O'Malley E K, Black I B, Dreyfus C F
Department of Neurology, Cornell University Medical College, New York City, New York 10021.
Exp Neurol. 1991 Apr;112(1):40-8. doi: 10.1016/0014-4886(91)90112-p.
Recent studies suggest that brain neurons require extracellular signals for continued survival during maturity as well as development. However, factors underlying the survival of specific populations of central neurons remain to be defined. To examine the regulation of neuronal survival, we have studied the substantia nigra (SN) dopaminergic (DA) system, in dissociated cell culture. DA neuron number was monitored immunocytochemically with antibody to tyrosine hydroxylase (TH), the DA biosynthetic enzyme. Initially, mixed cultures were grown at low, medium, and high densities in serum-containing media. After 7 days, the number of neuron-specific enolase (NSE)-positive cells, a measure of total neuron number, was proportional to cell plating density. In contrast, high density culture elicited a marked, disproportionate increase in TH-immunopositive cells, suggesting that high density conditions selectively enhanced the DA subpopulation. To define the role of cellular interactions in the selective increase in DA cells, virtually pure neuron cultures were compared to support cell-neuron cocultures, in fully defined medium. In support cell-neuron cocultures, SN support cells evoked a four-fold increase in TH cells, while NSE number did not differ from controls. Moreover, local support cells elicited a greater increase in TH cell number than support cells derived from other brain regions. To determine whether increased TH cell number reflected enhanced survival, or possibly expression of TH by new populations, we monitored the time course of this effect. TH cell number remained constant after 3 days in cocultures, while declining fourfold in controls. In parallel studies, support cells were added to SN dissociates at zero time or after 3 days.(ABSTRACT TRUNCATED AT 250 WORDS)
最近的研究表明，大脑神经元在成熟和发育过程中需要细胞外信号来维持生存。然而，中枢神经元特定群体生存的潜在因素仍有待确定。为了研究神经元生存的调节机制，我们在解离细胞培养中研究了黑质（SN）多巴胺能（DA）系统。用多巴胺生物合成酶酪氨酸羟化酶（TH）的抗体通过免疫细胞化学方法监测DA神经元数量。最初，混合培养物在含血清培养基中以低密度、中密度和高密度生长。7天后，神经元特异性烯醇化酶（NSE）阳性细胞的数量（衡量总神经元数量的指标）与细胞接种密度成正比。相比之下，高密度培养引起TH免疫阳性细胞显著且不成比例的增加，这表明高密度条件选择性地增强了DA亚群。为了确定细胞间相互作用在DA细胞选择性增加中的作用，在完全限定培养基中，将几乎纯的神经元培养物与支持细胞 - 神经元共培养物进行比较。在支持细胞 - 神经元共培养物中，SN支持细胞使TH细胞数量增加了四倍，而NSE数量与对照组无差异。此外，局部支持细胞比来自其他脑区的支持细胞引起TH细胞数量更大的增加。为了确定TH细胞数量的增加是反映了存活率的提高，还是可能是新群体表达TH，我们监测了这种效应的时间进程。共培养3天后TH细胞数量保持恒定，而对照组中则下降了四倍。在平行研究中，在零时间或3天后将支持细胞添加到SN解离物中。（摘要截断于250字）
