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. 2018 Jan 22;8:1381. doi: 10.1038/s41598-018-19794-0

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© The Author(s) 2018

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Examples of α-NeuN immunostaining in the hippocampus proper and dentate gyrus of Sema3E^0/0 (a–c) and Nestin-cre; PlxnD1^flox/flox (f) mice at different postnatal stages. Note the presence of numerous NeuN-positive cells in the molecular layer in absence of Sema3E/PlexinD1 signalling (arrowheads), especially in Sema3E-deficient mice. Also note the presence of several waves of the suprapyramidal blade of dentate gyrus. (d) Double immunolabeling of Calretinin (green) and Calbindin (red) in the dentate gyrus of Sema3E^0/0 mice at P9. Note the presence of numerous ectopic Calbindin-positive neurons in the IML of Sema3E^0/0 mice (arrowheads). (e) Prox-1 immunolabeling in the dentate gyrus of Sema3E^0/0 mice at P15 shows the presence of numerous Prox-1-positive granule cells in the molecular layer (arrowheads). (g,i) Double immunolabeling of Calretinin (green) and Calbindin (red) in the dentate gyrus of control (g) and Sema3E^0/0 adult mice (i). Note the presence of numerous ectopic Calbindin-positive neurons in the IML of Sema3E^0/0 mice (arrows in i). (h–j) Examples of BrdU-labeled neurons in the granule cell layer of control (h) and Sema3E^0/0 (j) mice. Numerous BrdU-positive cells forming columns in the granule cell layer can be seen in mutant mice (arrows in j). (k) Histogram illustrating the number of NeuN-positive cells in the IML of the suprapyramidal blade of the dentate gyrus in Sema3E^+/0 and Sema3E^0/0 mice. (l–m) Histograms illustrating the number of BrdU-positive cells counted in the whole granule cell layer (l) and in the outer portion of the granule cell layer of the dentate gyrus (labeled as ‘o’) (m) in Sema3E^+/+, Sema3E^+/0 and Sema3E^0/0 mice. Asterisks indicate statistical differences between groups. *P ≤ 0.05; **P ≤ 0.01; ***P ≤ 0.001; ****P ≤ 0.0001. ANOVA Bonferroni post hoc test. Abbreviations as in Figs 1–3 and i = inner portion of the granule cell layer; IML = inner molecular layer; o = outer portion of the granule cell layer and OML = outer molecular layer. Scale bars: a = b = 500μm; c = f = 500 μm; d = 200 μm; e = 200 μm; g and h = 150 μm pertains to i and j respectively.