Effect of glucose deprivation on rat glutamine synthetase in cultured astrocytes.
Glutamine synthetase was purified from the cerebral cortex of adult rats and characterized. Polyclonal rabbit antibodies were raised against the enzyme, purified and their specific anti-(glutamine synthetase) activity determined. A primary astroglial culture was prepared from newborn Sprague-Dawley rats. Astrocytes at different ages of development were incubated in the presence and absence of glucose. In glucose-deprived conditions the specific activity of glutamine synthetase decreased. This decrease was more pronounced in 8-day-old than in 21-day-old cultures. Kinetic analysis demonstrated that the reduction in activity was mainly related to a decrease in Vmax. By immunoprecipitation, it was shown that the number of enzyme molecules in astrocytes was decreased in glucose-deprived conditions. On addition of glucose, a total recovery of glutamine synthetase was obtained after 36 h in 8-day-old culture. Rates of degradation and synthesis were investigated. When compared with an incubation in the presence of glucose, glucose deprivation increased enzyme turnover, as estimated from the first-order disappearance of radioactivity from glutamine synthetase. Synthesis rate was estimated from the incorporation of [35S]methionine during a 2 h incubation period and was decreased in glucose-deprived conditions. Trichloroacetate-precipitable proteins changed only slightly in the experimental conditions, and total protein did not vary significantly during the experimental period. A mathematical model is presented which attempts to integrate degradation and synthesis in our experimental model.
Rosier
F
F
Laboratory of Pharmacology and Physiology, Facultés Universitaires, Namur, Belgium.
Lambert
D
D
Mertens-Strijthagen
M
M
D000367:Age Factors; D000818:Animals; D000831:Animals, Newborn; D000906:Antibodies; D001253:Astrocytes / Q000187:drug effects* / Q000201:enzymology*; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000201:enzymology; D005947:Glucose / Q000494:pharmacology*; D005974:Glutamate-Ammonia Ligase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D007700:Kinetics; D008954:Models, Biological; D011817:Rabbits; D051381:Rats
D000906:Antibodies; D005974:Glutamate-Ammonia Ligase; D005947:Glucose