Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: GRIP1 regulates synaptic plasticity and learning and memory

doi: 10.1073/pnas.2014827117

Figure Lengend Snippet: GRIP1 is recruited into synapses during cLTP. (A) Representative Western blots of proteins from PSD and total cell lysates (total) isolated from rat cortical neurons treated with (+) or without (−) glycine. (B) Quantification of protein levels in PSD (n = 5; Mann–Whitney U test). (C) Quantification of protein levels in total cell lysates. (n = 8; Student’s t test). (D) Representative Western blots of GRIP1 in P2 and S2 fractions isolated from rat cortical neurons treated with (+) or without (−) glycine. (E) Quantification of GRIP1 protein level in each fraction following cLTP (n = 15 to 16; Mann–Whitney U test). (F) Model of GRIP1 translocation during cLTP. Data are presented as mean ± SEM. n.s., not significant. **P < 0.01.

Article Snippet: The following antibodies were used: anti–beta-tubulin mAb (Sigma-Aldrich), anti-GluA1 N-terminal antibody mAb (4.9D, made in-house), anti-GluA2 N-terminal antibody mAb (032.19.9, made in-house), anti-GluA2 phospho-S880 specific mAb (02.22.4, made in-house), anti-GluA2 phospho-Y876 specific mAb (045.10.5, made in-house), anti-PSD95 mAb (NeuroMab), anti-GluA3 pAb (JH4300, made in-house), anti-GRIP1 mAb (BD Biosciences), anti-GRIP1 pAb (Chemicon), and anti-GRIP1 pAb (JH2260, made in-house).

Techniques: Western Blot, Isolation, MANN-WHITNEY, Translocation Assay

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: GRIP1 regulates synaptic plasticity and learning and memory

doi: 10.1073/pnas.2014827117

Figure Lengend Snippet: Activity-dependent up-regulation of synaptic AMPARs requires GRIP1 expression. (A) Representative Western blots of proteins from PSD and total cell lysates from WT or Grip1 KO mouse neurons treated with (+) or without (−) glycine. (B) Quantification of protein levels in PSD under basal conditions in WT and Grip1 KO mouse neurons (n = 9 to 11; Student’s t test). (C) Quantification of protein levels in PSD from WT or Grip1 KO mouse neurons treated with (+) or without (−) glycine (n = 15; Mann–Whitney U test). Data are presented as mean ± SEM. n.s., not significant. *P < 0.05; **P < 0.01; ***P < 0.001.

Article Snippet: The following antibodies were used: anti–beta-tubulin mAb (Sigma-Aldrich), anti-GluA1 N-terminal antibody mAb (4.9D, made in-house), anti-GluA2 N-terminal antibody mAb (032.19.9, made in-house), anti-GluA2 phospho-S880 specific mAb (02.22.4, made in-house), anti-GluA2 phospho-Y876 specific mAb (045.10.5, made in-house), anti-PSD95 mAb (NeuroMab), anti-GluA3 pAb (JH4300, made in-house), anti-GRIP1 mAb (BD Biosciences), anti-GRIP1 pAb (Chemicon), and anti-GRIP1 pAb (JH2260, made in-house).

Techniques: Activity Assay, Expressing, Western Blot, MANN-WHITNEY

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: GRIP1 regulates synaptic plasticity and learning and memory

doi: 10.1073/pnas.2014827117

Figure Lengend Snippet: Grip1 KO mice exhibit impaired NMDAR-dependent LTP. (A) Representative Western blots of total cell lysates from hippocampus in Nestin-Grip1fl/fl mice (Grip1 KO) and control Grip1fl/fl littermates (WT). (B) Representative evoked EPSCs obtained from CA1 pyramidal neurons before and after LTP induction in response to 0.1-Hz stimulation of Schaffer collaterals. Dash lines represent baseline EPSCs. Solid lines represent EPSCs 40 min after LTP induction. (C) Averaged EPSC amplitudes normalized to baseline responses. Arrow indicates the pairing induction (200 pulses at 2 Hz paired with 0 mV depolarization). (D) Statistics of LTP at 30–50 min (n = 16 cells from 8 control Grip1fl/fl littermate group; n = 17 cells from 8 Nestin-Grip1fl/fl mice; Mann–Whitney test). (E) Averaged EPSC amplitudes at baseline and during induction. Data are presented as mean ± SEM *P < 0.05.

