Electron microscopical study of a cytosolic enzyme in unfixed cryostat sections: demonstration of glycogen phosphorylase activity in rat liver and heart tissue |
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Authors: | Jacques P M Schellens Helena Vreeling-Sindelárová Rosier J M Van Den Munckhof and Wilma M Frederiks |
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Institution: | (1) Department of Cell Biology and Histology, Academic Medical Centre, University of Amsterdam, P.O. Box 22700, 1100 DE Amsterdam, The Netherlands |
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Abstract: | Summary Glycogen phosphorylase activity has been demonstrated at the ultrastructural level in liver and heart tissue of fasted rats.
Unfixed cryostat sections were incubated by mounting them on a semipermeable membrane stretched over a gelled incubation medium.
The medium contained a high concentration of glucose 1-phosphate which enables indirect detection of glycogen phosphorylase
activity on the basis of the synthesis of glycogen. Tissue fixation, dehydration and embedding for electron microscopical
study were performed after the incubation had been completed.
The ultrastructure of both liver and heart tissue was rather well preserved. Glycogen granules resulting from glycogen phosphorylase
activity were found in the cytoplasmic matrix of both hepatocytes and cardiomyocytes; no relationship with membranous structures
could be detected. It is concluded that the semipermeable membrane method is well suited for localizing cytosolic enzyme activities
at the ultrastructural level without prior tissue fixation; this opens further perspectives for correlations between histochemical
and biochemical data. |
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