Choline supplementation reduced ΔFosB expression in dorsal GCs of Tg2576 mice.
A. Representative images of ΔFosB staining in GCL..
1. A section from a low choline-treated Tg2576 mouse shows robust ΔFosB-ir in the GCL. The area outlined by the box is expanded below. Red arrows point to Δ FfosB-labeled cells. GCL=granule cell layer. Calibration for the top row, 100 µm; for the bottom row, 50 µm.2-3. Sections from intermediate (2) and high choline (3)-treated Tg2576 mice. Same calibrations as 1.
4, A section from a WT mouse treated with the intermediate diet. Same calibrations as 1.
B. Quantification methods. Representative images demonstrating the thresholding criteria established to quantify ΔFosB.
1. A ΔFosB -stained section shows strongly-stained cells (white arrows).
2. A strict thresholding criteria was used to make only the darkest stained cells red.
C. Use of the strict threshold to quantify ΔFosB-ir.
1. Anterior DG. Tg2576 mice treated with the choline supplemented diet had significantly less ΔFosB-ir compared to the Tg2576 mice fed the low or intermediate diets. Tg2576 mice fed the high choline diet were not significantly different from WT mice, suggesting a rescue of ΔFosB-ir.
2. There were no significant differences in ΔFosB-ir in posterior sections.
D. Methods are shown using a threshold that was less strict.
1. Some of the stained cells that were included are not as dark as those used for the strict threshold (white arrows).
2. All cells above the more permissive threshold are shown in red.
E. Use of the less strict threshold to quantify ΔFosB-ir.
1. Anterior DG. Tg2576 mice that were fed the high choline diet had less ΔFosB-ir pixels than the mice that were fed the other diets. There were no differences from WT mice, suggesting restoration of ΔFosB-ir by choline enrichment in early life.
2. Posterior DG. There were no significant differences between Tg2576 mice fed the 3 diets or WT mice.