Zaror-Behrens G, Depocas F, Lacelle S
Can J Physiol Pharmacol. 1982 Jan;60(1):13-22. doi: 10.1139/y82-002.
Rat interscapular brown adipose tissue (IBAT) was homogenized to release the noradrenergic vesicles present in its dense sympathetic innervation. The vesicles were then studied by several sedimentation techniques using noradrenaline (NA) and dopamine beta-hydroxylase (DBH) as markers. Sixty-three percent of the DBH activity and 29% of the NA in homogenates (0.25 M sucrose, 5 mM Tris, pH 7.4 at 21 degrees C) of IBAT from 28 degrees C acclimated rats sedimented in the microsomal fraction (226 600 X gmax, 60 min). Differential sedimentation of the microparticulate DBH in a low-speed supernatant fraction of the homogenate indicated at least two distinct populations of microparticles with average sedimentation coefficients of 80 +/- 11 and 255 +/- 42 S (4 degrees C) and containing, respectively, about 65 and 35% of the sedimentable DBH. Upon isopycnic, sucrose density centrifugation of the resuspended microsomal fraction, DBH peaked at a density of 1.091 but extended as a broad shoulder up to a density of about 1.19. During rate zonal centrifugation of the resuspended microsomal fraction on sucrose density gradients, microparticulate DBH and NA separated into slow and fast moving components. The modal density of the slow moving component upon isopycnic recentrifugation was 1.092, while the fast moving one, similarly treated, became almost equally distributed over a range of densities from 1.12 to 1.19. For the slow moving component, NA and DBH relative to protein were, respectively, 6.5 and 23 times more concentrated than in the IBAT homogenate. On the basis of its measured sedimentation characteristics, the slow moving component would correspond to vesicles having a calculated diameter of 66 nm. The data thus indicate that in IBAT, DBH and NA can be separated into two distinct populations of sedimentable particles. Whether or not these correspond to the small and large dense-cored vesicles observed by ultramicroscopy of IBAT remains to be demonstrated.
将大鼠肩胛间棕色脂肪组织（IBAT）匀浆，以释放其密集交感神经支配中存在的去甲肾上腺素能囊泡。然后使用去甲肾上腺素（NA）和多巴胺β-羟化酶（DBH）作为标志物，通过几种沉降技术对囊泡进行研究。来自28℃适应大鼠的IBAT匀浆（0.25M蔗糖，5mM Tris，21℃下pH 7.4）中，63%的DBH活性和29%的NA沉降到微粒体部分（226 600×gmax，60分钟）。匀浆低速上清液部分中微粒DBH的差异沉降表明，至少有两种不同的微粒群体，其平均沉降系数分别为80±1l和255±42 S（4℃），分别含有约65%和35%的可沉降DBH。对重悬的微粒体部分进行等密度蔗糖密度离心时，DBH在密度1.091处达到峰值，但作为一个宽峰延伸至约1.19的密度。在重悬的微粒体部分在蔗糖密度梯度上进行速率区带离心时，微粒DBH和NA分离为慢速和快速移动的组分。等密度再离心时，慢速移动组分模式密度为1.092，而同样处理的快速移动组分在1.12至1.19的密度范围内几乎均匀分布。对于慢速移动组分，相对于蛋白质的NA和DBH浓度分别比IBAT匀浆中高6.5倍和23倍。根据其测量的沉降特性，慢速移动组分将对应于计算直径为66nm的囊泡。因此，数据表明在IBAT中，DBH和NA可分离为两种不同的可沉降颗粒群体。这些是否对应于通过IBAT超微镜观察到的小和大的致密核心囊泡，仍有待证实。
