Journal: bioRxiv

Article Title: GluN2D-containing NMDA receptors regulate dentate gyrus function by facilitating granule cell activity and mediating synaptic plasticity

doi: 10.64898/2026.03.06.710109

Figure Lengend Snippet: (A) Grin2d f l/fl mice were injected with AAV5-CamKII-mCherry (Control) or AAV-CamKII-mCherry-Cre ( Grin2d cKO). NMDAR-LTP was abolished in Grin2d cKO compared with control mice (Control: 149.5 ± 6.0 %, p < 0.01, n = 5, paired t-test; cKO: 92.5 ± 5.3 %, p = 0.12201, n = 6, paired t-test; Control vs cKO: p < 0.001, unpaired t-test). (B) WT mice were bilaterally injected with an anti-GluN2D antibody or control Ab into the dentate gyrus. After one hour, animals were euthanized, and slices were prepared. Injection was confirmed by the presence of methylene blue. NMDAR-LTP was abolished in mice injected with the anti-GluN2D antibody (cKO: 110.4 ± 8.5 %, p = 0.2952, n = 6, paired t-test) compared with control mice (Control: 149.8 ± 8.1 %, p < 0.001, n = 7, paired t-test; Control vs cKO: p < 0.01, unpaired t-test). (C) NMDAR-LTP was impaired in Grid1 KO mice (KO: 117.7 ± 5.3, p < 0.05%, n = 8, Wilcoxon signed-rank test) compared with controls (Control: 147.5 ± 6.7 %, p < 0.001, n = 7, paired t-test; Control vs cKO: p < 0.05, Mann-Whitney U test). Data are presented as mean ± s.e.m.

Article Snippet: For GluN2D cross-linking experiments in C57BL/6J, the control group received 1 μL of anti-rabbit Alexa 568 (control IgG, 1/5), while the GluN2D-cross-link group received 1 μg of polyclonal rabbit anti-GluN2D subunit (Alomone Labs, cat #AGC-020), both diluted in PBS with 1% methylene blue (1 μL final volume).

Techniques: Injection, Control, MANN-WHITNEY

Journal: Journal of Affective Disorders Reports

Article Title: Long-term cyclosporine A treatment promotes anxiety-like behavior: Possible relation with glutamate signaling in rat hippocampus

doi: 10.1016/j.jadr.2022.100394

Figure Lengend Snippet: Fig. 4. Western blot analysis of NMDA receptors in the hippocampus of control and CsA-treated rats. (A) Representative images of the levels of total (pan) GluN2, GluN2A, GluN2B, p-GluN1 (Ser897) and total GluN1. (B-F) Respective quantifications of immunodetections normalized to β-actin and presented as a percentage of the control mean. Data represent the mean ± SEM. Statistics: ** *p ≤0.001 as determined by Student’s t-test; n = 6.

Article Snippet: The utilized primary antibodies and respective dilutions were as follows: anti-AMPAR (1:500, #13185 Cell Signaling Technology), phosphorylated AMPAR (pAMPAR-S831) (1:1000, A4352-Sigma–Aldrich), pAMPAR (S845) (1:1000, #8084 – Cell Signaling Technology), GluN1 (1:750, #G8913 – Sigma–Aldrich), pan-GluN2 (1:750, 244-0P – SYSY Synaptic Systems), GluN2A (1:1000, #4205 - Cell Signaling Technology), GluN2B (1:1000, #4207 - Cell Signaling Technology), pGluN1 (Ser897) (1:500, 3385S – Cell Signaling Technology), nNOS (1:500, SC648 – Santa Cruz Biotechnology), and iNOS (1:750, SC7271 – Santa Cruz Biotechnology). β-actin antibody (1:5000, A5441 – Sigma-Aldrich) was used as a loading control in the experiments.

