a, b, Representative immunoblot detecting TIMP2 from equal volumes of 1-month-old (n = 8), 3-month-old (n = 7), and 20-month-old (n = 8) WT mouse plasma with corresponding Ponceau S stain (a) and quantification (b). c, d, Representative immunoblot detecting TIMP2 and neuron-specific enolase loading control from hippocampal lysates (80 μg) from 1-, 12- and 20-month-old WT mice (c) with corresponding quantification (d; n = 7 per group). e, Representative confocal microscopy images from WT or TIMP2 KO mice demonstrating specificity of TIMP2 signal (green) in dentate gyrus (n = 4 mice per group; 3-month-old male mice; white arrowheads indicate TIMP2+ cells; scale bar, 100 μm). f, Representative confocal microscopy images showing TIMP2 (green), Iba1 (blue), and NeuN (red) staining in dentate gyrus/hilus of WT mice at various ages (C57Bl/6; National Institute on Aging colony; 1-month-old (n = 8), 2-month-old (n = 8), 6-month-old (n = 6), 12-month-old (n = 8), and 20-month-old mice (n = 9); scale bar, 100 μm). g, h, Quantification of TIMP2+Iba1+ cells (g) or TIMP2+ cells lacking Iba1 or NeuN staining (h) in the dentate gyrus/hilus of mice from f. i, Mean signal intensity per TIMP2+ cell for all counted cells within dentate gyrus/hilus in mice from f. j, NeuN+ cell counts per dentate gyrus/hilar area in the indicated ages. k, l, c-Fos-positive cell counts within dentate gyrus from WT mice treated once (k) or four times (l) with different rTIMP2 doses (n = 8 per group; 21-month-old). m, 64Cu-labelled BSA and 64Cu-labelled TIMP2 detected in blood of 21-month-old WT mice euthanized at the indicated time points following an injected dose of 64Cu-labelled BSA (n = 4 mice per time point (n = 3 mice at 24 h)) or 64Cu-labelled TIMP2 (n = 5 mice per time point). n, First-order elimination kinetics of 64Cu-labelled TIMP2 levels were analysed to approximate its blood half-life (curved line indicates confidence interval). o, Ex vivo autoradiography assessment of 64Cu-labelled TIMP2 localization in coronal or sagittal (right-most) brain sections from injected mice (top) with corresponding Nissl staining (middle) and 64Cu-labelled TIMP2/Nissl overlay to examine anatomical co-registration with radioactive signal (colour bar indicates radioactive signal from low (black) to high (white); n = 3 per group; ~20-month-old WT mice). p, Representative analytical high-performance liquid chromatography radioactivity chromatogram from mouse brain homogenates to assess stability of 64Cu-labelled TIMP2 24 h after injection. Dotted line corresponds to retention time of 11.5 min (n = 2 per group; ~20-month-old WT mice). q, Ultraviolet absorbance at 280 nm measured for TIMP2–DOTA before injection in vivo, exhibiting identical retention time as in m. r, Number of c-Fos+ cells in dentate gyrus from aged (21-month-old) WT mice treated seven times systemically every other day with vehicle (n = 8) or 50 μg kg−1 (intraperitoneal) recombinant TIMP2 (n = 7). s, t, Freezing levels for vehicle- or TIMP2-treated aged WT mice during baseline period (s) and during cued fear-conditioning task (t; n = 15 per group; eight intraperitoneal injections; 20-month-old). See Supplementary Fig. 1 for uncropped blot images. One-way ANOVA with Tukey’s post hoc test; two-way ANOVA with Bonferroni’s post hoc test for group × time comparisons; Student’s t-test for two-group comparisons. *P < 0.05, **P < 0.01, ****P < 0.0001; NS, not significant; mean ± s.e.m. Source data