Hsp90β negatively regulates GLT-1 protein levels. (A) Western blot of astrocytes after lentivirus-mediated Hsp90α or Hsp90β overexpression for 72 h. Lentivirus expressing GFP was used as a control (left). Bar graph shows relative fold change of GLT-1 (right; normalized to actin; n = 3; *, P < 0.05; **, P < 0.01; ***, P < 0.001, Student’s t test). (B) Western blot of astrocytes at 48 h after transfection with two independent siRNAs against Hsp90 (40 nM; left). Bar graph shows relative fold change of GLT-1 and Hsp90β (right; normalized to actin; n = 3; *, P < 0.05; ***, P < 0.001, Student’s t test). (C and D) Western blot and statistical analysis of astrocytes with treatment of (C) GA or (D) 17AAG at different concentrations for 48 h (n = 3 for each group; *, P < 0.05; **, P < 0.01; ***, P < 0.001, one-way ANOVA, post hoc Dunnett's test). (E and F) Expression of GLT-1 mRNA levels in astrocytes with treatment of 17AAG (100 nM, 24 h) and Hsp90 siRNA (40 nM, 48 h; Student’s t test). NTC, nontarget control. Data are representative of at least two independent experiments.