Localization of glucose-6-phosphate dehydrogenase activity on ribosomes ofgranular endoplasmic reticulum, in peroxisomes and peripheral cytoplasm ofrat liver parenchymal cells

Citation
Wm. Frederiks et H. Vreeling-sindelarova, Localization of glucose-6-phosphate dehydrogenase activity on ribosomes ofgranular endoplasmic reticulum, in peroxisomes and peripheral cytoplasm ofrat liver parenchymal cells, HISTOCHEM J, 33(6), 2001, pp. 345-353
Citations number
43
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Medical Research Diagnosis & Treatment
Journal title
HISTOCHEMICAL JOURNAL
ISSN journal
0018-2214 → ACNP
Volume
33
Issue
6
Year of publication
2001
Pages
345 - 353
Database
ISI
SICI code
0018-2214(2001)33:6<345:LOGDAO>2.0.ZU;2-B
Abstract
Glucose-6-phosphate dehydrogenase activity has been localized ultrastructur ally in fixed tissues. Activity was found in particular in association with ribosomes of granular endoplasmatic reticulum. Biochemical studies indicat ed that glucose-6-phosphate dehydrogenase activity is also present in the c ytoplasm and in peroxisomes. Fixation may be held responsible for selective inactivation of part of glucose-6-phosphate dehydrogenase activity. In the present study, we applied the ferricyanide method for the demonstration of glucose-6-phosphate dehydrogenase activity in unfixed cryostat sections of rat liver in combination with the semipermeable membrane technique and in isolated rat liver parenchymal cells. Isolated liver parenchymal cells were permeabilized with 0.025% glutaraldehyde after NADP(+) protection of the a ctive site of glucose-6-phosphate dehydrogenase. This treatment resulted in only slight inactivation of glucose-6-phosphate dehydrogenase activity. Th e composition of the incubation medium was optimized on the basis of rapid light microscopical analysis of the formation of reddish-brown final reacti on product in sections. With the optimized method, electron dense reaction product was observed in cryostat sections on granular endoplasmic reticulum , in mitochondria and at the cell border. However, the ultrastructural morp hology was rather poor. In contrast, the morphology of incubated isolated c ells was preserved much better. Electron dense precipitate was found on rib osomes of the granular endoplasmic reticulum, in peroxisomes and the cytopl asm, particularly at the periphery of cells. In conclusion, our ultrastruct ural study clearly demonstrates that it is essential to use mildly-fixed ce lls to allow detection of glucose-6-phosphate dehydrogenase activity in all cellular compartments where activity is present.