Article Snippet: The following antibodies were used: anti–beta-tubulin mAb (Sigma-Aldrich), anti-GluA1 N-terminal antibody mAb (4.9D, made in-house), anti-GluA2 N-terminal antibody mAb (032.19.9, made in-house), anti-GluA2 phospho-S880 specific mAb (02.22.4, made in-house), anti-GluA2 phospho-Y876 specific mAb (045.10.5, made in-house), anti-PSD95 mAb (NeuroMab), anti-GluA3 pAb (JH4300, made in-house), anti-GRIP1 mAb (BD Biosciences), anti-GRIP1 pAb (Chemicon), and anti-GRIP1 pAb (JH2260, made in-house).

Techniques: Western Blot, MANN-WHITNEY

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: GRIP1 regulates synaptic plasticity and learning and memory

doi: 10.1073/pnas.2014827117

Figure Lengend Snippet: Grip1 KO mice display impaired learning and memory. (A) Cartoon illustration of the IA task. (B) Quantifications of the latency to cross over to the dark chamber at training and 24 h later in CaMKII-Grip1fl/fl (Grip1 KO) and control Grip1fl/fl littermates (WT) (n = 15 WT; n = 13 Grip1 KO; Mann–Whitney U test). (C) Quantification of total, central, and peripheral ambulatory activities in open-field chambers (n = 15 WT; n = 13 Grip1 KO; Mann–Whitney U test). (D) Quantification of the anxiety index, calculated as the activity in the peripheral divided by the activity in the center for each mouse (n = 15 WT; n = 13 Grip1 KO; Student’s t test). Data are presented as mean ± SEM, n.s., not significant; ***P < 0.001.

Article Snippet: The following antibodies were used: anti–beta-tubulin mAb (Sigma-Aldrich), anti-GluA1 N-terminal antibody mAb (4.9D, made in-house), anti-GluA2 N-terminal antibody mAb (032.19.9, made in-house), anti-GluA2 phospho-S880 specific mAb (02.22.4, made in-house), anti-GluA2 phospho-Y876 specific mAb (045.10.5, made in-house), anti-PSD95 mAb (NeuroMab), anti-GluA3 pAb (JH4300, made in-house), anti-GRIP1 mAb (BD Biosciences), anti-GRIP1 pAb (Chemicon), and anti-GRIP1 pAb (JH2260, made in-house).

Techniques: MANN-WHITNEY, Activity Assay

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: GRIP1 regulates synaptic plasticity and learning and memory

doi: 10.1073/pnas.2014827117

Figure Lengend Snippet: The GRIP1–GluA2 association is enhanced during LTP. (A) Representative Western blots of proteins from P2 in rat cortical neurons treated with (+) or without (−) glycine. (B) Quantification of phospho-Y876 and phospho-S880 levels following cLTP (n = 7 to 12; Student’s t test). (C) GluA2 was immunoprecipitated with specific GluA2 antibody from P2 from rat cortical neurons treated with (+) or without (−) glycine, followed by Western blot analysis of GRIP1 and GluA2. (D) Quantification of relative GRIP1–GluA2 interactions in P2 during cLTP (n = 10; Mann–Whitney U test). (E) Model of GRIP1 regulation of AMPAR trafficking during LTP. Data are presented as mean ± SEM. n.s., not significant; *P < 0.05; ***P < 0.001.

Article Snippet: The following antibodies were used: anti–beta-tubulin mAb (Sigma-Aldrich), anti-GluA1 N-terminal antibody mAb (4.9D, made in-house), anti-GluA2 N-terminal antibody mAb (032.19.9, made in-house), anti-GluA2 phospho-S880 specific mAb (02.22.4, made in-house), anti-GluA2 phospho-Y876 specific mAb (045.10.5, made in-house), anti-PSD95 mAb (NeuroMab), anti-GluA3 pAb (JH4300, made in-house), anti-GRIP1 mAb (BD Biosciences), anti-GRIP1 pAb (Chemicon), and anti-GRIP1 pAb (JH2260, made in-house).

