Periventricular heterotopia (PH) is a disorder characterized by neuronal nodules, ectopically positioned along the lateral ventricles of the cerebral cortex. Mutations in either of two human genes, Filamin A (FLNA) or ADP-ribosylation factor guanine exchange factor 2 (ARFGEF2), cause PH (Fox et al. in 'Mutations in filamin 1 prevent migration of cerebral cortical neurons in human periventricular heterotopia'. Neuron, 21, 1315-1325, 1998; Sheen et al. in 'Mutations in ARFGEF2 implicate vesicle trafficking in neural progenitor proliferation and migration in the human cerebral cortex'. Nat. Genet., 36, 69-76, 2004). Recent studies have shown that mutations in mitogen-activated protein kinase kinase kinase-4 (Mekk4), an indirect interactor with FlnA, also lead to periventricular nodule formation in mice (Sarkisian et al. in 'MEKK4 signaling regulates filamin expression and neuronal migration'. Neuron, 52, 789-801, 2006). Here we show that neurons in post-mortem human PH brains migrated appropriately into the cortex, that periventricular nodules were primarily composed of later-born neurons, and that the neuroependyma was disrupted in all PH cases. As studied in the mouse, loss of FlnA or Big2 function in neural precursors impaired neuronal migration from the germinal zone, disrupted cell adhesion and compromised neuroepithelial integrity. Finally, the hydrocephalus with hop gait (hyh) mouse, which harbors a mutation in Napa [encoding N-ethylmaleimide-sensitive factor attachment protein alpha (alpha-SNAP)], also develops a progressive denudation of the neuroepithelium, leading to periventricular nodule formation. Previous studies have shown that Arfgef2 and Napa direct vesicle trafficking and fusion, whereas FlnA associates dynamically with the Golgi membranes during budding and trafficking of transport vesicles. Our current findings suggest that PH formation arises from a final common pathway involving disruption of vesicle trafficking, leading to impaired cell adhesion and loss of neuroependymal integrity.

Background: Malformations of cortical development (MCD) are increasingly recognized as an important cause of epilepsy and developmental delay. MCD encompass a wide spectrum of disorders with various underlying genetic etiologies and clinical manifestations. High resolution imaging has dramatically improved our recognition of MCD. Review Summary: This review will provide a brief overview of the stages of normal cortical development, including neuronal proliferation, neuroblast migration, and neuronal organization. Disruptions at various stages lead to characteristic MCD. Disorders of neurogenesis give rise to microcephaly (small brain) or macrocephaly (large brain). Disorders of early neuroblast migration give rise to periventricular heterotopia (neurons located along the ventricles), whereas abnormalities later in migration lead to lissencephaly (smooth brain) or subcortical band heterotopia (smooth brain with a band of heterotopic neurons under the cortex). Abnormal neuronal migration arrest give rise to over migration of neurons in cobblestone lissencephaly. Lastly, disorders of neuronal organization cause polymicrogyria (abnormally small gyri and sulci). This review will also discuss the known genetic mutations and potential mechanisms that contribute to these syndromes. Conclusion: Identification of various gene mutations has not only given us greater insight into some of the pathophysiologic basis of MCD, but also an understanding of the processes involved in normal cortical development.

The developmental mechanisms underlying the formation of human cortical convolutions (gyri and sulci) remain largely unknown. Genetic causes of lissencephaly (literally ‘smooth brain’) would imply that disorders in neuronal migration cause the loss of cortical convolutions. However, prior studies have suggested that loss of sulci and gyri can also arise from impaired proliferation, disrupted lamination and loss of intracortical connections. To gain further insight into the mechanisms underlying the formation of cortical convolutions, we examined the progressive brain development of the gyrencephalic ferret. In this study, we used magnetic resonance imaging to follow the temporal and spatial pattern of neuronal migration, proliferation and differentiation in relation to the onset and development of cortical convolutions. In this manner, we demonstrate that the onset of gyrification begins largely after completion of neuronal proliferation and migration. Gyrification occurs in a lateral to medial gradient, during the period of most rapid cerebral cortical growth. Cortical folding is also largely complete prior to myelination of the underlying cortical axons. These observations are consistent with gyrification arising secondary to cortical processes involving neuronal differentiation.