Techniques: Western Blot, Control

Journal: Frontiers in Neuroscience

Article Title: Spatiotemporal differential regulation of extrasynaptic GluN2B receptor subunits and PSA-NCAM in brain aging and Alzheimer’s disease

doi: 10.3389/fnins.2025.1649625

Figure Lengend Snippet: Regional expression changes in GluN2A, GluN2B, and GABA A R in WT and AD mice with aging. (A) Representative immunoblots showing GluN2A, GluN2B, and GABA A R expression across the hippocampus, prefrontal cortex, cortex, midbrain, and cerebellum in young and old WT mice. GluN2A expression remained largely stable with aging, except for an increase in the prefrontal cortex. GluN2B expression decreased with age across most regions, while GABA A R expression was reduced in the hippocampus and cortex in old WT mice. (B) Representative immunoblots from young and old AD mice showing GluN2A, GluN2B, and GABA A R levels across the same regions. GluN2A expression increased with aging in nearly all regions, while GluN2B and GABA A R levels decreased broadly.

Article Snippet: GluN2B , AB_1264223 , NMDA Receptor 2B (GluN2B) Antibody #4207 , H, M, R , (Cell Signaling Technology Cat# 4207, RRID: AB_1264223) , polyclonal , rabbit.

Techniques: Expressing, Western Blot

Journal: Frontiers in Neuroscience

Article Title: Spatiotemporal differential regulation of extrasynaptic GluN2B receptor subunits and PSA-NCAM in brain aging and Alzheimer’s disease

doi: 10.3389/fnins.2025.1649625

Figure Lengend Snippet: Comparative Analysis of GluN2A, GluN2B, and GABA A R Receptor Expression in Young and Old WT vs AD Mice. (A,B) Western blot analysis comparing GluN2A levels between WT and AD mice showed increased GluN2A expression in hippocampal and cortical regions of old WT mice compared to AD and a slight decrease in the prefrontal cortex of young AD mice. (C,D) No significant change region-wide between ages, except for an observed decrease in the prefrontal cortex of young AD mice. (E,F) Downregulated GABA A R expression is seen in old AD mice across all regions compared to WT. For each brain region, n = 4–5 mice per group. Bar graphs depict the mean ± SD normalized to β-actin. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: GluN2B , AB_1264223 , NMDA Receptor 2B (GluN2B) Antibody #4207 , H, M, R , (Cell Signaling Technology Cat# 4207, RRID: AB_1264223) , polyclonal , rabbit.

Techniques: Expressing, Western Blot

Journal: Frontiers in Neuroscience

Article Title: Spatiotemporal differential regulation of extrasynaptic GluN2B receptor subunits and PSA-NCAM in brain aging and Alzheimer’s disease

doi: 10.3389/fnins.2025.1649625

Figure Lengend Snippet: Increased extrasynaptic localization of GluN2B subunits with disease progression in AD mice and comparative analysis of ES-GluN2B and synaptic GluN2B expression in midbrain and cortex between WT and AD mice. (A,B) Immunoblot analysis showing significant increases in ES-GluN2B levels across brain regions of AD mice. Quantification reveals a marked increase in ES-GluN2B expression in these regions, indicating a shift from synaptic to extrasynaptic localization with disease. No significant changes in the cerebellum of AD mice. (C,D) Results demonstrate significant increases in ES-GluN2B levels in old AD mice compared to old WT mice, emphasizing the disease-dependent shift toward extrasynaptic localization, which becomes more pronounced with aging in these brain regions. Bar graphs represent the mean ± SD of ES-GluN2B levels normalized to Synaptic GluN2B. For each brain region, n = 4–5 mice per group. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: GluN2B , AB_1264223 , NMDA Receptor 2B (GluN2B) Antibody #4207 , H, M, R , (Cell Signaling Technology Cat# 4207, RRID: AB_1264223) , polyclonal , rabbit.

Techniques: Biomarker Discovery, Expressing, Western Blot

Journal: Frontiers in Neuroscience

Article Title: Spatiotemporal differential regulation of extrasynaptic GluN2B receptor subunits and PSA-NCAM in brain aging and Alzheimer’s disease

