Journal: Antioxidants

Article Title: Erythroid Differentiation Regulator 1 as a Regulator of Neuronal GSH Synthesis

doi: 10.3390/antiox13070771

Figure Lengend Snippet: Increased EAAC1 gene and protein expression in Neuro2a cells with Erdr1 siRNA knockdown. ( a ) Confocal images showing the effect of transfection with a specific siRNA targeting Erdr1 (siRNA) on the intensity of EAAC1 expression in Neuro2a cells compared with that in the negative control cells (scrambled siRNA). Scale bar, 100 μm. Insets in the panels are enlarged images corresponding to the box highlighted on the full images. Scale bar, 20 μm. ( b ) Quantification of relative EAAC1 densities in Neuro2a cells is shown. n = 5; 5–6 areas were measured in each sample. ** p < 0.01. ( c ) Protein expression of EAAC1 in Neuro2a cells transfected with a specific siRNA targeting Erdr1 (siRNA) and in the negative control cells (scrambled siRNA). Quantification of the EAAC1 protein expression (data in the left panel) by densitometry is shown in the right panel. n = 6. * p < 0.05.

Article Snippet: Non-specific staining was blocked with the reagent PBS containing 5% BSA, and the cells were incubated with anti-EAAC1 (Alomone Labs) at 1:1000 dilution for 1 h. After a wash with PBS-Tween20, the cells were labeled with fluorescent-labeled secondary antibodies Alexa-Fluor 488 anti-rabbit IgG (Invitrogen) at 1:1000 dilutions for 30 min.

Techniques: Expressing, Knockdown, Transfection, Negative Control

Journal: Antioxidants

Article Title: Erythroid Differentiation Regulator 1 as a Regulator of Neuronal GSH Synthesis

doi: 10.3390/antiox13070771

Figure Lengend Snippet: The schematic illustration indicates the relationship between Erdr1, GTRAP3-18, EAAC1, and GSH in the brain/hippocampus and Neuro2a/primary hippocampal neurons. In the brain/hippocampus of GTRAP3-18-deficient mice, GTRAP3-18 may be regulated by Erdr1 and exert compensatory feedback on Erdr1 expression. In Neuro2a/primary hippocampal neurons, knockdown of Erdr1 resulted in a decrease in GTRAP3-18, upregulation of EAAC1 expression, leading to an increase in GSH levels.

Article Snippet: Non-specific staining was blocked with the reagent PBS containing 5% BSA, and the cells were incubated with anti-EAAC1 (Alomone Labs) at 1:1000 dilution for 1 h. After a wash with PBS-Tween20, the cells were labeled with fluorescent-labeled secondary antibodies Alexa-Fluor 488 anti-rabbit IgG (Invitrogen) at 1:1000 dilutions for 30 min.

Techniques: Expressing, Knockdown

Journal: Neuropsychopharmacology : official publication of the American College of Neuropsychopharmacology

Article Title: Glutamatergic Dysfunction of Astrocytes in Paraventricular Nucleus of Thalamus Contributes to Adult Anxiety Susceptibility in Adolescent Ethanol Exposed Mice

doi: 10.1038/s41386-025-02264-3

Figure Lengend Snippet: (a) The quantification of glutamate levels in the dorsal thalamic area including PVT. (b-d) Representative expression of GLT1 (as known as EAAT2), GLAST (as known as EAAT1), and GAPDH (b) and pooled data (c-d) of western blots showing that the GLT1 expression in the PVT of AIE mice is selectively reduced compared to that of the CON mice. (e-f) Representative figures (e) and pooled data (f) confirming the reduction of GLT1 in the conditional GLT1 knockdown mice (GLT1 cHET ). (g-h) Representative figures (g) and pooled data (h) showing the selective reduction of GLT1 in astrocytes, not in neurons of PVT. (i-k) Non-invasive magnetic resonance spectroscopy (MRS) measurement showing the increase in glutamate levels in the dorsal thalamic area including PVT of GLT1 cHET . (l-q) Representative traces (l, o) and pooled data (m-n, p-q) showing the anxiogenic profiles of GLT1 cHET in the open field test (l-n) and elevated plus maze test (o-q). Female (circles) and male (triangles). Data represented as mean ± SEM. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001. Figure 3i , 3l , and 3o were created using BioRender.com .

Article Snippet: These membranes were incubated with anti-GLT1 antibody (1:2000, Guinea pig, Millipore Sigma Cat# AB1783, RRID:AB_90949), anti-GLAST antibody (1:2000, Rabbit, Alomone Labs Cat# AGC-021, RRID:AB_2039885), anti-GAPDH antibody (1:2000, Mouse, Sigma Aldrich Cat# MAB374, RRID:AB_2107445), and anti-xCT antibody (1:1000, Rabbit, ABclonal Cat# A2413, RRID:AB_2863004) overnight at 4 °C.

Techniques: Expressing, Knockdown, Western Blot, Spectroscopy