Fig. 1

Novel large clusters of NeuN⁺ puncta/swellings are found in the hippocampus of DPP6-KO mice. A In the CA1 region of the hippocampus of 12-month old DPP6-KO brain sections, immunofluorescence labeling for NeuN (green, 1:500, Millipore) is concentrated not only in the neuronal somas but also in novel structures made of enlarged, variable puncta formed in clusters (arrows). Scale bar = 100 μm, dg, dentate gyrus; so, stratum oriens; sp, stratum pyramidale; sr, stratum radiatum. B A higher magnification image of one of the clusters shown in A. C Even higher magnification of a single cluster from another section. Scale bar = 5 μm. D Immunogold labeling of NeuN (15 nm) in the CA1 stratum radiatum of the hippocampus of a WT 12-month old mouse. Da is a low magnification including two NeuN⁺ swellings. The top swelling is shown in higher magnification in Db, c and the bottom swelling is shown in Dd. Note how the immunogold labeling is concentrated mainly in the two adjacent swellings, which show a fibrous interior and traces of presynaptic vesicles (active zone; arrows) on the periphery, associated with postsynaptic spines (sp). The structure of these vesicles resembles those seen in adjacent separate presynaptic terminals (arrowheads); in Db, the cluster of presynaptic vesicles in the lower left (arrowhead) probably is a presynaptic zone of the swelling also, but the connection is not distinct in the image. In Dd, the NeuN⁺swelling extends off of an axon with presynaptic vesicles that makes a synapse with a spine in the image. (Da, d; the scale bar in d also applies to b) or 500 nm in Dc. Compare to examples from a DPP6-KO mouse shown in the Additional file 1: Fig. S1. And compare the presynaptic active zones in the periphery of the swellings in Figs. 1D, 3, and Additional file 1: Fig. S1. E These novel NeuN-labeled clusters are observed more in 12-month old DPP6-KO mice than in WT. Scale bar = 100 μm. F: Graph comparing the number of novel clusters in the CA1 region of the hippocampus during development from 8-week to 12-month old mice in WT and DPP6-KO mice. From 8-week to 12-month old, labeling in aged DPP6-KO mice is significantly increased compared to WT (n = 8–38 images per age from 3 to 8 mice each WT and DPP6-KO.*** = p < 0.001, **** = p < 0.0001)