We hypothesized that Sox14-positive and Dlx2-positive cells are two alternative GABAergic subtypes. To test whether an epistatic relationship exists between the cell types, we investigated the development of the Sox14-positive population in a mouse buy INCB024360 mutant for Dlx2 and for the similarly expressed Dlx1 genes (Dlx1/22KO). This double knockout mouse displays strongly impaired neuronal differentiation in the ventral telencephalon and prethalamus, with incomplete maturation and impaired

migration of GABAergic progenitors ( Anderson et al., 1997; Cobos et al., 2005). At E12.5 in the Dlx1/22KO, Tal1 and Sox14 are ectopically induced in the prethalamus, mirroring their position in the r-Th ( Figures 5A and 5B). Interestingly, the ectopic induction of Tal1 and Sox14 did not correlate with ectopic see more expression of Helt in the cycling progenitor domain of the prethalamus, an observation that helps place Helt function at an earlier stage than Dlx2 ( Figures 5A and 5B). By E16.5 of normal development, most Dlx2-positive GABAergic neurons in the prethalamus have formed the reticular nucleus, while those arising closer to the ZLI form the vLGN. As shown above, the vLGN is invaded at E14.5 by Sox14-expressing GABAergic neurons from the r-Th ( Figures 3B and 3C; Movie S4). This results in the intermixing of

Sox14-positive neurons within the largely Dlx2-expressing vLGN. However, in the double Dlx1/22KO mouse, the entire vLGN is occupied by Sox14-positive neurons ( Figures 5C and 5D). We then asked whether ectopic Sox14-positive neurons in the vLGN acquire their normal GABAergic fate. Indeed the panGABAergic marker Gad1 is highly expressed in these cells despite lack of expression of Dlx1 and Dlx2 ( Figure 5D). To assess whether these ectopic neurons have also acquired a full IGL character, we measured expression of the

IGL marker Npy. Ectopic Sox14 cells in the vLGN express a high level of Npy, resulting in a ∼5-fold increase of Npy-positive cells in the combined IGL/vLGN region compared to control littermates ( Non-specific serine/threonine protein kinase Figures 5E and 5F). We therefore conclude that ectopic Sox14-positive cells are true IGL cells and that Dlx1 and Dlx2 act to suppress IGL fate in the vLGN. Concomitant ectopic induction of Helt is not required for the acquisition of IGL marker expression or GABAergic fate, in agreement with the lack of any detectable phenotype in the rTh of MgntZ/tZ mice. This observation also rules out the possibility that Dlx1 and Dlx2 act as prepatterning genes before the onset of neurogenesis and that in their absence prethalamus is converted into thalamus. The ectopic IGL lineage in the prethalamus overlaps with expression of the Shh-induced gene Nkx2.2 ( Figures 5A and 5D). This suggests that on both sides of the ZLI, Nkx2.2 specifies GABAergic progenitors that differentiate either as IGL or vLGN neurons and that this decision is regulated at least in part by the transcription factors Dlx1 and Dlx2.