Techniques: Western Blot, Immunoprecipitation, MANN-WHITNEY

Journal: Frontiers in Molecular Neuroscience

Article Title: Secreted Amyloid Precursor Protein-Alpha Enhances LTP Through the Synthesis and Trafficking of Ca 2+ -Permeable AMPA Receptors

doi: 10.3389/fnmol.2021.660208

Figure Lengend Snippet: Secreted amyloid precursor protein-alpha enhances de novo GluA1 at the extrasynaptic membrane. Representative images showing de novo cell-surface GluA1 in (A) control and (B) sAPPα-treated conditions. Representative images show dendrites (50 μm; synapsin-1, ; GluA1, ). Scale bar = 10 μm. (C) No significant difference was observed in the Mander’s overlap coefficient following sAPPα-treatment (1 nM, 30 min; n = 20 cells, 58–65 dendrites). Significance was assessed by Mann-Whitney two-tailed U-test. (D) Frequency histogram of the distribution of de novo GluA1 puncta in relevance to synapsin-1 immunofluorescence. Distances were calculated for individual PLA puncta with respect to the closest synapsin-1 center of mass. Synaptic PLA showed overlapping centers within 0–2 μm, puncta within 2–4 μm were considered extrasynaptic, and puncta beyond 4 μm were considered non-synaptic ( n = 58–65 dendrites, 130–205 puncta). (E) Quantification of GluA1 puncta abundance at the synaptic, extrasynaptic, and non-synaptic membrane ( n = 58–65 dendrites, 130–205 puncta). Normality was determined by Shapiro-Wilk normality test. All data are expressed relative to control, as mean ± SEM from 3 experiments. Significance was assessed by two-way ANOVA and Šidák’s multiple comparisons, *** p ≤ 0.0005.

Article Snippet: For the detection of proteins in situ primary antibodies were used targeting Arc (rabbit polyclonal, 1:1000, synaptic systems, #156003), Biotin (mouse monoclonal, 1:1000, Sigma, #B7653), MAP2 (guinea-pig polyclonal, 1:1000, synaptic systems, #188004; monoclonal, 1:1000, Abcam, #AB11267), Synapsin-1 (mouse monoclonal, 1:1000, synaptic systems, #106011), GluA1 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB31232), GluA1 (N-terminal; mouse monoclonal, 1:500, #MAB2263), GluA2 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB1768-I), GluA2 (N-terminal; mouse polyclonal, 1:500, Abcam, #133477), and GluA3 (N-terminal; mouse monoclonal, 1:250, Thermofisher, #32-0400).

Techniques: MANN-WHITNEY, Two Tailed Test, Immunofluorescence

Journal: Frontiers in Molecular Neuroscience

Article Title: Secreted Amyloid Precursor Protein-Alpha Enhances LTP Through the Synthesis and Trafficking of Ca 2+ -Permeable AMPA Receptors

doi: 10.3389/fnmol.2021.660208

Figure Lengend Snippet: Secreted amyloid precursor protein-alpha enhances GluA1/2 AMPARs at synapses. Representative images showing cell-surface GluA1/2 AMPARs in (A) control and (B) sAPPα-treated conditions Representative images show dendrites (50 μm; synapsin-1, ; GluA1/2, ). Scale bar = 10 μm. (C) sAPPα treatment (1 nM, 2 h) showed a significant increase in Mander’s overlap coefficient ( n = 20 cells, 49–54 dendrites). Significance was assessed by unpaired t -test. (D) Frequency histogram of the distribution of GluA1/2 puncta in relevance to synapsin-1 immunofluorescence. Distances were calculated for individual PLA puncta with respect to the closest synapsin-1 center of mass. Synaptic PLA showed overlapping centers within 0–2 μm, puncta within 2–4 μm were considered extrasynaptic, and puncta beyond 4 μm were considered non-synaptic ( n = 49–54 dendrites, 291–389 puncta). (E) Quantification of GluA1/2 puncta abundance at the synaptic, extrasynaptic, and non-synaptic membrane ( n = 49–54 dendrites, 291–389 puncta). Normality was determined by Shapiro–Wilk normality test. All data are expressed relative to control, as mean ± SEM from 3 experiments. Significance was assessed by two-way ANOVA and Šidák’s multiple comparisons, * p = 0.0467, ** p ≤ 0.0029.

Article Snippet: For the detection of proteins in situ primary antibodies were used targeting Arc (rabbit polyclonal, 1:1000, synaptic systems, #156003), Biotin (mouse monoclonal, 1:1000, Sigma, #B7653), MAP2 (guinea-pig polyclonal, 1:1000, synaptic systems, #188004; monoclonal, 1:1000, Abcam, #AB11267), Synapsin-1 (mouse monoclonal, 1:1000, synaptic systems, #106011), GluA1 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB31232), GluA1 (N-terminal; mouse monoclonal, 1:500, #MAB2263), GluA2 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB1768-I), GluA2 (N-terminal; mouse polyclonal, 1:500, Abcam, #133477), and GluA3 (N-terminal; mouse monoclonal, 1:250, Thermofisher, #32-0400).

