hapln 771  (R&D Systems)

Journal: Nature Communications

Article Title: Midbrain extracellular matrix and microglia are associated with cognition in aging mice

doi: 10.1038/s41467-025-66434-z

Figure Lengend Snippet: a Left: photomicrographs of immunohistochemically labeled hyaluronan and proteoglycan link protein 1 (HAPLN1), aggrecan, and tyrosine hydroxylase (TH) from the ventral tegmental area (VTA) of a late-middle-aged mouse. Numbered images in middle panel correspond to numbered fields of view within white squares in the left panels. Right panels depict the fields of view delineated by white squares in middle panel. Right: photomicrographs depicting HAPLN1 and aggrecan deposition on dopamine neuron surfaces, and the peri-dopamine neuron region of interest used to estimate protein abundances at neuronal membranes. Experiments were performed once on a single cohort of mice. b Boxplots depicting field of view coverage of HAPLN1 and aggrecan in the VTA of young-adult (black; n = 8 mice), aging-unimpaired (AU; green; n = 6 mice), and aging-impaired (AI; yellow; n = 4 mice) mice (n-Way ANOVA; post-hoc Tukey-Kramer; HAPLN: p = 0.022; Aggrecan: p = 0.25). c Boxplots depicting HAPLN1-DA neuron, aggrecan-DA neuron, and HAPLN1-aggrecan-DA neuron puncta densities in young-adult ( n = 8 mice), aging-unimpaired (AU; n = 6 mice), and aging-impaired (AI; n = 4 mice) mice (n-Way ANOVA; two-sided; HAPLN1-DA: p = 0.029; Aggrecan-DA: p = 0.62; HAPLN-Aggrecan-DA: p = 0.031). d Schematic of the analysis strategy used to examine hyaluronan and Homer2 on dopamine neurons (Homer-DA neuron). Experiments were performed once on a single cohort of mice. e Boxplots depicting homer-DA neuron puncta densities in young-adult (black) and late-middle-aged (gray) sedentary (squares) and behavior-trained (triangles) mice (n-way ANOVA; two-sided; Behavior: p = 0.65; n = 8 young-sedentary; 12 young-behavior; 8 middle-aged-sedentary; 12 middle-aged-behavior mice). f Relationship between homer-DA neuron puncta densities and performance on foraging task (robust regression; two-sided; Young: p = 0.91; Middle-aged: p = 0.039). g Scatter plots of hyaluronan-DA neuron puncta densities plotted against Homer2-DA neuron puncta densities in sedentary and behavior-trained mice (robust regression; two-sided; Young Sedentary: p = 0.34; Middle-aged Sedentary: p = 0.14; Young Behavior: p = 0.0036; Middle-aged Behavior: p = 0.0068). In all boxplots, boxes represent interquartile ranges (IQR; 25–75 percentiles), middle lines represent medians, and whiskers extend +/− 1.5*IQR. Source data are provided in the file - Fig. 7 and all statistics are provided in Supplementary Data . * represents p < 0.05.

Article Snippet: Sections were washed in 1 M PBS (4 × 10 min) and then incubated with streptavidin-conjugated AlexaFluor-647 in the 1% BSA and 0.1% saponin block solution for 2 h. The tissue underwent another 4 × 10 min wash in 1 M PBS, and then was blocked again in 5% normal donkey serum with no additional permeabilization agent for 1 h and was then incubated overnight with mild agitation in a solution containing different combinations of goat anti-GFP (1:1000; Frontier Institute cat: GFP-go-Af1480), rabbit-anti-IBA1 (1:500; Wako cat: 019-19741), chicken anti-TH (1:500; Aves cat: TYH), rabbit anti-Homer2 (1:2000; Synaptic Systems cat: 160 003), mouse anti-Vglut1 (1:2000; DSBH cat: N28/9), biotinylated WFA (1:100; Vector Labs, B-1355), rabbit anti-aggrecan (1:200; Sigma Aldrich AB1031), goat-anti HAPLN (1:200; R&D Systems cat: AF2608) and the biotinylated HABP lectin in 5% NDS solution.

Techniques: Labeling