Schwarzenbrunner U, Schmidle T, Obendorf D, Scherman D, Hook V, Fischer-Colbrie R, Winkler H
Department of Pharmacology, University of Innsbruck, Austria.
Neuroscience. 1990;37(3):819-27. doi: 10.1016/0306-4522(90)90111-g.
Homogenates of bovine splenic nerve and of vas deferens were subjected to differential and density gradient centrifugation to investigate their noradrenaline-storing organelles. The subcellular fractions obtained were analysed by immunoblotting in order to define the presence of various antigens in small dense-core and large dense-core vesicles. In both large granule and microsomal fractions from splenic nerve only one type of noradrenaline-storing vesicle was found, which represents the large dense-core vesicles. These organelles contained chromogranin A, chromogranin B, cytochrome b-561, carboxypeptidase H, glycoprotein II, glycoprotein III, dopamine beta-hydroxylase and the monoamine carrier which are also present in adrenal chromaffin granules. The subcellular distribution of synaptin/synatophysin was more complex since this protein was apparently present in two organelles: in a light vesicle which did not contain significant amounts of antigens found in large dense-core vesicles (dopamine beta-hydroxylase, cytochrome b-561 and the monoamine carrier) and in the dense fractions of the gradient, possibly within large dense-core vesicles. In the microsomal gradient from vas deferens several markers (catecholamines, synaptin/synaptophysin and dopamine beta-hydroxylase) were found in a bimodal distribution, which is consistent with their presence in small and large dense-core vesicles. When the larger granules were removed with higher centrifugation speed a microsomal fraction containing only light vesicles was obtained. After gradient centrifugation of this fraction several components (catecholamines, dopamine beta-hydroxylase, cytochrome b-561, the monoamine carrier and synaptin/synaptophysin) were concentrated in a peak at low density; apparently only small dense-core vesicles were now present.(ABSTRACT TRUNCATED AT 250 WORDS)
对牛脾神经和输精管的匀浆进行差速离心和密度梯度离心，以研究其去甲肾上腺素储存细胞器。通过免疫印迹分析获得的亚细胞组分，以确定小致密核心囊泡和大致密核心囊泡中各种抗原的存在情况。在脾神经的大颗粒和微粒体组分中，仅发现一种去甲肾上腺素储存囊泡，即大致密核心囊泡。这些细胞器含有嗜铬粒蛋白A、嗜铬粒蛋白B、细胞色素b-561、羧肽酶H、糖蛋白II、糖蛋白III、多巴胺β-羟化酶和单胺载体，这些物质也存在于肾上腺嗜铬颗粒中。突触素/突触囊泡蛋白的亚细胞分布更为复杂，因为这种蛋白显然存在于两种细胞器中：一种轻囊泡中，其不含有大致密核心囊泡中发现的大量抗原（多巴胺β-羟化酶、细胞色素b-561和单胺载体），另一种存在于梯度的致密组分中，可能在大致密核心囊泡内。在输精管的微粒体梯度中，几种标志物（儿茶酚胺、突触素/突触囊泡蛋白和多巴胺β-羟化酶）呈双峰分布，这与它们存在于小和大致密核心囊泡中一致。当以更高的离心速度去除较大颗粒时，获得仅含轻囊泡的微粒体组分。对该组分进行梯度离心后，几种成分（儿茶酚胺、多巴胺β-羟化酶、细胞色素b-561、单胺载体和突触素/突触囊泡蛋白）集中在低密度峰处；显然现在仅存在小致密核心囊泡。（摘要截断于250字）