Techniques: Immunofluorescence

Journal: Frontiers in Molecular Neuroscience

Article Title: Secreted Amyloid Precursor Protein-Alpha Enhances LTP Through the Synthesis and Trafficking of Ca 2+ -Permeable AMPA Receptors

doi: 10.3389/fnmol.2021.660208

Figure Lengend Snippet: Rapid increase in cell-surface de novo GluA1 following treatment. Representative images showing cell-surface de novo GluA1 levels in the soma (top panels) and dendrites (lower panels) from 30 min (left) and 2 h (right) control and sAPPα-treated conditions. (A) Average data showing 1 nM sAPPα promotes an increase in the soma ( n = 50–71 cells) following 30 min but not 2 h. (B) Average data showing 1 nM sAPPα (30 min) promotes an increase in de novo GluA1 at the dendritic cell surface follow 30 min but not 2 h treatments ( n = 104–128). Outliers were removed from each experiment prior to amalgamation using Grubb’s tests, and normality was detected by D’Agostino and Pearson omnibus normality tests. All data are expressed relative to control, as mean ± SEM from 3 experiments. Significance relative to control was assessed by student’s t -test, significance between sAPPα-treated groups was assessed by Mann-Whitney two-tailed U-test. ** p ≤ 0.01, *** p ≤ 0.001. Representative images show neuronal soma (upper panels) and dendrites (lower panels; MAP2, ; GluA1, ; DAPI, ). Scale bars = 10 μm.

Article Snippet: For the detection of proteins in situ primary antibodies were used targeting Arc (rabbit polyclonal, 1:1000, synaptic systems, #156003), Biotin (mouse monoclonal, 1:1000, Sigma, #B7653), MAP2 (guinea-pig polyclonal, 1:1000, synaptic systems, #188004; monoclonal, 1:1000, Abcam, #AB11267), Synapsin-1 (mouse monoclonal, 1:1000, synaptic systems, #106011), GluA1 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB31232), GluA1 (N-terminal; mouse monoclonal, 1:500, #MAB2263), GluA2 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB1768-I), GluA2 (N-terminal; mouse polyclonal, 1:500, Abcam, #133477), and GluA3 (N-terminal; mouse monoclonal, 1:250, Thermofisher, #32-0400).

Techniques: MANN-WHITNEY, Two Tailed Test

Journal: Frontiers in Molecular Neuroscience

Article Title: Secreted Amyloid Precursor Protein-Alpha Enhances LTP Through the Synthesis and Trafficking of Ca 2+ -Permeable AMPA Receptors

doi: 10.3389/fnmol.2021.660208

Figure Lengend Snippet: Rapid decrease in de novo cell-surface GluA2 following treatment. Representative images showing cell-surface de novo GluA2 levels in the soma (top panels) and dendrites (lower panels) from 30 min (left) and 2 h (right) controls and sAPPα treated conditions. (A) Average data showing 1 nM sAPPα does not affect somatic ( n = 25–31 cells) de novo GluA2. (B) Average data showing 1 nM sAPPα (30 min) significantly decreases dendritic de novo GluA2 ( n = 71–76). All data are expressed relative to control, as mean ± SEM from 3 experiments. Outliers were removed from each experiment prior to amalgamation using Grubb’s tests, and normality was detected by D’Agostino and Pearson omnibus normality tests. Significance was assessed by student’s t -test, ** p ≤ 0.01. Representative images show neuronal soma (upper panels) and dendrites (lower panels; MAP2; ; GluA1, ; DAPI; ). Scale bars = 10 μm.

Article Snippet: For the detection of proteins in situ primary antibodies were used targeting Arc (rabbit polyclonal, 1:1000, synaptic systems, #156003), Biotin (mouse monoclonal, 1:1000, Sigma, #B7653), MAP2 (guinea-pig polyclonal, 1:1000, synaptic systems, #188004; monoclonal, 1:1000, Abcam, #AB11267), Synapsin-1 (mouse monoclonal, 1:1000, synaptic systems, #106011), GluA1 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB31232), GluA1 (N-terminal; mouse monoclonal, 1:500, #MAB2263), GluA2 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB1768-I), GluA2 (N-terminal; mouse polyclonal, 1:500, Abcam, #133477), and GluA3 (N-terminal; mouse monoclonal, 1:250, Thermofisher, #32-0400).

