Fig. 1

Effect of AA147 and AA263 on the total and surface protein levels of trafficking-deficient mutant GABA_A receptors. A Dose–response analysis of AA147 and AA263 treatment (24 h) on the total protein levels of α1(D219N) subunits in HEK293T cells expressing α1(D219N)β2γ2 GABA_A receptors. B Time-course analysis of AA147 (5 µM) and AA263 (5 µM) treatment on the total protein levels of α1(D219N) subunits in HEK293T cells expressing α1(D219N)β2γ2 GABA_A receptors. C Effect of AA147 (2.5 µM, 24 h) and AA263 (2.5 µM, 24 h) on the total protein level of α1(D219N) subunits in neuronal SH-SY5Y cells stably expressing α1(D219N)β2γ2 GABA_A receptors. β-actin serves as total protein loading control. D Effect of AA147 (5 µM, 24 h) and AA263 (5 µM, 24 h) on the surface protein expression of the α1(D219N) subunits in HEK293T cells stably expressing α1(D219N)β2γ2 GABA_A receptors according to surface biotinylation analysis. E Effect of AA147 (2.5 µM, 24 h) and AA263 (2.5 µM, 24 h) on the surface protein expression of the α1(D219N) subunits in neuronal SH-SY5Y cells stably expressing α1(D219N)β2γ2 GABA_A receptors according to surface biotinylation analysis. Na⁺/K⁺-ATPase serves as membrane protein loading control. Quantification of the band intensities is shown on the bottom panels (n = 3). F HEK293T cells stably expressing α1(D219N)β2γ2 receptors were treated with DMSO vehicle control, AA147 (5 µM, 24 h) or AA263 (5 µM, 24 h). Surface α1 staining was in green (column 1), and plasma membrane marker Na⁺/K⁺-ATPase staining in red (column 2). Merge of these two signals and nucleus staining by DAPI in blue was shown in column 3. Scale bar = 20 μm. Quantification of the fluorescence intensity of the surface subunits from 30–40 cells per condition is shown on the right. IB: immunoblotting. Each data point is reported as mean ± SEM. One-way ANOVA followed by post-hoc Tukey test was used for statistical analysis. *, p < 0.05; **, p < 0.01; ***, p < 0.001