Fig. 2: PSD-95 turnover depends on experience.

a, Experimental design for turnover measurement in a PSD-95–HT knock-in mouse. b–e, Example coronal sections showing the pulse (b), chase (c) and calculated lifetime aligned to the Allen CCFv3 (d and e). Note the lifetime gradient that separates the CA1 stratum radiatum (sr) and stratum lacunosum moleculare (slm; long lifetime) from dentate gyrus (DG) and hilus (all shorter lifetimes). f, Average ± s.e. lifetimes of control animals (n = 4). Cortical layers and subfields of the hippocampus (HC) were significantly different (mixed-effects linear model with layers as fixed effects (means: 10.0–16.5 days, s.e.: ~0.5, all P < 0.0001) and animal ID as a random effect (mean, 1.42 days; residual error, 1.39 days)). All mixed-effects models are double sided and without multiple-comparison corrections. g, Average ± s.e. lifetimes for 12 large brain regions were also significantly different in control mice (n = 4; mixed-effects linear model with brain regions as fixed effects (means: 11.9–15.2 days, s.e.: ~0.23, all P < 0.0001); animal ID as a random effect (mean: 0.46 days); residual error: 1.8 days). h, Average ± s.e. lifetimes of four mice under EE and four mice under control conditions. EE increased protein turnover and shortened the average lifetime of PSD-95–HT. Individual animals in gray (EE: 11.8 ± 0.2 days, n = 4; control: 14.2 ± 0.7 days, n = 4; two-sided Wilcoxon rank-sum test W = 26, P = 0.0286). i, Percentage change ± s.e. in control versus EE animals for 12 different brain regions; Mixed-effects linear model for each brain region with group assignment as fixed effects and animal as a random effect. j, Same as i for cortical layers and HC subfields. k, Example images using Airyscan imaging of ExM tissue (maximum projection of five z-planes, 0.3 μm apart) from layer 1, HC CA1 subfield and basal dendrites of CA3 showing both pulse, chase and lifetime (τ). l, Quantification of segmented single-synapse turnover (median lifetime in days; L1, 10.97; L5, 9.5; CA1, 13.78; CA3, 7.32). Boxes show the interquartile range and whiskers mark the 5th and 95th percentiles.