Techniques:

Journal: Frontiers in Molecular Neuroscience

Article Title: Secreted Amyloid Precursor Protein-Alpha Enhances LTP Through the Synthesis and Trafficking of Ca 2+ -Permeable AMPA Receptors

doi: 10.3389/fnmol.2021.660208

Figure Lengend Snippet: Secreted amyloid precursor protein-alpha promoted Arc expression is affected by Accell TM Arc siRNA. Representative images show Arc protein levels the soma (upper panels) and dendrites (lower panels) in control and sAPPα-treated primary hippocampal neurons ± co-treatment with Arc siRNA or non-targeted (NT) control siRNA. In both the (A) soma and (B) dendrites ( n = 19–30), sAPPα treatment significantly enhanced Arc expression in sAPPα-only and sAPPα + NT siRNA conditions. This effect was inhibited by co-treatment with the Arc siRNA. Outliers were removed from each experiment prior to amalgamation using Grubb’s tests, and normality was detected by D’Agostino and Pearson omnibus normality tests. Significance was calculated using a Kruskal–Wallis one-way ANOVA with Dunn’s multiple comparisons test on raw data, and expressed as fold change relative to the experimental control. All data are expressed relative to control, as mean ± SEM from 3 experiments. Hashes (#) denote significant difference from control, asterisks (*) denote significant difference from sAPPα-treated. # /* p ≤ 0.05, **/ ## p ≤ 0.005. Representative images show neuronal soma, dendrites (MAP2, ), Arc protein ( ), nuclei (DAPI, ). Scale bars = 10 μm.

Article Snippet: For the detection of proteins in situ primary antibodies were used targeting Arc (rabbit polyclonal, 1:1000, synaptic systems, #156003), Biotin (mouse monoclonal, 1:1000, Sigma, #B7653), MAP2 (guinea-pig polyclonal, 1:1000, synaptic systems, #188004; monoclonal, 1:1000, Abcam, #AB11267), Synapsin-1 (mouse monoclonal, 1:1000, synaptic systems, #106011), GluA1 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB31232), GluA1 (N-terminal; mouse monoclonal, 1:500, #MAB2263), GluA2 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB1768-I), GluA2 (N-terminal; mouse polyclonal, 1:500, Abcam, #133477), and GluA3 (N-terminal; mouse monoclonal, 1:250, Thermofisher, #32-0400).

Techniques: Expressing

Journal: Frontiers in Molecular Neuroscience

Article Title: Secreted Amyloid Precursor Protein-Alpha Enhances LTP Through the Synthesis and Trafficking of Ca 2+ -Permeable AMPA Receptors

doi: 10.3389/fnmol.2021.660208

Figure Lengend Snippet: de novo GluA1 persist at the cell surface following siRNA-mediated knockdown of Arc protein. Representative images of somatic (upper panels) and dendritic (lower panels) of control and sAPPα-treated conditions ± treatment with the Arc siRNA. (A) Average data showing treatment with sAPPα (1 nM, 2 h) and Arc siRNA (1 μM, pre-treatment: 60 min, co-treatment: 2 h) promotes an increase in (A) somatic ( n = 28–35 cells) and (B) dendritic ( n = 79–132) de novo cell-surface GluA1. Outliers were removed from each experiment prior to amalgamation using Grubb’s tests, and normality was detected by D’Agostino and Pearson omnibus normality tests. All data are expressed relative to control, as mean ± SEM from 3 experiments. Data assessing the effect of siRNA are expressed as mean ratio of sAPPα + Arc siRNA/Arc siRNA alone ± SEM. Significance between control and treatment was assessed by student’s t -test, significance between sAPPα-treated and sAPPα + siRNA-treated was assessed by Mann-Whitney two-tailed U-test, ** p ≤ 0.001, **** p ≤ 0.0001. Representative images show neuronal soma (upper panels) and dendrites (lower panels; MAP2, ; GluA1, ; DAPI, ). Scale bars = 10 μm.

Article Snippet: For the detection of proteins in situ primary antibodies were used targeting Arc (rabbit polyclonal, 1:1000, synaptic systems, #156003), Biotin (mouse monoclonal, 1:1000, Sigma, #B7653), MAP2 (guinea-pig polyclonal, 1:1000, synaptic systems, #188004; monoclonal, 1:1000, Abcam, #AB11267), Synapsin-1 (mouse monoclonal, 1:1000, synaptic systems, #106011), GluA1 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB31232), GluA1 (N-terminal; mouse monoclonal, 1:500, #MAB2263), GluA2 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB1768-I), GluA2 (N-terminal; mouse polyclonal, 1:500, Abcam, #133477), and GluA3 (N-terminal; mouse monoclonal, 1:250, Thermofisher, #32-0400).

