Large dense core vesicle (LDCVs) biogenesis in neuroendocrine cells involves: (a) production of cargo peptides processed in the Golgi; (b) fission of cargo loaded LDCVs undergoing maturation steps; (c) movement of these LDCVs to the plasma membrane. These steps have been resolved over several decades in PC12 cells and in bovine chromaffin cells. More recently, the molecular machinery involved in LDCV biogenesis has been examined using genetically modified mice, generating contradictory results. To address these contradictions, we have used NPY‐mCherry electroporation combined with immunolabeling and super‐resolution structured illumination microscopy. We show that LDCVs separate from an intermediate Golgi compartment, mature in its proximity for about 1 hour and then travel to the plasma membrane. The exocytotic machinery composed of vSNAREs and synaptotagmin1, which originate from either de novo synthesis or recycling, is most likely acquired via fusion with precursor vesicles during maturation. Finally, recycling of LDCV membrane protein is achieved in less than 2 hours. With this comprehensive scheme of LDCV biogenesis we have established a framework for future studies in mouse chromaffin cells.

中文翻译：

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小鼠嗜铬细胞中大密集核心囊泡的生物发生

神经内分泌细胞中大的致密核心囊泡（LDCV）的生物发生涉及：（a）在高尔基体中加工的货物肽的产生；（b）进行成熟步骤的载有货物的最不发达国家的裂变；（c）这些LDCV移动到质膜。这些步骤已经在PC12细胞和牛嗜铬细胞中解决了数十年。最近，已经使用转基因小鼠检查了涉及LDCV生物发生的分子机制，产生了矛盾的结果。为了解决这些矛盾，我们将NPY-mCherry电穿孔与免疫标记和超分辨率结构照明显微镜相结合。我们显示，LDCVs从中间的高尔基体隔间中分离出来，在其附近成熟约1小时，然后到达质膜。由vSNARE和synaptotagmin1组成的胞外机制很可能是通过在成熟过程中与前体囊泡融合而获得的，它们是从头合成或再循环产生的。最终，LDCV膜蛋白的回收利用不到2小时即可实现。通过这种全面的LDCV生物发生方案，我们为小鼠嗜铬细胞的未来研究建立了框架。