HIGH-RESOLUTION IMMUNOGOLD ANALYSIS REVEALS DISTINCT SUBCELLULAR COMPARTMENTATION OF PROTEIN-KINASE-C-GAMMA AND PROTEIN-KINASE-DELTA IN RATPURKINJE-CELLS

High resolution immunogold cytochemistry was used to investigate the s ubcellular distribution of protein kinase C gamma and delta in Purkinj e cells of the rat cerebellum. Postembedding incubation with an antibo dy raised to a peptide sequence near the C-terminus of protein kinase C gamma resulted in strong labelling, along the dendrosomatic plasma m embrane. A quantitative analysis indicated that this labelling reflect ed the existence of two pools of protein kinase C gamma; one membrane associated pool and one cytoplasmic pool located within 50 nm of the p lasma membrane. The labelling along the plasma membrane showed a prono unced and abrupt increase when moving from the cell body into the axon initial segment. Gold particles signalling protein kinase C gamma wer e also enriched in putative Purkinje axon terminals in the dentate nuc leus. The only organelle showing a consistent immunolabelling for prot ein kinase C gamma was the Golgi apparatus where the gold particles we re restricted to the trans face. Protein kinase C gamma immunoreactivi ty also occurred in the Purkinje cell spines, with an enrichment in or near the postsynaptic density. Antibodies to protein kinase C delta p roduced a very different labelling pattern in the Purkinje cells. Most of the gold particles were associated with rough endoplasmic reticulu m, particularly with those cisternae that were located close to the nu cleus or in the nuclear indentations. No significant protein kinase C delta immunolabelling was detected al the plasma membrane or in Purkin je cell spines. The present data point to a highly specific compartmen tation of the two major protein kinase C isozymes in Purkinje cells an d suggest that these isozymes act on different substrates and hence ha ve different regulatory functions within these neurons. (C) 1997 IBRO. Published by Elsevier Science Ltd.