Nissl-stained histological sections revealed that cKO embryos displayed a dysgenesis of the olfactory bulbs (data not shown) and SE, including an almost complete absence of the midline (15% penetrance, n = 25) that resulted in a mild rostral holoprosencephaly (Figure 1e, e’, e”; Additional data file 2). (a’ -f’) cKO, (a” -f”) cKO no midline (most severe phenotype observed). At E12.5, conditional Sp8 mutant tissue sections revealed a significant size reduction of the telencephalon and the morphological absence of the septum at rostral levels (a’, a”). Caudally, the basal ganglia appear as a single eminence (b’, b”) when compared to controls (b). At E15.5, cKO forebrains lacked the septum (c, c’). Due to an almost complete disgenesis of the midline, the lateral ventricles appear as a single aqueduct space (compare (c) with the area indicated by the asterisk in (c”)). At more caudal levels, there is p32 Inhibitor M36 no obvious difference between Sp8cKO and Sp8cKO no midline. However, when compared to wild-type embryos, the internal capsule of conditional mutants showed aberrant fiber bundles extending towards the basal telencephalon (arrows in (d, d’, d”)). At E18.5, further reduction of the forebrain size and a variable dysgenesis of the midline were observed (e, e’, e”). em Sp8 /em -deficient embryos typically lack a discernable corpus callosum (CC); instead, unilaterally probst bundles (PB) were apparent (e, e’, e”). 1749-8104-2-8-S2.jpeg (654K) GUID:?69F8D3F8-920D-4714-8F02-C5519D61C1EB Additional data file 3 No evidence for cell cycle alterations in Sp8 mutants. Immunohistochemistry for (a, a’, b, b’) Tuj and (c, c’) Tuj/Pax6 antibodies on coronal sections. Tuj staining revealed no difference in the thickness of the PP/CP at E10.5 (a, a’), E12.5 (b, b’), and E15.5 (c, c’). Therefore, premature differentiation is not evident in cKO. Estimation of cell cycle parameters, using (e, e’, f) BrdU, (e, e’, g) phosphor-histone H3, and (d, d’, h) BrdU/IdU staining on E12.5 sections. None of the tested cell cycle parameters were significantly abnormal in Sp8 conditional mutants. 1749-8104-2-8-S3.jpeg (395K) GUID:?9C6A7FB3-B9CD-4B27-973C-2C3214386D12 Abstract Background The forebrain consists of multiple structures necessary to achieve elaborate functions. Proper patterning is, therefore, a prerequisite for the generation of optimal functional areas. Only a few factors have been shown to control the genetic networks that establish early forebrain patterning. Results p32 Inhibitor M36 and conclusion Using conditional inactivation, we show that the transcription factor Sp8 Cryaa has an essential role in the molecular and functional patterning of the developing telencephalon along the anteroposterior axis by modulating the expression gradients of em Emx2 /em and em Pax6 /em . Moreover, Sp8 is essential for the maintenance of ventral cell identity in the septum and medial ganglionic eminence (MGE). This is probably mediated through a positive regulatory interaction with Fgf8 in the medial wall, and Nkx2.1 in the rostral MGE anlage, and independent of SHH and WNT signaling. Furthermore, em Sp8 /em is required during corticogenesis to sustain a normal progenitor pool, and to control preplate p32 Inhibitor M36 splitting, as well as the specification of cellular diversity within distinct cortical layers. Background The mammalian forebrain, with its components the basal ganglia (subpallium) and cortex (pallium), is a result of advanced evolutionary processes. Although several genetic pathways that establish cell diversity within the developing telencephalon have been identified, only a few factors have been shown to control the earliest steps of anteroposterior (A/P) and dorsoventral (D/V) patterning . From embryonic stage E7.5 onwards, the telencephalic vesicles are progressively regionalized through complex interactions of secreted ligands from inductive centers, and by the regionalized or graded expression of transcription factors [1-3]. FGF (Fibroblast Growth Factor) signaling acts downstream of SHH (Sonic Hedgehog) and is required to both specify and promote the proliferation and/or survival of ventral cell types in p32 Inhibitor M36 the telencephalon [4-7], while WNT (Wint) signaling apparently elaborates archicortical morphogenesis [8,9]. Interestingly, modulation of the normal expression gradient of em Fgf8 /em in the early cortical primordium alters the molecular location and the size of cortical domains along the A/P axis [10,11]. At the beginning of cortical neurogenesis, several transcription factors display graded expression in cortical progenitors along the main axes, which seems to confer cortical regional specificity [1-3]. The transcription factors Emx2 and Pax6 exhibit an opposing expression gradient along the A/P axis of the forebrain. Accordingly, single mutants of either em Emx2 /em or em Pax6 /em show a severe shrinkage of the corresponding cortical area, which normally expresses these genes at high levels . Of note, Emx2 is p32 Inhibitor M36 the only factor that has been shown to additionally affect the innervation of thalamic axons into the cortex [1-3]. The zinc-finger transcription factor Sp8 is expressed in the developing nervous system, limbs and the tail bud. Analysis of em Sp8 /em knockout mice revealed severe truncations of the limbs and.