Journal: International Journal of Molecular Sciences

Article Title: Pregabalin Depresses Cerebellar Parallel Fiber–Purkinje Cell Synaptic Transmission by Modulating Glun2a-Containing Nmda Receptors in Mice In Vitro

doi: 10.3390/ijms27114660

Figure Lengend Snippet: The inhibitory effect of PGB on PF–PC synaptic transmission persisted following blockade of postsynaptic NMDA receptors but was abolished by disruption of the α 2 δ-1–NMDAR interaction. ( A ) Representative traces of evoked EPSCs in response to paired-pulse stimulation (0.2 ms pulse duration, 50 ms inter-pulse interval) under control conditions, during pregabalin (PGB; 10 μM) application, and following washout (recovery). Recordings were performed with MK-801-containing pipette solution (left) or following extracellular application of the α2δ-1 Tat peptide (right; 1 μM). ( B ) Pooled data showing normalized N1 amplitude under these conditions. ( C , D ) Bar graphs with individual data points illustrating normalized N1 amplitude ( C ) and paired-pulse ratio (PPR; ( D )) under these conditions. * p < 0.05 versus control; n = 8 cells/8 slices/5 mice for the MK-801 group; n = 8 cells/8 slices/6 mice for the α2δ-1 Tat peptide group.

Article Snippet: The α 2 δ-1 Tat peptide (VSGLNPSLWSIFGLQFILLWLVSGSRHYLW) was purchased from MedChemExpress LLC. (Shanghai, China).

Techniques: Transmission Assay, Disruption, Control, Transferring

Journal: International Journal of Molecular Sciences

Article Title: Pregabalin Depresses Cerebellar Parallel Fiber–Purkinje Cell Synaptic Transmission by Modulating Glun2a-Containing Nmda Receptors in Mice In Vitro

doi: 10.3390/ijms27114660

Figure Lengend Snippet: The effect of PGB on mEPSCs was prevented by blockade of GluN2A-containing NMDA receptors or disruption of α 2 δ-1–NMDA receptor coupling. ( A ) In the presence of a mixture of gabazine (20 μM), TTX (1 μM), and α2δ-1Tat peptide (1 μM), representative membrane current traces of a cerebellar PC recorded under control conditions, during pregabalin (PGB; 10 μM) application, and following washout (recovery). ( B , C ) In the presence of α2δ-1Tat peptide, cumulative probability–interevent interval curves ( B ) and cumulative probability–amplitude curves ( C ) of mEPSCs under control, PGB treatment, and recovery conditions. ( D , E ) Bar graphs showing the mean ± S.E.M. and individual data points for the normalized mEPSC frequency ( D ) and amplitude ( E ) in PCs in the presence of α2δ-1Tat peptide (1 μM; left panel) and PEAQX (1 µM; right panel) under each condition. n = 7 cells/7 slices/4 mice in PEAQX group; n = 7 cells/7 slices/6 mice in α2δ-1Tat peptide group.

Article Snippet: The α 2 δ-1 Tat peptide (VSGLNPSLWSIFGLQFILLWLVSGSRHYLW) was purchased from MedChemExpress LLC. (Shanghai, China).

Techniques: Disruption, Membrane, Control

Journal: International Journal of Molecular Sciences

Article Title: Pregabalin Depresses Cerebellar Parallel Fiber–Purkinje Cell Synaptic Transmission by Modulating Glun2a-Containing Nmda Receptors in Mice In Vitro

doi: 10.3390/ijms27114660

Figure Lengend Snippet: Co-expression of α 2 δ-1 and GluN2A subunits in the molecular layer of the mouse cerebellum. ( A ) Left panel: confocal micrograph showing DAPI staining (blue) in the mouse cerebellar cortex; right panel: magnified view of DAPI staining (blue) corresponding to the boxed area in the left panel. DAPI is a blue nucleic acid dye that preferentially stains double-stranded DNA (dsDNA) in cells. ( B ) Higher-magnification images of the boxed area in the right panel of ( A ), showing DAPI staining (blue), α 2 δ-1 subunit immunoreactivity (red), GluN2A immunoreactivity (green), and the merged immunofluorescence image in the Purkinje cell layer (PCL) and molecular layer (ML). Red fluorescence signals for the α 2 δ-1 subunit and green fluorescence signals for GluN2A immunoreactivity were detected in the ML, surrounding the dendrites of PCs. ML, molecular layer; PCL, Purkinje cell layer; GCL, granular cell layer. ( C ) Co-immunoprecipitation analysis revealed the protein–protein interaction between α 2 δ-1 and NMDAR in membrane extracts from the molecular layer of the cerebellar cortex. Proteins were first immunoprecipitated with rabbit anti-GluN1, anti-α 2 δ-1, or control IgG antibodies. Western immunoblotting (IB) was then performed using mouse anti-α 2 δ-1 (upper panel) and anti-GluN2A antibodies (lower panel). IgG and input samples (tissue lysates without immunoprecipitation) served as the negative and positive controls, respectively.

Article Snippet: The α 2 δ-1 Tat peptide (VSGLNPSLWSIFGLQFILLWLVSGSRHYLW) was purchased from MedChemExpress LLC. (Shanghai, China).

