Journal: bioRxiv
Article Title: C1q and immunoglobulins mediate activity-dependent synapse loss in the adult brain
doi: 10.1101/2024.12.18.629085
Figure Lengend Snippet: (A) Representative Airyscan images of VGLUT1 puncta at contralateral and ipsilateral DGML of hM3Dq+CNO WT mouse, scale bar = 2 μm. (B) Quantification of VGLUT1 puncta density between hemispheres in each mouse group. n = 6 EGFP+Veh mice (contra = 33 ROIs, ipsi = 33 ROIs), n = 6 EGFP+CNO mice (contra = 48 ROIs, ipsi = 48 ROIs), n = 9 hM3Dq+Veh mice (contra = 36 ROIs, ipsi = 36 ROIs) and n = 7 hM3Dq+CNO mice (contra = 48 ROIs, ipsi = 48 ROIs), 1-3 brain sections per hemisphere, 3 ROIs analysed per section. p-values from Bonferroni post-hoc test after 2-way mixed ANOVA with group as between- and hemisphere as within-subject factor (interaction p = 0.019). (C) Representative Airyscan images of VGLUT2 puncta at contralateral and ipsilateral DGML of hM3Dq+CNO WT mouse, scale bar = 2 μm. (D) Quantification of VGLUT2 puncta density between hemispheres in each mouse group. Data are mean ± SEM, n = 5 EGFP+Veh mice (contra = 36 ROIs, ipsi = 36 ROIs), n = 5 EGFP+CNO mice (contra = 42 ROIs, ipsi = 42 ROIs), n = 5 hM3Dq+Veh mice (contra = 36 ROIs, ipsi = 36 ROIs) and n = 7 hM3Dq+CNO mice (contra = 39 ROIs, ipsi = 39 ROIs), 2-4 brain sections per hemisphere, 2-3 ROIs analysed per section. p-values from Bonferroni post-hoc test after 2-way mixed ANOVA with group as between- and hemisphere as within-subject factor (interaction p = 0.015). (E) Representative Airyscan images of Homer1 puncta at contralateral and ipsilateral DGML of hM4Di+CNO J20 mouse, scale bar = 2 μm. (F) Quantification of Homer1 puncta density between hemispheres in hM4Di+CNO J20 mouse. n = 3 EGFP+Veh mice (contra = 17 ROIs, ipsi = 17 ROIs), n = 3 EGFP+CNO mice (contra = 18 ROIs, ipsi = 18 ROIs), n = 3 hM4Di+Veh mice (contra = 15 ROIs, ipsi = 18 ROIs) and n = 6 hM4Di+CNO mice (contra = 40 ROIs, ipsi = 40 ROIs), 2 brain sections per hemisphere, 2-3 ROIs analysed per section. p-values from Bonferroni post-hoc test after 2-way mixed ANOVA with group as between- and hemisphere as within-subject factor (interaction p = 0.014). (G) Representative images of C1q and VGLUT2 colocalization at the contralateral and ipsilateral DGML, colocalized puncta indicated by yellow arrowheads, scale bar = 2 μm. (H) Quantification of percentage total VGLUT2 puncta colocalizing with C1q at contralateral and ipsilateral DGML of hM3Dq+CNO WT mouse. n = 5 mice (contra = 28 ROIs, ipsi = 27 ROIs), 1 brain section per hemisphere, 3 ROIs analysed per section. p-value from paired t test. (I) Schematic of C1qa KO experimental paradigm. Created in https://BioRender.com . (J) Representative widefield images of c-Fos labeling at contralateral and ipsilateral MEC of hM3Dq+CNO C1qa KO mouse, scale bar = 50 μm. (K) Quantification of c-Fos immunoreactive area between hemispheres in hM3Dq+CNO C1qa KO mouse. n = 5 C1qa KO mice (contra = 42 ROIs, ipsi = 42 ROIs), 2-3 brain sections per hemisphere, 3 ROIs analysed per section. p-values from paired t test. (L) Representative Airyscan images of VGLUT2 puncta at contralateral and ipsilateral DGML of hM3Dq+CNO C1qa KO mouse, scale bar = 2 μm. (M) Quantification of VGLUT2 puncta density between hemispheres in hM3Dq+CNO C1qa KO mouse. n = 5 C1qa KO mice (contra = 39 ROIs, ipsi = 39 ROIs), 2-5 brain sections per hemisphere, 3 ROIs analysed per section. p-values from paired t test. Throughout, square points represent males and circular points represent females, linked points indicate data from the same mouse brain, where points are linked 1 point = 1 hemisphere average = average of brain sections. Data shown as mean ± SEM. ns p > 0.05, *p < 0.05, **p < 0.01, ***p < 0.001.
Article Snippet: Pulled long-shaft borosilicate pipettes (3.5” tubes, #3-000-203-G/X, Drummond Scientific) were backfilled with mineral oil (330779, Sigma-Aldrich) before being loaded with either DREADD virus (AAV8- CaMKIIa-HA-hM3Dq-IRES-mCitrine, #50466-AAV8, Addgene; titre = 1.6×10 13 gc/mL or AAV8-CaMKIIa-HA-hM4Di-IRES-mCitrine, #50467-AAV8, Addgene; titre = 1.6×10 13 gc/mL) or control EGFP virus (AAV8-CaMKIIa-EGFP, #50469-AAV8, Addgene; titre = 2.3×10 13 gc/mL).
Techniques: Labeling
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