VALPROATE INCREASES GLUTAMINASE AND DECREASES GLUTAMINE-SYNTHETASE ACTIVITIES IN PRIMARY CULTURES OF RAT-BRAIN ASTROCYTES

It has been proposed that hyperammonemia may be associated with valpro ate therapy. As astrocytes are the primary site of ammonia detoxificat ion in brain, the effects of valproate on glutamate and glutamine meta bolism in astrocytes were studied. It is well established that, becaus e of compartmentation of glutamine synthetase, astrocytes are the site of synthesis of glutamine from glutamate and ammonia. The reverse rea ction is catalyzed by the ubiquitous enzyme glutaminase, which is pres ent in both neurons and astrocytes. In astrocytes exposed to 1.2 mM va lproate, glutaminase activity increased 80% by day 2 and remained elev ated at day 4; glutamine synthetase activity was decreased 30%. Direct addition of valproate to assay tubes with enzyme extracts from untrea ted astrocytes had significant effects only at concentrations of 10 an d 20 mM. When astrocytes were exposed for 4 days to 0.3, 0.6, or 1.2 m M valproate and subsequently incubated with L-[U-C-14]glutamate, label incorporation into [C-14]glutamine was decreased by 11, 25, and 48%, respectively, and is consistent with a reduction in glutamine syntheta se activity. Label incorporation from L[U-C-14]glutamate into [C-14]as partate also decreased with increasing concentrations of valproate. Fo llowing a 4-day exposure to 0.6 mM valproate, the glutamine levels inc reased 40% and the glutamate levels 100%. These effects were not direc tly proportional to valproate concentration, because exposure to 1.2 m M valproate resulted in a 15% decrease in glutamine levels and a 25% i ncrease in glutamate levels compared with control cultures. Intracellu lar aspartate was inversely proportional to all concentrations of extr acellular valproate, decreasing 60% with exposure to 1.2 mM valproate. These results indicate that valproate increases glutaminase activity, decreases glutamine synthetase activity, and alters Krebs-cycle activ ity in astrocytes, suggesting a possible mechanism for hyperammonemia in brain during valproate therapy.