Life Science Institute, Sophia University, 7-1 Kioi-cho, Chiyoda-ku, Tokyo 102-8554, Japan.
Neuropharmacology. 2011 Jun;60(7-8):1364-70. doi: 10.1016/j.neuropharm.2011.03.005. Epub 2011 Mar 12.
Synaptotagmins (Syts) serve as a Ca²+ sensor in the release of neurotransmitters and hormones. Inositol polyphosphates (InsPPs) such as Inositol 1,3,4,5,6-pentakisphosphate (InsP₅) and inositol hexakisphosphate (InsP₆) bind to Ca²+-binding C2B domain of Syt I and II, and inhibit transmitter release. We have shown that the inhibition by InsPPs is reversed by Ca²+ in adrenal chromaffin cells, while a rapid accumulation of endogenous InsP₅ and InsP₆ upon depolarizing stimuli have been reported in these and some other cells. Such a rapid accumulation of InsPPs, if not all, might reflect their dissociation from C2B domain of Syt. To elucidate the functional relevance, we studied the effects of antibodies against C2A and C2B domains (anti-C2A Ab, anti-C2B Ab) on the accumulation of InsPPs induced by Ca²+ in digitonin-permeabilized adrenal chromaffin cells. Anti-C2B Ab by itself caused an accumulation of InsPPs in the permeabilizing medium, and increased spontaneous release of catecholamines (CA). Anti-C2A Ab abolished Ca²+-induced increase of InsPPs in cytosolic component, and inhibited Ca²+-evoked release of CA with little effect on the spontaneous release. Microinjection of InsP₆ but not inositol hexakissulfate into intact chromaffin cells inhibited both spontaneous and nicotine-evoked exocytotic events. These results suggest that endogenous InsPPs bound to the C2B domain clamp spontaneous fusion of the docked or primed vesicles at resting level of intracellular Ca²+, and binding of Ca²+ to the C2A or/and C2B domain facilitate fusion dissociating InsPPs from Syt in adrenal chromaffin cells. This article is part of a Special Issue entitled 'Trends in neuropharmacology: in memory of Erminio Costa'.
突触结合蛋白（Syts）作为神经递质和激素释放的 Ca²+ 传感器。肌醇多磷酸盐（InsPPs），如肌醇 1,3,4,5,6-五磷酸（InsP₅）和肌醇六磷酸（InsP₆），与 Syt I 和 II 的 Ca²+-结合 C2B 结构域结合，并抑制递质释放。我们已经表明，在肾上腺嗜铬细胞中，InsPPs 的抑制作用可被 Ca²+ 逆转，而在这些细胞和一些其他细胞中，当去极化刺激时，内源性 InsP₅和 InsP₆ 的快速积累已经被报道。如果不是全部，这种 InsPPs 的快速积累可能反映了它们从 Syt 的 C2B 结构域解离。为了阐明其功能相关性，我们研究了针对 C2A 和 C2B 结构域的抗体（抗-C2A Ab、抗-C2B Ab）对细胞通透化的肾上腺嗜铬细胞中由 Ca²+ 诱导的 InsPPs 积累的影响。抗-C2B Ab 本身在通透化介质中引起 InsPPs 的积累，并增加儿茶酚胺（CA）的自发性释放。抗-C2A Ab 消除了细胞溶质成分中 Ca²+-诱导的 InsPPs 的增加，并抑制了 Ca²+-诱发的 CA 释放，对自发性释放几乎没有影响。将 InsP₆而不是肌醇六磺酸微注射到完整的嗜铬细胞中，抑制了自发性和尼古丁诱发的胞吐事件。这些结果表明，内源性 InsPPs 与 C2B 结构域结合，在细胞内 Ca²+ 的静息水平下夹住停靠或引发的囊泡的自发融合，并且 Ca²+ 与 C2A 或/和 C2B 结构域的结合促进融合，使 InsPPs 从肾上腺嗜铬细胞中的 Syt 解离。本文是题为“神经药理学趋势：纪念 Erminio Costa”的特刊的一部分。
