Satb2 is a DNA-binding protein that regulates chromatin organization and gene expression. In the developing brain, Satb2 is expressed in cortical neurons that extend axons across the corpus callosum. To assess the role of Satb2 in neurons, we analyzed mice in which the Satb2 locus was disrupted by insertion of a LacZ gene. In mutant mice, beta-galactosidase-labeled axons are absent from the corpus callosum and instead descend along the corticospinal tract. Satb2 mutant neurons acquire expression of Ctip2, a transcription factor that is necessary and sufficient for the extension of subcortical projections by cortical neurons. Conversely, ectopic expression of Satb2 in neural stem cells markedly decreases Ctip2 expression. Finally, we find that Satb2 binds directly to regulatory regions of Ctip2 and induces changes in chromatin structure. These data suggest that Satb2 functions as a repressor of Ctip2 and regulatory determinant of corticocortical connections in the developing cerebral cortex.

Recently, we and others reported that the doublecortin gene is responsible for X-linked lissencephaly and subcortical laminar heterotopia. Here, we show that Doublecortin is expressed in the brain throughout the period of corticogenesis in migrating and differentiating neurons. Immunohistochemical studies show its localization in the soma and leading processes of tangentially migrating neurons, and a strong axonal labeling is observed in differentiating neurons. In cultured neurons, Doublecortin expression is highest in the distal parts of developing processes. We demonstrate by sedimentation and microscopy studies that Doublecortin is associated with microtubules (MTs) and postulate that it is a novel MAP. Our data suggest that the cortical dysgeneses associated with the loss of Doublecortin function might result from abnormal cytoskeletal dynamics in neuronal cell development.

The use of genetics to study the development of the telencephalon and derivatives such as the cerebral cortex has been limited. The telencephalon begins to form midway through gestation, and targeted mutations in genes suspected of playing roles in its development often lead to early phenotypes that preclude analysis of their role at later stages. This problem can be circumvented using a Cre/loxP recombination system. A mouse line was produced in which cre was targeted to the Foxg1 (BF-1) locus, a gene expressed specifically in the telencephalon and discrete head structures. Crosses between Foxg1-Cre mice and three separate loxP reporter mice generated embryos with recombination patterns matching that expected from the normal pattern of Foxg1 expression. Recombination occurs invariably in the telencephalon, anterior optic vesicle, otic vesicle, facial and head ectoderm, olfactory epithelium, mid-hindbrain junction, and pharyngeal pouches. Recombination in some animals also occurs less efficiently in tissues not known to express Foxg1. We show that the genetic background of the parental mice and the loxP target allele can each contribute to differences in the exact pattern of recombination. Collectively, these data show that Foxg1-Cre mice should be useful in the deletion or ectopic expression of any floxed target gene in a Foxg1-like pattern.

Neurons in the mammalian central nervous system are generated from progenitor cells near the lumen of the neural tube. Time-lapse microscopy of dividing cells in slices of developing cerebral cortex reveals that cleavage orientation predicts the fates of daughter cells. Vertical cleavages produce behaviorally and morphologically identical daughters that resemble precursor cells; these symmetric divisions may serve to expand or maintain the progenitor pool. In contrast, horizontally dividing cells produce basal daughters that behave like young migratory neurons and apical daughters that remain within the proliferative zone. Notch1 immunoreactivity is distributed asymmetrically in mitotic cells, with Notch1 inherited selectively by the basal (neuronal) daughter of horizontal divisions. These results provide cellular and molecular evidence that cortical neurons are generated from asymmetric divisions.

Mouse embryos bearing hypomorphic and conditional null Fgf8 mutations have small and abnormally patterned telencephalons. We provide evidence that the hypoplasia results from decreased Foxg1 expression, reduced cell proliferation and increased cell death. In addition, alterations in the expression of Bmp4, Wnt8b, Nkx2.1 and Shh are associated with abnormal development of dorsal and ventral structures. Furthermore, nonlinear effects of Fgf8 gene dose on the expression of a subset of genes, including Bmp4 and Msx1, correlate with a holoprosencephaly phenotype and with the nonlinear expression of transcription factors that regulate neocortical patterning. These data suggest that Fgf8 functions to coordinate multiple patterning centers, and that modifications in the relative strength of FGF signaling can have profound effects on the relative size and nature of telencephalic subdivisions.KEY WORDS: Fgf8, Forebrain, Patterning, Mouse Development 133, 1831Development 133, -1844Development 133, (2006 The previous studies concentrated on the phenotype of the Fgf8 TelKO and Fgf8 Null/Neo mutant telencephalon beginning at E10.5 and did not examine primary phenotypes in the neural plate or just following neural tube closure. Because prosencephalic expression of Fgf8 begins at neural plate stages (Crossley and Martin, 1995;Shimamura and Rubenstein, 1997; Crossley et al., 2001), it is essential to investigate the mutant phenotypes shortly after this stage to elucidate the mechanisms underlying Fgf8 TelKO and Fgf8 Null/Neo phenotypes. Therefore, here we report studies of Fgf8 dose-dependent effects on neural plate and early post-neurulation stage embryos. Furthermore, Storm et al. (Storm et al., 2003) focused on the effects of reducing Fgf8 dose on telencephalic midline development; here we concentrate on the effect of reducing Fgf8 dose on telencephalic patterning centers, regionalization and growth.We report our finding that specification of the prosencephalon is intact in Fgf8 mutants; however, a major reduction in Foxg1 expression, a reduced mitotic index, and increased apoptosis contribute to telencephalic hypoplasia. We also demonstrate that Fgf8 regulates the expression of Bmp4, Wnt8b and Shh, which in turn affect patterning of both dorsal and ventral structures. Nonlinear effects of Fgf8 dose on Bmp4 expression correlate with a holoprosencephaly phenotype and alterations in the expression of transcription factors that regulate neocortical patterning. The nexus of regulatory interactions between patterning centers that control gradients of transcription factor expression demonstrates that modifications in the relative strength of FGF/BMP/WNT/SHH signaling have profound effects on the relative size and nature of telencephalic subdivisions that are likely to contribute to their phylogenetic and intra-individual diversity. MATERIALS AND METHODS Mice and genotypingAll Fgf8 mutant alleles were maintained on a mixed 129/CD1 Swiss genetic background. Fgf8 Null/+ )embryos. PCR genotyping was performed as described pr...