doi: 10.3389/fnins.2025.1649625

Figure Lengend Snippet: Region-specific increases in GluN2B phosphorylation and casein kinase IIα (CK2α) expression in AD brain regions. (A,B) Immunoblot and quantification of CK2α protein levels across brain regions (hippocampus, prefrontal cortex, cortex, and midbrain) of aged AD mice compared to aged WT mice. CK2α expression significantly increases with age in AD mice, whereas WT mice exhibit stable or reduced levels. n = 4–5 per group. (C,D) Western blot and quantification showing significant increases in phosphorylation of GluN2B at Ser1480 (pGluN2B) across brain regions (hippocampus, prefrontal cortex, cortex, and midbrain) of aged AD mice compared to aged WT mice. Old AD mice also showed higher pGluN2B levels than their WT counterparts in all regions. The ratio of pGluN2B to total GluN2B is markedly elevated in old AD mice, supporting increased extrasynaptic signaling activity with disease progression. n = 4–5 per group. All bar graphs depict mean ± SD of protein levels normalized to loading controls (GluN2B or β-actin). * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: GluN2B , AB_1264223 , NMDA Receptor 2B (GluN2B) Antibody #4207 , H, M, R , (Cell Signaling Technology Cat# 4207, RRID: AB_1264223) , polyclonal , rabbit.

Techniques: Phospho-proteomics, Expressing, Western Blot, Activity Assay, Biomarker Discovery

Journal: Frontiers in Neuroscience

Article Title: Spatiotemporal differential regulation of extrasynaptic GluN2B receptor subunits and PSA-NCAM in brain aging and Alzheimer’s disease

doi: 10.3389/fnins.2025.1649625

Figure Lengend Snippet: Region-specific expression of polysialylation enzymes ST8Sia2 and ST8Sia4 in AD and normal aging. (A,B) Western blot images and quantitative analysis of ST8Sia4 protein expression in the hippocampus, prefrontal cortex, cortex, and midbrain of young and aged WT and AD mice. ST8Sia4 expression significantly increased with age in WT mice but declined in aged AD mice across all regions, notably in the prefrontal cortex and cortex. Similarly, ST8Sia4 showed a significant decrease in aged AD mice compared to aged WT mice. (C,D) ST8Sia2 protein levels showed no significant regional differences. No significant changes were observed across age groups region-wide. (E) Parental IMR32 cells are unmodified. In gRNA-ST conditions, IMR32 cells are transfected with plasmid DNA that expresses dCas9-VP64 and the guide RNA targeting the ST8sia4 promoter, enabling CRISPR-mediated transcriptional activation of ST8sia4. (F) Western blot analysis revealed significantly higher GluN2B phosphorylation at Ser1480 in parental cells compared to PSA–NCAM–overexpressing cells (gRNA-ST), (G) increased PSA-NCAM expression in gRNA-ST cells, (H) with no significant difference in CKIIα levels. Bar graphs depict mean ± SD of protein levels normalized to loading controls (GluN2B or β-actin), n = 3 per group. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: GluN2B , AB_1264223 , NMDA Receptor 2B (GluN2B) Antibody #4207 , H, M, R , (Cell Signaling Technology Cat# 4207, RRID: AB_1264223) , polyclonal , rabbit.

Techniques: Expressing, Western Blot, Transfection, Plasmid Preparation, CRISPR, Activation Assay, Phospho-proteomics

Journal: Frontiers in Neuroscience

Article Title: Spatiotemporal differential regulation of extrasynaptic GluN2B receptor subunits and PSA-NCAM in brain aging and Alzheimer’s disease

doi: 10.3389/fnins.2025.1649625

Figure Lengend Snippet: Summary of NMDA-GABA receptor-mediated E/I Balance in Normal Aging and AD. The figure illustrates the shift in E/I balance during normal aging (top) and Alzheimer’s disease (bottom). In normal aging, the balance is maintained by decreased total GluN2B level and increasing GluN2A subunit expression. In contrast, AD shows a significant decrease in GluN2B expression and an upregulation of GluN2A, alongside a marked reduction in GABA A R expression, reflecting diminished inhibitory signaling and overall E/I imbalance. Additionally, increased ES-GluN2B localization further contributes to synaptic dysfunction and excitotoxicity in AD.

Article Snippet: GluN2B , AB_1264223 , NMDA Receptor 2B (GluN2B) Antibody #4207 , H, M, R , (Cell Signaling Technology Cat# 4207, RRID: AB_1264223) , polyclonal , rabbit.

Techniques: Expressing