Bader M F, Ciesielski-Treska J, Thierse D, Hesketh J E, Aunis D
J Neurochem. 1981 Oct;37(4):917-33. doi: 10.1111/j.1471-4159.1981.tb04479.x.
Bovine adrenal chromaffin cells were maintained in culture in Dulbecco's modified Eagle's medium containing 20% foetal calf serum and 10 units per ml of Nerve Growth Factor. Under these conditions, chromaffin cells developed up to five neurites per cell. The neurites showed lateral branches and varicosities along their trunk which ended with thick growth cone-like structures. Cultures of chromaffin cells were stained by indirect immunofluorescence with antibodies against (a) chromogranin A to follow the distribution of chromaffin granules, the catecholamine-storing organelles, and (b) tubulin, to study the microtubular system during outgrowth of neurites. Chromogranin A antibodies showed a very intensely staining punctate pattern, not randomly distributed but localized in neurites. Chromaffin granules were found to migrate from the cell body to reach neurite endings where they were densely packed. Intense staining was also observed in varicosities; a linear arrangement of granules was evident along neurite trunks. Tubulin antibodies decorated a complex network, clearly visible at the cell periphery and also in the growth cone-like structures, in the palm region of the growth cone. Colchicine treatment effected retraction of neurites and disappearance of organized microtubule networks; chromaffin granules were found in the perinuclear region of the cell. Some tubulin (0.2% of total membrane proteins) was found in the purified chromaffin granule membrane preparation; however, this tubulin is probably associated with contaminating plasma membranes. By the criteria of morphology and staining with antitubulin antibodies, adult bovine chromaffin cells in culture display characteristics similar to those of sympathetic neurones. In addition, they showed an exaggerated transport of granules. Adult bovine chromaffin cells in culture offer an excellent model for studying the role of microtubules and the contractile apparatus in relation to cell morphological changes and neurosecretion.
牛肾上腺嗜铬细胞在含有20%胎牛血清和每毫升10单位神经生长因子的杜尔贝科改良伊格尔培养基中进行培养。在这些条件下，嗜铬细胞每个细胞可长出多达五条神经突。神经突在其主干上有侧支和膨体，末端为粗大的生长锥样结构。嗜铬细胞培养物用针对以下物质的抗体进行间接免疫荧光染色：(a) 嗜铬粒蛋白A，以追踪嗜铬颗粒（储存儿茶酚胺的细胞器）的分布；(b) 微管蛋白，以研究神经突生长过程中的微管系统。嗜铬粒蛋白A抗体显示出非常强烈的点状染色模式，并非随机分布，而是定位于神经突中。发现嗜铬颗粒从细胞体迁移至神经突末端，并在那里密集堆积。在膨体中也观察到强烈染色；沿神经突主干可见颗粒呈线性排列。微管蛋白抗体勾勒出一个复杂的网络，在细胞周边以及生长锥样结构、生长锥掌状区域清晰可见。秋水仙碱处理导致神经突回缩以及有组织的微管网络消失；在细胞的核周区域发现了嗜铬颗粒。在纯化的嗜铬颗粒膜制剂中发现了一些微管蛋白（占总膜蛋白的0.2%）；然而，这种微管蛋白可能与污染的质膜有关。根据形态学标准以及用抗微管蛋白抗体染色的结果，培养的成年牛嗜铬细胞表现出与交感神经元相似的特征。此外，它们显示出颗粒的过度运输。培养的成年牛嗜铬细胞为研究微管和收缩装置在细胞形态变化和神经分泌方面的作用提供了一个极好的模型。