During the development of the cerebral cortex, progenitor cells produce neurons that migrate to laminar positions appropriate for their birth dates, adopt specific neuronal identities, and form appropriate local and long-distance axonal connections. Here, we report that forebrain embryonic zinc-finger-like protein (Fezl), a putative zinc-finger transcriptional repressor, is required for the differentiation of projection neurons in cortical layer 5. In Fezldeficient mice, these neurons display molecular, morphological, and axonal targeting defects. The corticospinal tract was absent in Fezl ؊/؊ mice, corticotectal and pontine projections were severely reduced, and Fezl-expressing neurons formed aberrant axonal projections. The expression of many molecular markers for deeplayer neurons was reduced or absent in the Fezl ؊/؊ cerebral cortex. Most strikingly, Ctip2, a transcription factor required for the formation of the corticospinal tract, was not expressed in the Fezl-deficient cortex. These results suggest that Fezl regulates the differentiation of layer 5 subcortical projection neurons.axon guidance ͉ cell fate ͉ corticospinal tract ͉ zinc-finger transcription factor

During the development of the nervous system, growing axons must traverse considerable distances to find their targets. In insects, this problem is solved in part by pioneer neurons, which lay down the first axonal pathways when distances are at a minimum. Here the existence of a similar kind of neuron in the developing mammalian telencephalon is described. These are the subplate cells, the first postmitotic neurons of the cerebral cortex. Axons from subplate neurons traverse the internal capsule and invade the thalamus early in fetal life, even before the neurons of cortical layers 5 and 6, which will form the adult subcortical projections, are generated. During postnatal life, after the adult pattern of axonal projections is firmly established, most subplate neurons disappear. These observations raise the possibility that the early axonal scaffold formed by subplate cells may prove essential for the establishment of permanent subcortical projections.

Discoveries from human and mouse genetics have identified cytoskeletal and signaling proteins that are essential for neuronal migration in the developing brain. To provide a meaningful context for these studies, we took an unbiased approach of correlative electron microscopy of neurons migrating through a three-dimensional matrix, and characterized the cytoskeletal events that occur as migrating neurons initiate saltatory forward movements of the cell nucleus. The formation of a cytoplasmic dilation in the proximal leading process precedes nuclear translocation. Cell nuclei translocate into these dilations in saltatory movements. Time-lapse imaging and pharmacological perturbation suggest that nucleokinesis requires stepwise or hierarchical interactions between microtubules, myosin II, and cell adhesion. We hypothesize that these interactions couple leading process extension to nuclear translocation during neuronal migration.actin ͉ microtubules ͉ myosin II ͉ blebbistatin ͉ motility N euronal migration is a central feature of mammalian brain development. Most neurons are born in proliferative zones that line the ventricles of the brain and travel long distances before settling into their final positions. Imaging studies of neurons migrating in the developing cerebral cortex reveal that individual neurons can switch between radial and tangential modes of migration (1-4). These observations imply that different types of migrating neurons use a common core mechanism for motility that is modulated by external guidance cues and substrates for migration.Migrating neurons show a number of behaviors that distinguish them from other types of migrating cells. First, migrating neurons have a characteristic leading process that extends over relatively long distances away from the nucleus and cell soma (5). By contrast, fibroblasts and neutrophils studied in vitro have broad lamellae at the leading edge and narrow, retractile tails (6). The movement of the nucleus in these cells is closely coupled to that of the leading edge. The leading process of a migrating neuron, however, seems to move almost autonomously with respect to the nucleus and cell soma. Leading processes exhibit a number of complex behaviors, including the extension of multiple processes and even polarity reversals in which the leading process is withdrawn completely and a new process extends from the cell rear (1). During such maneuvers, the cell body and nucleus remain largely stationary. It is only when a single leading process is consolidated and executes sustained movement in one direction that the cell body and nucleus rapidly advance to a discrete point, and the cycle of leading process motility begins again (7,8).The ability of neurons to ''uncouple'' movements of the leading process from cell soma translocation may reflect the importance of detecting guidance cues in vivo. The extension of a long leading process may enable the neuron to sample the environment before initiating somal migration, a strategy that would likely require subcellular speciali...