Down syndrome (DS) is caused by trisomy of chromosome 21 and is characterized by mental retardation, seizures and premature Alzheimer’s disease. To examine neuropathological mechanisms giving rise to this disorder, we generated multiple human DS neural progenitor cell (NPC) lines from the 19–21 week frontal cortex and characterized their genomic and functional properties. Microarray profiling of DS progenitors indicated that increased levels of gene expression were not limited to chromosome 21, suggesting that increased expression of genes on chromosome 21 altered transcriptional regulation of a subset of genes throughout the entire genome. Moreover, many transcriptionally dysregulated genes were involved in cell death and oxidative stress. Network analyses suggested that upregulated expression of chromosome 21 genes such as S100B and amyloid precursor protein activated the stress response kinase pathways, and furthermore, could be linked to upregulation of the water channel aquaporin 4 (AQP4). We further demonstrate in DS NPCs that S100B is constitutively overexpressed, that overexpression leads to increased reactive oxygen species (ROS) formation and activation of stress response kinases, and that activation of this pathway results in compensatory AQP4 expression. In addition, AQP4 expression could be induced by direct exposure to ROS, and siRNA inhibition of AQP4 resulted in elevated levels of ROS following S100B exposure. Finally, elevated levels of S100B-induced ROS and loss of AQP4 expression led to increased programmed cell death. These findings suggest that dysregulation of chromosome 21 genes in DS neural progenitors leads to increased ROS and thereby alters transcriptional regulation of cytoprotective, non-chromosome 21 genes in response to ongoing cellular insults.

Cytoskeleton-associated proteins play key roles not only in regulating cell morphology and migration but also in proliferation. Mutations in the cytoskeleton-associated genefilamin A(FlnA) cause the human disorder periventricular heterotopia (PH). PH is a disorder of neural stem cell development that is characterized by disruption of progenitors along the ventricular epithelium and subsequent formation of ectopic neuronal nodules. FlnA-dependent regulation of cytoskeletal dynamics is thought to direct neural progenitor migration and proliferation. Here we show that embryonicFlnA-null mice exhibited a reduction in brain size and decline in neural progenitor numbers over time. The drop in the progenitor population was not attributable to cell death or changes in premature differentiation, but to prolonged cell cycle duration. Suppression of FlnA led to prolongation of the entire cell cycle length, principally in M phase. FlnA loss impaired degradation of cyclin B1-related proteins, thereby delaying the onset and progression through mitosis. We found that the cdk1 kinase Wee1 bound FlnA, demonstrated increased expression levels after loss of FlnA function, and was associated with increased phosphorylation of cdk1. Phosphorylation of cdk1 inhibited activation of the anaphase promoting complex degradation system, which was responsible for cyclin B1 degradation and progression through mitosis. Collectively, our results demonstrate a molecular mechanism whereby FlnA loss impaired G2 to M phase entry, leading to cell cycle prolongation, compromised neural progenitor proliferation, and reduced brain size.

Filamin B (FLNB) is a cytoplasmic protein that regulates the cytoskeletal network by cross-linking actin, linking cell membrane to the cytoskeleton and regulating intracellular signaling pathways responsible for skeletal development (Stossel, T.P., Condeelis, J., Cooley, L., Hartwig, J.H., Noegel, A., Schleicher, M. and Shapiro, S.S. (2001) Filamins as integrators of cell mechanics and signalling. Nat. Rev. Mol. Cell Biol. , 2 , 138–145). Mutations in FLNB cause human skeletal disorders [boomerang dysplasia, spondylocarpotarsal (SCT), Larsen, and atelosteogenesis I/III syndromes], which are characterized by disrupted vertebral segmentation, joint formation and endochondral ossification [Krakow, D., Robertson, S.P., King, L.M., Morgan, T., Sebald, E.T., Bertolotto, C., Wachsmann-Hogiu, S., Acuna, D., Shapiro, S.S., Takafuta, T. et al. (2004) Mutations in the gene encoding filamin B disrupt vertebral segmentation, joint formation and skeletogenesis. Nat. Genet. , 36, 405–410; Bicknell, L.S., Morgan, T., Bonafe, L., Wessels, M.W., Bialer, M.G., Willems, P.J., Cohn, D.H., Krakow, D. and Robertson, S.P. (2005) Mutations in FLNB cause boomerang dysplasia. J. Med. Genet. , 42, e43]. Here we show that Flnb deficient mice have shortened distal limbs with small body size, and develop fusion of the ribs and vertebrae, abnormal spinal curvatures, and dysmorphic facial/calvarial bones, similar to the human phenotype. Characterization of the mutant mice demonstrated increased apoptosis along the bone periphery of the distal appendages, consistent with reduced bone width. No changes in the initial proliferative rate of chondrocytes were observed, but the progressive differentiation of chondrocyte precursors was impaired, consistent with reduced bone length. The extracellular matrix appeared disrupted and phosphorylated β1-integrin (a collagen receptor and Flnb binding partner) expression was diminished in the mutant growth plate. Like integrin-deficient chondrocytes, adhesion to the ECM was decreased in Flnb (−/−) chondrocytes, and inhibition of β1-integrin in these cells led to further impairments in cell spreading. These data suggest that disruption of the ECM-β1-integrin-Flnb pathway contributes to defects in vertebral and distal limb development, similar to those seen in the human autosomal recessive SCT due to Flnb mutations.