild type pups of the same age. Our data show that: a) motor-cortical outgrowth is already detectable after 1 d in culture and is source specific; b) treatment with neurotrophin-3 and C3 transferase from Clostridium botulinum significantly enhances axonal outgrowth during the course of cultivation; c) outgrowing axons form synaptic connections, as demonstrated by immunohistochemistry and calcium imaging; and d) migrating cells of motor-cortical origin can be reliably identified without previous tracing and are mostly neural precursors that survive and mature in the spinal cord parenchyma. Thus, our model is suitable for screening for candidate substances that enhance outgrowth and regeneration of the corticospinal tract and for studying the role of endogenous neural precursors after lesion induction.