DOCKING OF CHROMAFFIN GRANULES - A NECESSARY STEP IN EXOCYTOSIS
DOCKING OF CHROMAFFIN GRANULES - A NECESSARY STEP IN EXOCYTOSIS
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DOI：
10.1007/bf01121448
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发表时间：
1987-04-01
影响因子：
4
通讯作者：
BURGER, MM
中科院分区：
文献类型：
--
作者：
SCHAFER, T;KARLI, UO;BURGER, MM
在牛肾上腺髓质染色质细胞和大鼠嗜铬细胞瘤（pc12）细胞中研究了分泌囊泡的对接，包括对未来融合位点的识别和附着。洋地黄苷渗透后，通过将游离Ca2+浓度提高到μM水平，可以刺激两种细胞的分泌。尽管可溶性细胞质成分的丧失表明颗粒稳定附着在质膜上，等待融合触发，但在渗透作用开始后1小时仍可激发分泌活性。透过性pc12细胞在电子显微镜下可以观察到停靠的颗粒，其中大部分颗粒排列在质膜下。在染色质细胞中，由于颗粒在细胞质中分散，推测停靠的颗粒的数量不能很容易地识别。进一步比较pc12和染色质细胞的实验表明，在Ca2+存在的情况下，渗透细胞仍然可以进行主动对接，而不是颗粒的运输：pc12细胞中短（2分钟）的Ca2+脉冲导致在15分钟的观察期内几乎所有可释放激素的分泌，而在染色质细胞中，只有一小部分颗粒对接，Ca2+的退出导致分泌立即停止。染色质颗粒从高尔基体转运到质膜对接位点似乎依赖于一种对渗透敏感的机制。这可以通过两种渗透细胞类型释放的激素量的差异来证明，这反映了pc12或染色质细胞中停靠在质膜上的颗粒比例的差异。细胞chalasine B和秋水仙碱都不影响对接和对接状态。渗透细胞系统是一种有价值的体外研究分泌囊泡与其靶膜相互作用的技术。
Putative docking of secretory vesicles comprising recognition of and attachment to future fusion sites in the plasma membrane has been investigated in chromaffin cells of the bovine adrenal medulla and in rat phaeochromocytoma (PC 12) cells. Upon permeabilization with digitonin, secretion can be stimulated in both cell types by indreasing the free Ca2+-concentration to μM levels. Secretory activity can be elicited up to 1 hr after starting permeabilization and despite the loss of soluble cytoplasmic components indicating a stable attachment of granules to the plasma membrane awaiting the trigger for fusion. Docked granules can be observed in the electron microscope in permeabilized PC 12 cells which contain a large proportion of their granules aligned underneath the plasma membrane. The population of putatively docked granules in chromaffin cells cannot be as readily discerned due to the dispersal of granules throughout the cytoplasm. Further experiments comparing PC 12 and chromaffin cells suggest that active docking but not transport of granules can still be performed by permeabilized cells in the presence of Ca2+: a short (2 min) pulse of Ca2+in PC 12 cells leads to the secretion of almost all releasable hormone over a 15 min observation period whereas, in chromaffin cells, with only a small proportion of granules docked, withdrawal of Ca2+leads to an immediate halt in secretion. Transport of chromaffin granules from the Golgi to the plasma membrane docking sites seems to depend on a mechanism sensitive to permeabilization. This is shown by the difference in the amount of hormone released from the two permeabilized cell types, reflecting the contrast in the proportion of granules docked to the plasma membrane in PC 12 or chromaffin cells. Neither docking nor the docked state are influenced by cytochalasine B or colchicine. The permeabilized cell system is a valuable technique for thein vitrostudy of interaction between secretory vesicles and their target membrane.