rat anti-cfos primary antibody synaptic systems 226 017  (Synaptic Systems)

Journal: bioRxiv

Article Title: Mapping memories: pulse-chase labeling reveals AMPA receptor dynamics during memory formation

doi: 10.1101/2023.05.26.541296

Figure Lengend Snippet: (A) Expression of HT-GluA1 in pyramidal neurons in mouse hippocampus CA1. (Top left) Coronal section showing overall expression pattern. Arrowhead indicates an expressing cell. (Bottom left) Single neuron expressing HT-GluA1. (Right) Dendrites co-expressing membrane-localized GFP (cyan) and HT-GluA1 stained with AF 647 -HTL, magenta. Scale bars, 200 μm (top left and bottom left), and 5 μm (right). (B) EPSILON scheme for tagging spines potentiated during CFC. IHC, immunohistochemistry. (C) Pyramidal neuron in CA1 after EPSILON tagging, CFC, and cFos immunostaining. Preexisting HT-GluA1 stained with AF 647 (green) and immunostained cFos (cyan) are shown here. Scale bar, 200 μm. (D) Inset from C showing spines labeled with both dyes. (Left) Preexisting HT-GluA1 stained with AF 647 -HTL. (Middle) HT-GluA1 newly exposed during CFC, stained with JF 549i -HTL. (Right) Merge. Scale bars, 10 μm. (E)-(G) Density plots of spine fluorescence in (E) home cage control, (F) context-only, and (G) CFC animals (home cage control: n = 43,659 spines from 3 mice; context-only: n = 74,801 spines from 4 mice; CFC: n = 75,200 spines from 4 mice). Thresholds indicated with dashed lines (see for calculation of threshold). (H) Map of all spines (grey) and spines above Dye 2 threshold (color). The color scale indicates the fluorescence ratio, Dye 2/Dye 1. Scale bar, 200 μm. (I) Percentage of spines with Dye 2 above threshold (home cage control: n = 12 cells from 3 mice; context-only: n = 17 cells from 4 mice; CFC: n = 19 cells from 4 mice). Distinct mice indicated by different shape symbols. Error bars show mean ± s.e.m., twosided Wilcoxon rank-sum test. (J) Dye 2 intensity above threshold in potentiated spines (home cage control: n = 215 spines from 12 cells from 3 mice; context-only: n = 683 spines from 16 cells from 4 mice; CFC: n = 1,016 spines from 19 cells from 4 mice). Distinct mice indicated with marker shape and distinct cells by marker color. Lower and upper bounds of the box plot: 25 th and 75 th percentiles; lower and upper whiskers: minimum and maximum; and center lines: median. Two-sided Wilcoxon rank-sum test. (K) , (L) HT-GluA1 expressing CA1 neurons (green) with (K) high and (L) low cFos expression (cyan). Scale bars, 10 μm. (M) Distributions of cFos intensities in individual cells. Green: no exposure ( n = 2,285 cells from 3 mice); cyan: home cage control ( n = 1,922 cells from 3 mice); magenta: contextonly ( n = 5,074 cells from 4 mice); blue: CFC ( n = 7,208 cells from 4 mice). (N), (O) Correlation between cFos intensity and the fraction of spines with high HT-GluA1 exocytosis from (N) home cage control ( n = 12 cells from 3 mice) and context-only ( n = 17 cells from 4 mice) and (O) CFC ( n = 19 cells from 4 mice). Distinct mice indicated by different shape symbols. R , Pearson’s linear correlation coefficient, P value from two-sided Student’s t-test.

Article Snippet: For cFos immunostaining, the slices were incubated with a rat anti-cFos primary antibody (1:1000 dilution in 1% BSA in PBST, Synaptic Systems, 226 017) for 24 hours at room temperature on a shaker.

Techniques: Expressing, Membrane, Staining, Immunohistochemistry, Immunostaining, Labeling, Fluorescence, Control, Marker