Thioflavin s

Tissue sections (40 μm thick) were stained with thioflavin-S to visualize and quantify fibrillar amyloid plaques (Christensen and Pike, 2020 (link)) in contrast to the soluble Aβ₄₂ described above. Briefly, sections containing the PFC (bregma: 2.10 to 1.94 mm) and hippocampus (bregma: −1.22 to −2.54 mm) were washed with 1xPBS, stained for 5 min in 1% thioflavin-S (Sigma, Cat# T1892) aqueous solution, and differentiated for 3 min in 70% alcohol (Franklin and Paxinos, 2008 ; Guntern et al., 1992 (link)). Then, sections were mounted onto slides coated with 2% pig skin gelatin (Sigma, Cat# G1890), coverslipped with Fluorescence Mounting Medium (Abcam, Cat# ab104135), and stored overnight at 4°C. We used a Nikon Eclipse-Ni-E fluorescent microscope to image thioflavin-S-stained plaques in 4 sections of the PFC and dorsal hippocampus per mouse at 10x magnification for quantification. In the PFC, we counted plaques from the rhinal sulcus to the anterior cortex. In the hippocampus, we focused our plaque quantification in the CA1 subregion (from the end of the subiculum to the start of CA2 subregion) given its functional connectivity with the PFC (Li et al., 2015 (link)). Using ImageJ Software (v1.53), we determined the mean number and size of Aβ plaques in all PFC and CA1 sections per mouse according to published methods (Locci et al., 2021 (link)).