Cardell M, Landsend A S, Eidet J, Wieloch T, Blackstad T W, Ottersen O P
Department of Anatomy, University of Oslo, Norway.
Neuroscience. 1998 Feb;82(3):709-25. doi: 10.1016/s0306-4522(97)00305-9.
High resolution immunogold cytochemistry was used to investigate the subcellular distribution of protein kinase C gamma and delta in Purkinje cells of the rat cerebellum. Postembedding incubation with an antibody raised to a peptide sequence near the C-terminus of protein kinase C gamma resulted in strong labelling along the dendrosomatic plasma membrane. A quantitative analysis indicated that this labelling reflected the existence of two pools of protein kinase C gamma; one membrane associated pool and one cytoplasmic pool located within 50 nm of the plasma membrane. The labelling along the plasma membrane showed a pronounced and abrupt increase when moving from the cell body into the axon initial segment. Gold particles signalling protein kinase C gamma were also enriched in putative Purkinje axon terminals in the dentate nucleus. The only organelle showing a consistent immunolabelling for protein kinase C gamma was the Golgi apparatus where the gold particles were restricted to the trans face. Protein kinase C gamma immunoreactivity also occurred in the Purkinje cell spines, with an enrichment in or near the postsynaptic density. Antibodies to protein kinase C delta produced a very different labelling pattern in the Purkinje cells. Most of the gold particles were associated with rough endoplasmic reticulum, particularly with those cisternae that were located close to the nucleus or in the nuclear indentations. No significant protein kinase C delta immunolabelling was detected at the plasma membrane or in Purkinje cell spines. The present data point to a highly specific compartmentation of the two major protein kinase C isozymes in Purkinje cells and suggest that these isozymes act on different substrates and hence have different regulatory functions within these neurons.
采用高分辨率免疫金细胞化学技术研究蛋白激酶Cγ和δ在大鼠小脑浦肯野细胞中的亚细胞分布。用针对蛋白激酶Cγ C末端附近肽序列产生的抗体进行包埋后孵育，结果显示沿树突体细胞膜有强烈标记。定量分析表明，这种标记反映了蛋白激酶Cγ的两个池的存在；一个是膜相关池，另一个是位于质膜50 nm内的细胞质池。当从细胞体移向轴突起始段时，质膜上的标记显示出明显且突然的增加。在齿状核中假定的浦肯野轴突终末中，标记蛋白激酶Cγ的金颗粒也富集。唯一对蛋白激酶Cγ持续显示免疫标记的细胞器是高尔基体，其中金颗粒局限于反面。蛋白激酶Cγ免疫反应性也出现在浦肯野细胞棘中，在突触后致密区或其附近富集。蛋白激酶Cδ抗体在浦肯野细胞中产生了非常不同的标记模式。大多数金颗粒与粗面内质网相关，特别是与那些靠近细胞核或位于核凹陷处的扁平囊泡相关。在质膜或浦肯野细胞棘中未检测到明显的蛋白激酶Cδ免疫标记。目前的数据表明，浦肯野细胞中两种主要的蛋白激酶C同工酶存在高度特异性的区室化，提示这些同工酶作用于不同的底物，因此在这些神经元内具有不同的调节功能。
