Fig. 1: Approach for analyses and quantification of changes in synaptic protein levels in AD and DS. | Nature Communications

Fig. 1: Approach for analyses and quantification of changes in synaptic protein levels in AD and DS.

From: Increased excitatory to inhibitory synaptic ratio in parietal cortex samples from individuals with Alzheimer’s disease

Fig. 1

a Schematic illustrating the methodological approach for processing fresh-frozen parietal cortex subsections from the same tissue block (step 1) for either fluorescence deconvolution tomography (FDT; step 2) or microtransplantation of synaptic membranes (MSM; step 3) analyses. See “Methods” for details. P2, synaptosomal fraction. b Top: Representative pseudocolor image of amyloid plaques in parietal cortex of an Alzheimer’s disease (AD) case (calibration bar, 35 μm). Bottom: Bar graph showing density of amyloid plaques in the three groups (mean ± SEM; quantified from 20 individual 574,200 μm2 images per case). Each dot represents the density of amyloid plaques in each subject (n = 5 controls, 5 AD and 6 DS). CTRL, control; DS, Down syndrome. Controls had no plaques (indicated by #). All AD and DS cases exhibited plaques, but there were no group differences (AD vs DS, P = 0.800, two-sided Student’s t-test). Photomicrographs showing PSD-95 (green) and gephyrin (GPHN, red) immunoreactive puncta in layers 1 (c) and 2 (d) parietal cortex from control (CTRL), AD, and DS subjects. Calibration bars, 2 μm. Synaptic immunolabeling intensity frequency distributions from the FDT analyses show the proportion of PSDs (Y-axis) in layers 1 (e,f) and 2 (g,h) that were immunolabeled for PSD-95 or GPHN at different immunoreactivity (IR) intensities (X-axis) for the three groups; data are expressed as percentages of total labeled PSDs and plotted as group mean values ± SEM (n = 5 controls, 5 AD, and 6 DS). Layer 1 results are based on 10 individual 42,840 μm3 3-D builds per subject with the following exceptions: 12 builds (two AD cases), 11 builds (one AD, two DS cases), and 9 builds (one DS case). Layer 2 results are based on 10 individual 3-D builds per subject, except for one AD case with 12 builds. Note the leftward skew in the immunolabeling intensity frequency distributions for the AD and DS groups relative to controls; this was most pronounced for layer 2 (two-way RM-ANOVA, P < 0.0001 for interaction between groups and intensities for both GPHN and PSD-95 in layer 2; P = 0.0066 for GPHN and P = 0.0419 for PSD-95 in layer 1). Source data for this and all figures are found in the source data file.

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