Techniques: MANN-WHITNEY, Two Tailed Test

Journal: Frontiers in Molecular Neuroscience

Article Title: Secreted Amyloid Precursor Protein-Alpha Enhances LTP Through the Synthesis and Trafficking of Ca 2+ -Permeable AMPA Receptors

doi: 10.3389/fnmol.2021.660208

Figure Lengend Snippet: Secreted amyloid precursor protein-alpha enhances cell-surface GluA1/2-containing AMPARs. Representative images showing cell-surface GluA1/2 levels in the soma (top panels) and dendrites (lower panels) from 30 min (left) and 2 h (right) controls and sAPPα treated conditions. Average data showing 1 nM sAPPα promotes an increase in the (A) soma ( n = 28–31 cells) and (B) dendrites ( n = 98–142) following 2 h- but not 30 min treatments. All data are expressed relative to control, as mean ± SEM from 3 experiments. Normality was detected by D’Agostino and Pearson omnibus normality tests and significance was assessed by student’s t -test, ** p ≤ 0.001, **** p ≤ 0.0001. Representative images show neuronal soma (upper panels) and dendrites (lower panels; MAP2, ; GluA1/2, ; DAPI, ). Scale bars = 10 μm.

Article Snippet: For the detection of proteins in situ primary antibodies were used targeting Arc (rabbit polyclonal, 1:1000, synaptic systems, #156003), Biotin (mouse monoclonal, 1:1000, Sigma, #B7653), MAP2 (guinea-pig polyclonal, 1:1000, synaptic systems, #188004; monoclonal, 1:1000, Abcam, #AB11267), Synapsin-1 (mouse monoclonal, 1:1000, synaptic systems, #106011), GluA1 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB31232), GluA1 (N-terminal; mouse monoclonal, 1:500, #MAB2263), GluA2 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB1768-I), GluA2 (N-terminal; mouse polyclonal, 1:500, Abcam, #133477), and GluA3 (N-terminal; mouse monoclonal, 1:250, Thermofisher, #32-0400).

Techniques:

Journal: Frontiers in Molecular Neuroscience

Article Title: Secreted Amyloid Precursor Protein-Alpha Enhances LTP Through the Synthesis and Trafficking of Ca 2+ -Permeable AMPA Receptors

doi: 10.3389/fnmol.2021.660208

Figure Lengend Snippet: Secreted amyloid precursor protein-alpha downregulates cell-surface GluA2/3-containing AMPAR expression. Representative images showing cell-surface GluA2/3 levels in the soma (top panels) and dendrites (lower panels) from 30 min (left) and 2 h (right) controls and sAPPα treated conditions. (A) Average data showing sAPPα (1 nM) does not affect somatic GluA2/3 following 30- and 2 h treatments ( n = 30–32 cells). (B) Average data showing sAPPα (1 nM; 30 min, 2 h) promotes a decrease in the dendrites ( n = 109–111). All data are expressed relative to control, as mean ± SEM from 3 experiments. Normality was detected by D’Agostino and Pearson omnibus normality tests and significance was assessed by student’s t -test, ** p = 0.0033, *** p = 0.0002. Representative images show neuronal soma (upper panels) and dendrites (lower panels; MAP2, , GluA2/3, ; DAPI, ). Scale bars = 10 μm.

Article Snippet: For the detection of proteins in situ primary antibodies were used targeting Arc (rabbit polyclonal, 1:1000, synaptic systems, #156003), Biotin (mouse monoclonal, 1:1000, Sigma, #B7653), MAP2 (guinea-pig polyclonal, 1:1000, synaptic systems, #188004; monoclonal, 1:1000, Abcam, #AB11267), Synapsin-1 (mouse monoclonal, 1:1000, synaptic systems, #106011), GluA1 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB31232), GluA1 (N-terminal; mouse monoclonal, 1:500, #MAB2263), GluA2 (C-terminal; rabbit polyclonal, 1:500, Abcam, #AB1768-I), GluA2 (N-terminal; mouse polyclonal, 1:500, Abcam, #133477), and GluA3 (N-terminal; mouse monoclonal, 1:250, Thermofisher, #32-0400).

Techniques: Expressing