Quantitative Immunoblot Analysis of GluA2

Immunoblot analysis was performed as described previously (Gonzalez-Lozano et al., 2021 (link)) on 5 μg of synaptosomal protein from each experimental group. After electrophoresis, gels were scanned using a Gel Doc EZ imager (Bio-Rad) and electro-transferred onto a PVDF membrane overnight at 40 V. After blocking in 5% milk, membranes were incubated with a primary antibody against GluA2 (1:1,000, Cat nr. 75-002, Neuromab) overnight at 4°C and with a matching HRP-conjugated secondary antibody for 2 h at room temperature. The membranes were then scanned with Femto ECL Substrate (Thermo Fisher Scientific, Waltham, MA, USA) on an Odyssey Fc system (LI-COR Bioscience, Lincoln, NA, USA) and quantified using Image Studio software (version 2.0.38). Input differences were corrected using total protein amounts loaded on the gel (Gonzalez-Lozano et al., 2021 (link)).