FIG. 1.

Evidence of GS activity in LPS-activated microglia. (A) GS activity was measured in LPS-activated microglia at 16, 24, 48, and 72 h from the applied proinflammatory stimulus. The results are presented as mean ± S.D. of four independent experiments. Activity levels significantly higher than those measured at time zero are denoted as follows: ***p < 0.0001. (B) GS and β-actin protein level was assessed by Western blotting analysis on LPS-activated microglia harvested at the same time points. (C) Viability was tested on resting microglia incubated up to 48 h with increasing MSO concentrations, ranging from 1 to 5 mM. The results are presented as mean ± S.D. of five independent experiments. (D) LPS challenge was carried out for 24 h on microglial cells pretreated or not with 1 mM MSO. Intracellular (D) and extracellular (E) glutamine and glutamate levels and extracellular quinolinic acid (F) were measured by LC-MS/MS analysis. The results are presented as mean ± S.D. of four independent experiments. Glutamine/glutamate ratios and quinolinic acid levels are significantly higher than those measured in untreated control microglia cells and are denoted as follows: *p < 0.05, **p < 0.001, and ***p < 0.0001. GS, glutamine synthetase; LPS, lipopolysaccharide; MSO, methionine sulfoximine.