Deletion of the Nucleotide Exchange Factor Vav3 Enhances Axonal Complexity and Synapse Formation but Tampers Activity of Hippocampal Neuronal Networks In Vitro
Deletion of the Nucleotide Exchange Factor Vav3 Enhances Axonal Complexity and Synapse Formation but Tampers Activity of Hippocampal Neuronal Networks In Vitro
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DOI：
10.3390/ijms21030856
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发表时间：
2020-01
影响因子：
5.6
通讯作者：
David Wegrzyn;C. Wegrzyn;K. Tedford;K. Fischer;A. Faissner
中科院分区：
文献类型：
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作者：
David Wegrzyn;C. Wegrzyn;K. Tedford;K. Fischer;A. Faissner
Vav 蛋白激活 RhoA 亚家族的 GTP 酶，调节细胞骨架并参与粘附、迁移、分化、极性和细胞周期。虽然 RhoA GTPases 对神经元形态的重要性是无可争议的，但它们的调节机制尚不清楚。从这个角度来看，我们研究了删除 Vav2、Vav3 和 Vav2 和 3（Vav2−/−、Vav3−/−、Vav2−/−/3−/−）对胚胎海马神经元体外发育的影响。使用海马神经元与原代野生型（WT）皮质星形胶质细胞的间接共培养系统，我们通过免疫细胞化学和多电极阵列分析（MEA）分析了轴突和树突参数、结构突触数量和自发网络活动。在这里，我们在体外（DIV）三天和五天后观察到 Vav3−/− 神经元中的过程复杂性较高，但 Vav2−/− 神经元中没有。此外，培养 14 天后，在 Vav3−/− 中观察到突触形成增强。值得注意的是，Vav2−/−/3−/− 双敲除神经元没有表现出协同效应。有趣的是，这些差异是暂时的，并在 21 天的培养期后得到补偿。网络分析显示，21 DIV 后，Vav3−/− 神经元中自发发生的动作电位数量减少。基于这些结果，Vav3 似乎参与神经元分化的关键事件。
Vav proteins activate GTPases of the RhoA subfamily that regulate the cytoskeleton and are involved in adhesion, migration, differentiation, polarity and the cell cycle. While the importance of RhoA GTPases for neuronal morphology is undisputed, their regulation is less well understood. In this perspective, we studied the consequences of the deletion of Vav2, Vav3 and Vav2 and 3 (Vav2−/−, Vav3−/−, Vav2−/−/3−/−) for the development of embryonic hippocampal neurons in vitro. Using an indirect co-culture system of hippocampal neurons with primary wild-type (WT) cortical astrocytes, we analysed axonal and dendritic parameters, structural synapse numbers and the spontaneous network activity via immunocytochemistry and multielectrode array analysis (MEA). Here, we observed a higher process complexity in Vav3−/−, but not in Vav2−/− neurons after three and five days in vitro (DIV). Furthermore, an enhanced synapse formation was observed in Vav3−/− after 14 days in culture. Remarkably, Vav2−/−/3−/− double knockout neurons did not display synergistic effects. Interestingly, these differences were transient and compensated after a cultivation period of 21 days. Network analysis revealed a diminished number of spontaneously occurring action potentials in Vav3−/− neurons after 21 DIV. Based on these results, it appears that Vav3 participates in key events of neuronal differentiation.