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. Author manuscript; available in PMC: 2012 Jul 1.
Published in final edited form as: J Neurochem. 2011 May 19;118(1):140–152. doi: 10.1111/j.1471-4159.2011.07289.x

Fig. 5. Hypoosmotic cell swelling suppresses enzymatic activity of glutamine synthetase (GS) and glutaminase (GLNase) in intact cultured astrocytes.

Fig. 5

(a) The activity of GS was measured as enzymatic production of L-[3H]glutamine from L[3H]glutamate in intact astrocytes incubated under basal, HYPO and Low-Na conditions. To prevent enzymatic hydrolysis of L-[3H]glutamine by GLN and loss of L[3H]glutamate via VRAC, cells were preincubated with 1 mM DON, and 20 μM DCPIB was additionally added to all reaction media. Intracellular L-[3H]glutamine and L[3H]glutamate were extracted, separated, and quantified as described in Materials and methods. Pretreatment of cells with 1 mM MSO was used as a positive control for GS activity. Data are the mean values ± SE of 6–9 experiments. ***p<0.001 vs. basal. (b) The activity of glutaminase was analyzed in similar fashion but cells were preincubated with the irreversible inhibitor of GS 1 mM MSO. 20 μM DCPIB was added to all reaction media. Pretreatment of cells with 1 mM DON was used as a positive control for GLNase activity. Data are the mean values ± SE of 5–6 experiments. *p<0.05, ***p<0.001 vs. basal. Inset in each figure briefly summarizes experimental design.