A regulated secretory pathway in cultured hippocampal astrocytes
A regulated secretory pathway in cultured hippocampal astrocytes
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DOI：
10.1074/jbc.274.32.22539
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发表时间：
1999-08-06
影响因子：
4.8
通讯作者：
Rosa, P
中科院分区：
文献类型：
--
作者：
Calegari, F;Coco, S;Rosa, P
据报道，神经胶质细胞表达最初在神经元和神经内分泌细胞中发现的分子，如神经肽、神经肽加工酶和离子通道。为了验证星形胶质细胞是否可以调节分泌囊泡，从胚胎和新生大鼠的海马制备的原代培养物被用来研究亚细胞定位和分泌途径，其次是分泌颗粒蛋白II，一个众所周知的标记致密核心颗粒。通过间接免疫荧光法，在大量培养的海马星形胶质细胞中检测到SgII，在GoIgi复合物和储存在细胞中的致密核心囊泡群体中检测到颗粒蛋白的免疫反应性。亚细胞分离实验表明，SgII存储在一个囊泡人口密度相同的致密核心分泌颗粒存在于大鼠嗜铬细胞瘤细胞。与这些数据一致，生化结果表明，40-50%的分泌颗粒蛋白II在18小时标记期间合成的细胞内保留超过4小时的追逐期，并释放后，用不同的促分泌素处理。最有效的刺激似乎是佛波酯与离子霉素在细胞外Ca 2+存在下的组合，发现该处理产生细胞内钙[Ca 2 +]的大的和持续的增加（i）瞬变。我们的研究结果表明，受调节的分泌途径的特征在于（i）受调节的分泌颗粒的典型标志物的表达和刺激的胞吐，（ii）致密核心囊泡的存在，和（iii）经历[Ca 2 +]（i）在特定刺激后增加的能力存在于培养的海马星形胶质细胞中。
Glial cells have been reported to express molecules originally discovered in neuronal and neuroendocrine cells, such as neuropeptides, neuropeptide processing enzymes, and ionic channels. To verify whether astrocytes may have regulated secretory vesicles, the primary cultures prepared from hippocampi of embryonic and neonatal rats were used to investigate the subcellular localization and secretory pathway followed by secretogranin II, a well known marker for dense-core granules. By indirect immunofluorescence, SgII was detected in a large number of cultured hippocampal astrocytes, Immunoreactivity for the granin was detected in the GoIgi complex and in a population of dense-core vesicles stored in the cells. Subcellular fractionation experiments revealed that SgII was stored in a vesicle population with a density identical to that of the dense-core secretory granules present in rat pheochromocytoma cells. In line with these data, biochemical results indicated that 40-50% of secretogranin II synthesized during 18-h labeling was retained intracellularly over a 4-h chase period and released after treatment with different secretagogues. The most effective stimulus appeared to be phorbol ester in combination with ionomycin in the presence of extracellular Ca2+, a treatment that was found to produce a large and sustained increase in intracellular calcium [Ca2+](i) transients, Our findings indicate that a regulated secretory pathway characterized by (i) the expression and stimulated exocytosis of a typical marker for regulated secretory granules, (ii) the presence of dense-core vesicles, and (iii) the ability to undergo [Ca2+](i) increase upon specific stimuli is present in cultured hippocampal astrocytes.