Unit of Pharmacology, Medical School, La Laguna University, 38071 La Laguna, Tenerife, Spain.
J Neurosci. 2010 Jan 20;30(3):950-7. doi: 10.1523/JNEUROSCI.2894-09.2010.
Chromogranins/secretogranins (Cgs) are the major soluble proteins of large dense-core secretory vesicles (LDCVs). We have recently reported that the absence of chromogranin A (CgA) caused important changes in the accumulation and in the exocytosis of catecholamines (CAs) using a CgA-knock-out (CgA-KO) mouse. Here, we have analyzed a CgB-KO mouse strain that can be maintained in homozygosis. These mice have 36% less adrenomedullary epinephrine when compared to Chgb(+/+) [wild type (WT)], whereas the norepinephrine content was similar. The total evoked release of CA was 33% lower than WT mice. This decrease was not due to a lower frequency of exocytotic events but to less secretion per quantum (approximately 30%) measured by amperometry; amperometric spikes exhibited a slower ascending but a normal decaying phase. Cell incubation with L-DOPA increased the vesicle CA content of WT but not of the CgB-KO cells. Intracellular electrochemistry, using patch amperometry, showed that L-DOPA overload produced a significantly larger increase in cytosolic CAs in cells from the KO animals than chromaffin cells from the WT. These data indicate that the mechanisms for vesicular accumulation of CAs in the CgB-KO cells were saturated, while there was ample capacity for further accumulation in WT cells. Protein analysis of LDCVs showed the overexpression of CgA as well as other proteins apparently unrelated to the secretory process. We conclude that CgB, like CgA, is a highly efficient system directly involved in monoamine accumulation and in the kinetics of exocytosis from LDCVs.
嗜铬粒蛋白/分泌粒蛋白（Cgs）是大致密核心分泌小泡（LDCVs）的主要可溶性蛋白。我们最近报道，缺乏嗜铬粒蛋白 A（CgA）会导致儿茶酚胺（CAs）的积累和胞吐发生重要变化，这是使用 CgA 敲除（CgA-KO）小鼠得出的结果。在这里，我们分析了一种可以纯合维持的 CgB-KO 小鼠品系。与 Chgb(+/+) [野生型 (WT)]相比，这些小鼠的肾上腺髓质肾上腺素减少了 36%，而去甲肾上腺素含量相似。CA 的总诱发释放比 WT 小鼠低 33%。这种减少不是由于胞吐事件的频率降低，而是由于每个量子的分泌量减少（约 30%），通过安培法测量；安培法尖峰显示出较慢的上升但正常的衰减阶段。细胞用 L-DOPA 孵育会增加 WT 细胞但不会增加 CgB-KO 细胞的囊泡 CA 含量。使用贴附安培法的细胞内电化学显示，L-DOPA 过载会导致 KO 动物的细胞内胞质 CA 明显增加，而 WT 细胞的细胞内 CA 增加则不明显。这些数据表明，CgB-KO 细胞中 CA 囊泡积累的机制已经饱和，而 WT 细胞仍有足够的容量进行进一步积累。LDCVs 的蛋白分析显示 CgA 以及其他显然与分泌过程无关的蛋白的过表达。我们得出结论，CgB 与 CgA 一样，是一种直接参与单胺积累和从 LDCVs 中进行胞吐的高效系统。
