Fig. 3. Degenerative cortical neurons in Vps35^Neurod6 brain.

a Representative Nissl stains of control (Vps35^f/f) and Vps35^Neurod6 animals at indicated ages. b Representative images of active caspase-3 immunostaining analysis of neocortex at indicated ages. c Quantification analysis of Nissl stains that revealed a significant reduction in cortical thickness of Vps35^Neurod6 animals at P14/P21, but not at P0/P7, as compared with those of control mice (n = 3–4 animals per genotype; two-tailed unpaired t-test). d Quantification analysis of NeuN⁺ neuron numbers in the neocortex that revealed no significant change at P7, but losses of 12% and 29% of cortical neurons at P14 and P21, respectively (n = 3–4 animals per genotype; two-tailed unpaired t-test). e Quantification analysis of active caspase-3⁺ cells that showed an age-dependent increase in apoptotic cells in Vps35^Neurod6 neocortex at indicated ages (n = 3–4 animals per genotype; two-tailed unpaired t-test). f Images of NeuroSilver-stained brain sections from control and Vps35^Neurod6 mice. Positive signals (dark gray/black staining) were detected in cortical L2–3 neurons and corpus callosum [major white matter (WM) axonal pathways], but not in striatum. Representative images of Jade C staining (g) and anti-Rtn3 immunostaining (h) in cortical L2–3, corpus callosum, and striatum of Control and Vps35^Neurod6 mice. i–k Quantification analysis of data from (f–h). (n = 4 animals per genotype; two-tailed unpaired t-test). Scale bars: in a, b, and f, 100 μm; and in g and h, 20 μm. Individual data points were shown as dots with group mean ± s.e.m; *P < 0.05; ***P < 0.001; n.s. not significant.