Angevin V, Salecker I, Vaillant C, Le Guen J, Branchereau P, Tiaho F, Van Eyseren I, Pichon Y
Groupe de Neurobiologie, Equipe C.R.M., UPRESA-6026 CNRS, Université de Rennes 1, France.
Cell Tissue Res. 2000 Jan;299(1):129-43. doi: 10.1007/s004419900106.
Neurons dissociated from the brain of embryonic cockroaches (Periplaneta americana) can be maintained in culture for several weeks. The survival as well as the progressive organization of the neurons into a complex network was studied during a 5-week period under different culture conditions. About 10% of the dissociated cells adhered to the culture dish. This figure remained constant throughout the culture. The cell diameter ranged from 10 to 20 microns and did not change significantly over time in culture. Whereas only a few cells exhibited neurites at the start of the culture, the number of cells exhibiting neurites increased to reach about 99% after 2 weeks. The different cells were then connected to each other, forming a network, which became more and more complex. The number of cells per cluster as well as the length and the diameter of the "connectives" that linked the different clusters were found to increase with time. The morphology of individual neurons within the network was visualized after intracellular injection of biocytin. Labeling with antibodies raised against serotonin or GABA indicated that neurons were able to differentiate and to acquire specific neurotransmitter fates. The serotonergic phenotype was found to appear progressively throughout the culture, in parallel with the formation of the network. Cell density, addition of fetal calf serum, and ecdysone were shown to influence the development of the network.
从胚胎蟑螂（美洲大蠊）大脑中分离出的神经元可以在培养物中维持数周。在5周的时间里，研究了在不同培养条件下神经元的存活情况以及它们逐渐组织成复杂网络的过程。大约10%的解离细胞附着在培养皿上。在整个培养过程中，这个数字保持不变。细胞直径在10到20微米之间，在培养过程中不会随时间发生显著变化。在培养开始时，只有少数细胞表现出神经突，而在2周后，表现出神经突的细胞数量增加到约99%。然后，不同的细胞相互连接，形成一个网络，这个网络变得越来越复杂。发现每个簇中的细胞数量以及连接不同簇的“连接物”的长度和直径都随时间增加。在细胞内注射生物素后，可以观察到网络中单个神经元的形态。用针对血清素或γ-氨基丁酸的抗体进行标记表明，神经元能够分化并获得特定的神经递质命运。发现血清素能表型在整个培养过程中逐渐出现，与网络的形成同步。细胞密度、添加胎牛血清和蜕皮激素被证明会影响网络的发育。
