Mazuir Elisa, Dubessy Anne-Laure, Wallon Loane, Aigrot Marie-Stéphane, Lubetzki Catherine, Sol-Foulon Nathalie
Sorbonne Université, Inserm, CNRS, UMR7225, Institut du Cerveau et de la Moelle épinière, ICM.
Sorbonne Université, Inserm, CNRS, UMR7225, Institut du Cerveau et de la Moelle épinière, ICM; Assistance Publique-Hôpitaux de Paris, GH Pitié Salpêtrière.
J Vis Exp. 2020 Feb 9(156). doi: 10.3791/60912.
In the central nervous system, oligodendrocytes are well-known for their role in axon myelination, that accelerates the propagation of action potentials through saltatory conduction. Moreover, an increasing number of reports suggest that oligodendrocytes interact with neurons beyond myelination, notably through the secretion of soluble factors. Here, we present a detailed protocol allowing purification of oligodendroglial lineage cells from glial cell cultures also containing astrocytes and microglial cells. The method relies on overnight shaking at 37 °C, which allows selective detachment of the overlying oligodendroglial cells and microglial cells, and the elimination of microglia by differential adhesion. We then describe the culture of oligodendrocytes and production of oligodendrocyte-conditioned medium (OCM). We also provide the kinetics of OCM treatment or oligodendrocytes addition to purified hippocampal neurons in co-culture experiments, studying oligodendrocyte-neuron interactions.
在中枢神经系统中，少突胶质细胞因其在轴突髓鞘形成中的作用而闻名，轴突髓鞘形成通过跳跃传导加速动作电位的传播。此外，越来越多的报告表明，少突胶质细胞除了髓鞘形成外还与神经元相互作用，特别是通过分泌可溶性因子。在这里，我们提供了一个详细的方案，可从也含有星形胶质细胞和小胶质细胞的胶质细胞培养物中纯化少突胶质谱系细胞。该方法依赖于在37°C下过夜振荡，这允许上层的少突胶质细胞和小胶质细胞选择性脱离，并通过差异黏附消除小胶质细胞。然后我们描述了少突胶质细胞的培养和少突胶质细胞条件培养基（OCM）的制备。我们还提供了在共培养实验中用OCM处理或添加少突胶质细胞至纯化的海马神经元的动力学，以研究少突胶质细胞与神经元的相互作用。
