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. 2016 Jun 27;6:28750. doi: 10.1038/srep28750

Figure 4. Ablation of netrin-G1 in excitatory neurons in the cortex and thalamus.

Figure 4

(a) X-gal staining of coronal sections of a NetrinG1-NLS-LacZ mouse. (b–d) X-gal staining of Rosa-NLS-LacZ reporter mice crossed with Emx1-Cre (b), 5HTT-Cre (c), and Pkcd-Cre (d) lines, respectively. (e–j) Representative images from ISH of netrin-G1 (Ntng1). Horizontal sections from control (e) and Emx1-G1-cKO mice (f) show that cortical expression of netrin-G1 was abolished. Coronal sections from control (g) and 5HTT-G1-cKO (h) mice show the selective reduction of netrin-G1 in the ventral basal nucleus (Vbn) and lateral geniculate nucleus (Lgn). Coronal sections from control (i) and Pkcd-G1-cKO (j) mice show the dramatic reduction of netrin-G1 in almost the entire thalamus. (k–p) Representative images from immunohistochemistry of netrin-G1. In Emx-G1-cKO (l) mice, netrin-G1 disappeared in the Slm and Oml of the hippocampus and piriform cortex layer I compared with the control (k). In 5HTT-G1-cKO (n) mice, netrin-G1 dramatically decreased in cortex layer IV compared with the control (m). In Pkcd-G1-cKO (p) mice, netrin-G1 disappeared from cortical layers IV and I compared with the control (o). Scale bar: 1 mm (a–f); 800 μm (g–j); 750 μm (k–p). (q–s) Behaviors in the EPM. (q) The littermate control mice spent a significantly greater amount of time in the closed arms while Emx1-G1-cKO mice showed no preference between open and closed arms (n = 11/genotype). (r) 5HTT-G1-cKO mice showed no differences from the control mice (n = 12 per genotype). (s) Pkcd-G1-cKO mice showed no differences from the control mice (n = 12 per genotype). Shown are the mean + SEM. Unpaired two-tailed Student’s t test; ***p < 0.001; ns, not significant. (t–v) Percent freezing behavior in the fear conditioning (day 1), context-dependent memory (day 2), and cue-dependent memory (day 3) tests. (t) No significant effect of genotype was detected in Emx-G1-cKO mice in all tests (n = 14 per genotype). (u) No significant effect of genotype was detected in 5HTT-G1-cKO mice in all tests (n = 12 per genotype). (v) No significant effect of genotype was detected in Pkcd-G1-cKO mice in all tests (n = 12 per genotype). Shown are the mean + SEM. Two-way ANOVA was conducted between time and genotype.