To test arealization at the functional level in the double cortex of cKO mice, we examined the primary visual (V1) and somatosensory (S1) cortex by measuring immediate-early gene (IEG) expression triggered by light exposure for 1.5 hr under normal lighting

conditions after 3 days of complete darkness. Immunostaining for the IEG products c-Fos and Egr-1 revealed a strong U0126 concentration signal in visual cortex areas of WT animals, while expression of both genes was significantly lower in both the upper and lower cortex of cKO mice (Figures S5A and S5B). However, light-induced IEG expression was confined to visual cortical areas as no upregulation of c-Fos was detectable

in the primary somatosensory (S1) cortex (Figure S5A). Thus, despite its severe disorganization, the cKO cortex still possesses functional arealization. PF-02341066 clinical trial Lastly, we examined the number of GABAergic neurons in the NC and HC, as their number is crucial for maturation of intracortical neural networks and plasticity (Gandhi et al., 2008, Huang et al., 1999 and Lodato et al., 2011), and their migration into the cerebral cortex may have been affected by these severe alterations. However, GABAergic GAD67-positive neurons were present in normal numbers in the SBH (Figure S5D) and only slightly reduced in the NC of the cKO

mice compared to controls (Figure S5D), suggesting that migration of GABAergic neurons is largely normal, but their number is not entirely sufficient to cover the increased size of the cKO cerebral cortex (1.32×, N = 4, p = 0.0001), intriguingly largely affecting the NC rather than the HC. Taken together, despite the profound alteration of a double cortex formation, neuronal subtype composition appears Mdm2 antagonist relatively normal. In order to understand when and how the prominent SBH is formed in the cKO mice, we first examined when RhoA protein disappears. While RhoA immunoreactivity was present throughout the WT cerebral cortex at embryonic day (E)12, it was largely absent in the cKO mice at this time (Figures 2A and 2B) but not yet at E11 (data not shown). As observed before (Cappello et al., 2006 and Iwasato et al., 2004), Emx1::Cre-mediated recombination occurs specifically in the cerebral cortex, such that RhoA was still present in the neighboring plexus choroideus (Figure 2B, arrowhead) or the ganglionic eminence (GE, data not shown). Moreover, blood vessels depicted by arrows in Figure 2B also maintained RhoA, as Cre is not expressed in these cells.