Fig. 4
Morphology of septal ChAT-positive neurons during aging and the impact of chronic neuroinflammation. A–F Representative confocal images of reconstructed medial septum (MS) cholinergic cells from control and GFAP-IL6 cohorts. The extended depth of focus images (A–E) obtained from collapsing 3D tile scanned confocal microscopy images of ChAT + cholinergic cells obtained under 63 × oil immersion objective, along with the corresponding manually reconstructed images (a-e) in Neurolucida 360, of young, adult and old mice from control and GFAP-IL6 cohorts (scale bar 20 μm). G–L Grouped bar graphs representing the effects of aging and neuroinflammation on ChAT+ cholinergic cell morphology in the medial septum (MS), measured by branched structure analysis. Quantitative analysis of the morphological changes in ChAT+ cholinergic cells, including G soma area (µm2), H soma perimeter (µm), I soma circularity, J number of processes from soma (n), K total length of processes (µm) and L number of nodes (n), of young (n = 25), adult (n = 25) and old (n = 25) mice from control (black) and GFAP-IL6 (green) cohorts. Data presented as mean ± SEM and analysed with two-way ANOVA followed by Tukey’s post hoc tests. Post hoc effects of ‘genotype’ are represented in asterisks (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001); post hoc effects of ‘age’ are represented by hash symbols (#p < 0.05, ##p < 0.01, ###p < 0.001, ####p < 0.0001). M–O Grouped bar graphs representing the effects of aging and neuroinflammation on ChAT+ cholinergic cell spatial distribution in the medial septum (MS), measured by convex-hull analysis. Quantitative analysis of spatial distribution changes in ChAT+ cholinergic cells, including M convex 3D surface area (µm2), N convex 3D volume (µm3) and O convex perimeter (µm) of young (n = 25), adult (n = 25) and old (n = 25) mice from control (black) and GFAP-IL6 (green) cohorts. Data presented as mean ± SEM and analysed with two-way ANOVA followed by Tukey’s post hoc tests. Post hoc effects of ‘genotype’ are represented in asterisks (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001); post hoc effects of ‘age’ are represented by hash symbols (#p < 0.05, ##p < 0.01, ###p < 0.001, ####p < 0.0001)