Techniques: Expressing, Staining, Immunofluorescence, Fluorescence, Immunoprecipitation, Membrane, Control, Western Blot

Journal: Scientific Reports

Article Title: Prediction of SARS-CoV-2 exposure through T cell activation profiles

doi: 10.1038/s41598-026-48323-7

Figure Lengend Snippet: Comparison of naïve and COVID-19 convalescent donor T cell activation for select AIM pairs. ( a ) Flow cytometry plots comparing naïve and COVD-19 convalescent donor PBMCs with or without SARS-CoV-2 peptide pool stimulation . Five CD4 + T cell ( b ) and five CD8 + T cell ( c ) AIM pairs discriminate naïve (circle) and COVID-19 convalescent (square) donors. ( b , c ) Graphs indicate significant differential expression between naïve and COVID-19 convalescent donors, at 5% FDR (*), ns = not significant.

Article Snippet: Curated SARS-CoV-2 peptide pools (130-127-309, Miltenyi and 3622-1, Mabtech) containing 9-22 mer peptides were used to activate SARS-CoV-2 antigen-responsive CD4 + and CD8 + T cells.

Techniques: Comparison, Activation Assay, Flow Cytometry, Quantitative Proteomics

Journal: Scientific Reports

Article Title: Prediction of SARS-CoV-2 exposure through T cell activation profiles

doi: 10.1038/s41598-026-48323-7

Figure Lengend Snippet: Comparison of naïve and COVID-19 convalescent donor T cell activation using fold-change and difference analyses. ( a ) Difference analysis for AIM pairs identified from the CD4 + T cell population; wherein, the median background subtracted activation marker pair signal from the naïve donor samples was subtracted from the background subtracted activation AIM pair signals detected from each COVID convalescent donor sample. ( b ) Fold-change analysis for marker pairs identified from the CD4 + T cell population; wherein, the background subtracted activation AIM pair signals detected from each COVID convalescent donor sample were divided by the median background subtracted activation marker pair signal from the naïve donor samples. ( c ) Difference analysis for AIM pairs identified from the CD8 + T cell population, analyzed similarly to the CD4 + T cell population. ( d ) Fold-change analysis for marker pairs identified from the CD8 + T cell population, analyzed similarly to the CD4 + T cell population. Each data point represents cells from a single COVID-19 convalescent donor treated with a single SARS-CoV-2 peptide pool in comparison to the median signal from the naïve donors. For all graphs, each unique symbol color represents data from a unique COVID-19 convalescent donor.

Article Snippet: Curated SARS-CoV-2 peptide pools (130-127-309, Miltenyi and 3622-1, Mabtech) containing 9-22 mer peptides were used to activate SARS-CoV-2 antigen-responsive CD4 + and CD8 + T cells.

Techniques: Comparison, Activation Assay, Marker

Journal: Scientific Reports

Article Title: Prediction of SARS-CoV-2 exposure through T cell activation profiles

doi: 10.1038/s41598-026-48323-7

Figure Lengend Snippet: Cytokine production is increased in COVID-19 convalescent donors following stimulation with SARS-CoV-2 peptide pools. ( a ) IFN-γ cytokine production is increased in COVID-19 convalescent donor samples following stimulation with SARS-CoV-2 pp B as monitored via an Enzymatic 2-step ELISpot assay. Shown are a graph and a representative image of ELISpot results. Note: due to a technical issue, pp A failed to elicit an ELISpot response in 2 experiments, even for positive controls, which impacted the naïve to COVID-19 convalescent comparison for the pp A ELISpot data. ( b ) IFN-γ, IL-2, IL-6, and IL-10 cytokine production is increased in COVID-19 convalescent donor samples following SARS-CoV-2 peptide stimulation as monitored via a bead-based flowcytometry assay. ( a, b ) Graphs indicate significant differential expression between naïve and COVID-19 convalescent donors, at 5% FDR (*), ns = not significant. ( c ) Spearman correlation analysis of AIM pairs described in Fig. that significantly discriminated naïve and COVID-19 convalescent samples with cytokine release assay and ELISpot.

Article Snippet: Curated SARS-CoV-2 peptide pools (130-127-309, Miltenyi and 3622-1, Mabtech) containing 9-22 mer peptides were used to activate SARS-CoV-2 antigen-responsive CD4 + and CD8 + T cells.

Techniques: Enzyme-linked Immunospot, Comparison, Quantitative Proteomics, Release Assay

Journal: Scientific Reports

Article Title: Prediction of SARS-CoV-2 exposure through T cell activation profiles

doi: 10.1038/s41598-026-48323-7

Figure Lengend Snippet: Unsupervised hierarchical clustering of all 35 CD4 + and CD8 + T cell AIM pair combinations. Using data from normalized samples, unsupervised hierarchical clustering of all 35 CD4 + and CD8 + T cell marker pair combinations separated naïve and COVID-19 convalescent donor samples, except for a single COVID-19 convalescent donor sample stimulated with SARS-CoV-2 pp A (blue arrow). The same donor sample stimulated with SARS-CoV-2 pp B clustered with the COVID-19 convalescent donor samples. Obtaining this clear separation of naïve and convalescent donor samples by chance is very unlikely, p < 2 -10 .

Article Snippet: Curated SARS-CoV-2 peptide pools (130-127-309, Miltenyi and 3622-1, Mabtech) containing 9-22 mer peptides were used to activate SARS-CoV-2 antigen-responsive CD4 + and CD8 + T cells.

Techniques: Marker