Mundorf M L, Troyer K P, Hochstetler S E, Near J A, Wightman R M
Department of Chemistry, University of North Carolina, Chapel Hill, North Carolina 27599-3290, USA.
J Biol Chem. 2000 Mar 31;275(13):9136-42. doi: 10.1074/jbc.275.13.9136.
Effects of vesicular monoamine transporter inhibitors on catecholamine release from bovine chromaffin cells have been examined at the level of individual exocytotic events. As expected for a depletion of vesicular stores, release evoked by depolarizing agents was decreased following 15-min incubations with reserpine and tetrabenazine, as evidenced by a decrease in exocytotic frequency and amount released per event. In contrast, two reserpine derivatives, methyl reserpate and reserpic acid, were much less effective. Surprisingly, the incubations also decreased the accompanying rise in intracellular Ca(2+) evoked by depolarizing agents. Subcellular studies revealed that reserpine and tetrabenazine at concentrations near their K(i) values not only could increase cytoplasmic catecholamines but also could displace Ca(2+) from vesicles. Furthermore, transient exposure to tetrabenazine and reserpine, but not methyl reserpate and reserpic acid, induced exocytotic release of catecholamines. Reserpine induced a rise in intracellular Ca(2+), as detected by whole-cell measurements with Fura-2. It could induce exocytosis, albeit at a lower frequency, in Ca(2+)-free solutions, supporting an internal Ca(2+) source. Depletion of endoplasmic reticulum and mitochondrial Ca(2+) pools did not eliminate the reserpine-activated release. These results indicate that vesicular Ca(2+) can play an important role in exocytosis and under some conditions may be involved in initiating this process.
在单个胞吐事件层面，已对囊泡单胺转运体抑制剂对牛嗜铬细胞儿茶酚胺释放的影响进行了研究。正如囊泡储存耗竭所预期的那样，在用利血平和丁苯那嗪孵育15分钟后，去极化剂诱发的释放减少，这表现为胞吐频率降低以及每个事件释放量减少。相比之下，两种利血平衍生物，利血甲酯和利血酸，效果要差得多。令人惊讶的是，孵育还减少了去极化剂诱发的细胞内Ca(2+)伴随升高。亚细胞研究表明，利血平和丁苯那嗪在接近其K(i)值的浓度下不仅可以增加细胞质儿茶酚胺，还可以从囊泡中置换出Ca(2+)。此外，短暂暴露于丁苯那嗪和利血平，但不包括利血甲酯和利血酸，会诱导儿茶酚胺的胞吐释放。用Fura-2进行全细胞测量检测到，利血平会引起细胞内Ca(2+)升高。在无Ca(2+)溶液中，它可以诱导胞吐作用，尽管频率较低，这支持了内部Ca(2+)来源。内质网和线粒体Ca(2+)池的耗竭并没有消除利血平激活的释放。这些结果表明，囊泡Ca(2+)在胞吐作用中可以发挥重要作用，并且在某些情况下可能参与启动这一过程。
