Adult B6 and D2 strains were used for protein quantification with three biological replicates of hippocampus. The hippocampal tissues were dissected as previously described50 and lysed in lysis buffer (8 M urea, 0.5% sodium deoxycholate, 50 mM HEPES and pH 8.5). In short, fixed brains were bisected along the midline. Left and right hippocampal regions were dissected under a dissecting microscope by inserting fine blunt forceps into the ventricular cavity just dorsal to the hippocampus and removing overlying cortex and callosum. The surface of the hippocampus and dentate gyrus was used to guide removal of cortex along the septotemporal axis. The exposed hippocampus and dentate gyrus was pulled free of the hemisphere in a ventral-to-dorsal direction. The dorsoanterior aspect of each hippocampus was trimmed free of septum and dorsal fornix, rolled quickly in tissue paper, and immediately weighed to the nearest 0.1 mg. The dissection includes a small part of the subiculum adjacent to CA1 and occasionally a small strand of the fimbria. For each sample, 100 μg of extracted proteins were digested with LysC (1:100, w/w) for 3 h.
