Journal: Journal of neuroinflammation
Article Title: β-arrestin1 regulates astrocytic reactivity via Drp1-dependent mitochondrial fission: implications in postoperative delirium.
doi: 10.1186/s12974-023-02794-x
Figure Lengend Snippet: Fig. 3 β-arrestin1 in astrocytes modulates cognitive impairments and astrocytic reactivity in the mouse model for POD. A β-arrestin1 protein levels in brain lysate of WT and β-arrestin1−/− mice. B Experimental protocol and timeline of the POD mouse model. C Representative moving track plots (red curve) of mice in the second trial of Y-maze test. Blue box represents the novel arm. D Time (%) spent in the novel arm in the Y-maze test. E Bouts of novel arm entry in the Y-maze test. F Representative moving track plots (red curve) of mice in the probe trial of Morris water maze test. Black circle represents the invisible platform. G Latency (s) to reach the hidden platform in the probe test of Morris water maze test. H Crossing times in target quadrant in the probe test of Morris water maze test. I Representative immunofluorescent staining of GFAP in the hippocampus. J Analysis of GFAP-positive cell body area in the hippocampus. K Analysis of GFAP-positive cell numbers in the hippocampus. L Heatmap of the expression level of the neurotoxic astrocytes-specific transcripts in the hippocampus. M Expression of C3, Serping1 and Psmb8 in the hippocampus. N Densitometric analysis of C3, Serping1 and Psmb8. O Schematic diagram of the mice model with micro-injection of AAV-siβ-arrestin1 into the hippocampus. P Immunofluorescent co-localization of GFP and GFAP (red) after the AAV micro-injection. Q Representative immunofluorescent staining of GFAP (red) in the hippocampus. R Analysis of astrocytic reactivity by GFAP-positive cell body area and GFAP-positive cell numbers in the hippocampus. S Bouts of novel arm entry in the Y-maze test. T Time (%) spent in the novel arm in the Y-maze test. U Latency (s) to reach the hidden platform in the probe test of Morris water maze test. V Crossing times in target quadrant in the probe test of Morris water maze test. For C-M except for R, data were analyzed by two-way ANOVA followed by Tukey’s multiple comparisons test. *P < 0.05, **P < 0.01 and ***P < 0.001 vs. the WT-CON group or NC AAV CON group. #P < 0.05, ##P < 0.01 and ###P < 0.001 vs. the WT-POD mice or NC AAV POD. For R, data are analyzed by unpaired Student’s t-test. *P < 0.05 and ***P < 0.001 vs. the NC AAV POD group. n = 6 mice per group for immunofluorescent staining. n = 3 for western blotting. n = 7–10 mice for behavioral tests. Values are presented as means ± SEM
Article Snippet: After blocking with 10% nonfat dry milk in Tris-buffered saline (20 mM Tris– HCl, 500 mM NaCl, pH 7.4) with Tween 20 (Aladdin, #T104863), the membranes were then probed with the following primary antibodies overnight at 4 °C: mouse anti-GFAP antibody (1:1000, Cell Signaling Technology, #3670), rabbit anti-Iba-1 antibody (1:1000, Wako, #019-19741), rabbit anti-β-arrestin1 (1:1000, Cell Signaling Technologies, #12697), rabbit anti-βarrestin2 (1:200, Cell Signaling Technologies, #3857), rabbit anti-C3 antibody (1:1000, abcam, #ab11887), rabbit anti-Serping1 antibody (1:1000, proteintech, #12259-1-AP), rabbit anti-Psmb8 antibody (1:1000, proteintech, #14859-1-AP), mouse anti-Fis1 antibody (1:1000, Santa Cruz, #sc-376447), mouse anti-Drp1 antibody (1:1000, Santa Cruz, #sc-101270), rabbit anti-Drp1 antibody (1:1000, proteintech, #12957-1- AP), rabbit anti-COX IV antibody (1:1000, Cell Signaling Technology, #4850), mouse anti-β-actin antibody (1:3000, sigma, #a1978).
Techniques: Staining, Expressing, Microinjection, Western Blot
