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1.
J Neurochem ; 149(2): 269-283, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30664245

RESUMO

Contributions of damaged mitochondria to neuropathologies have stimulated interest in mitophagy. We investigated triggers of neuronal mitophagy by disruption of mitochondrial energy metabolism in primary neurons. Mitophagy was examined in cultured murine cerebellar granule cells after inhibition of mitochondrial respiratory chain by drugs rotenone, 3-nitropropionic acid, antimycin A, and potassium cyanide, targeting complexes I, II, III, and IV, respectively. Inhibitor concentrations producing slow cellular demise were determined from analyses of cellular viability, morphology of neuritic damage, plasma membrane permeability, and oxidative phosphorylation. Live cell imaging of dissipation of mitochondrial membrane potential (ΔΨm ) by drugs targeting mitochondrial complexes was referenced to complete depolarization by carbonyl cyanide m-chlorophenyl hydrazone. While inhibition of complexes I, III and IV effected rapid dissipation of ΔΨm , inhibition of complex II using 3-nitropropionic acid led to minimal depolarization of mitochondria. Nonetheless, all respiratory chain inhibitors triggered mitophagy as indicated by increased aggregation of mitochondrially localized PINK1. Mitophagy was further analyzed using a dual fluorescent protein biosensor reporting mitochondrial relocation to acidic lysosomal environment. Significant acidification of mitochondria was observed in neurons treated with rotenone or 3-nitropropionic acid, revealing mitophagy at distal processes. Neurons treated with antimycin A or cyanide failed to show mitochondrial acidification. Minor dissipation of ΔΨm by 3-nitropropionic acid coupled with vigorous triggering of mitophagy suggested depolarization of mitochondria is not a necessary condition to trigger mitophagy. Moreover, weak elicitation of mitophagy by antimycin A, subsequent to loss of ΔΨm , suggested that mitochondrial depolarization is not a sufficient condition for triggering robust neuronal mitophagy. Our findings provide new insight into complexities of mitophagic clearance of neuronal mitochondria.

2.
Exp Neurol ; 295: 36-45, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28511841

RESUMO

The two neuronal populations in the cortex, pyramidal neurons and interneurons, can be separated based on neurotransmitter identity, however, within this segregation a large degree of diversity exists. Investigations into the molecular diversity of neurons are impeded by the inability to isolate cell populations born at different times for gene expression analysis. Developing interneurons may be distinguished by the expression of Glutamic Acid Decarboxylase-67 (GAD67). Neuronal birthdating using nucleoside analogs is an effective means of identifying coetaneous interneurons. Using these two features, neurotransmitter identity and birthdating, we have developed a method to isolate migrating interneurons using fluorescent-activated cell sorting (FACS) for RNA extraction and gene expression analysis. We utilized 5-ethynyl-2'-deoxyuridine (EdU) to birthdate interneuron cohorts and the GAD67 knock-in GFP transgenic mice to identify interneurons. In combination, we achieved simultaneous detection of GFP and EdU signals during FACS sorting of coetaneous interneurons with minimum loss of RNA integrity. RNA quality was deemed to be satisfactory by quantitative polymerase chain reaction (qPCR) for the interneuron-specific transcript Gad67.


Assuntos
Separação Celular/métodos , Córtex Cerebral/citologia , Expressão Gênica , Técnicas Genéticas , Interneurônios , Animais , Permeabilidade da Membrana Celular , Citometria de Fluxo/métodos , Regulação da Expressão Gênica no Desenvolvimento , Glutamato Descarboxilase/genética , Proteínas de Fluorescência Verde , Humanos , Camundongos , Camundongos Transgênicos , Células Piramidais , RNA/biossíntese , RNA/genética
3.
Cereb Cortex ; 27(1): 576-588, 2017 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-26503265

RESUMO

Mutations of the reelin gene cause severe defects in cerebral cortex development and profound intellectual impairment. While many aspects of the reelin signaling pathway have been identified, the molecular and ultimate cellular consequences of reelin signaling remain unknown. Specifically, it is unclear if termination of reelin signaling is as important for normal cortical neuron migration as activation of reelin signaling. Using mice that are single or double deficient, we discovered that combined loss of the suppressors of cytokine signaling, SOCS6 and SOCS7, recapitulated the cortical layer inversion seen in mice lacking reelin and led to a dramatic increase in the reelin signaling molecule disabled (DAB1) in the cortex. The SRC homology domains of SOCS6 and SOCS7 bound DAB1 ex vivo. Mutation of DAB1 greatly diminished binding and protected from degradation by SOCS6. Phosphorylated DAB1 was elevated in cortical neurons in the absence of SOCS6 and SOCS7. Thus, constitutive activation of reelin signaling was observed to be equally detrimental as lack of activation. We hypothesize that, by terminating reelin signaling, SOCS6 and SOCS7 may allow new cycles of reelin signaling to occur and that these may be essential for cortical neuron migration.


Assuntos
Moléculas de Adesão Celular Neuronais/metabolismo , Córtex Cerebral/embriologia , Córtex Cerebral/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Serina Endopeptidases/metabolismo , Proteínas Supressoras da Sinalização de Citocina/deficiência , Animais , Moléculas de Adesão Celular Neuronais/genética , Movimento Celular/fisiologia , Córtex Cerebral/patologia , Proteínas da Matriz Extracelular/genética , Células HEK293 , Humanos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas do Tecido Nervoso/genética , Neurônios/metabolismo , Fosforilação , Serina Endopeptidases/genética , Proteínas Supressoras da Sinalização de Citocina/genética
4.
PLoS One ; 9(10): e110415, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25334023

RESUMO

The characteristic six-layered appearance of the neocortex arises from the correct positioning of pyramidal neurons during development and alterations in this process can cause intellectual disabilities and developmental delay. Malformations in cortical development arise when neurons either fail to migrate properly from the germinal zones or fail to cease migration in the correct laminar position within the cortical plate. The Reelin signalling pathway is vital for correct neuronal positioning as loss of Reelin leads to a partially inverted cortex. The precise biological function of Reelin remains controversial and debate surrounds its role as a chemoattractant or stop signal for migrating neurons. To investigate this further we developed an in silico agent-based model of cortical layer formation. Using this model we tested four biologically plausible hypotheses for neuron motility and four biologically plausible hypotheses for the loss of neuron motility (conversion from migration). A matrix of 16 combinations of motility and conversion rules was applied against the known structure of mouse cortical layers in the wild-type cortex, the Reelin-null mutant, the Dab1-null mutant and a conditional Dab1 mutant. Using this approach, many combinations of motility and conversion mechanisms can be rejected. For example, the model does not support Reelin acting as a repelling or as a stopping signal. In contrast, the study lends very strong support to the notion that the glycoprotein Reelin acts as a chemoattractant for neurons. Furthermore, the most viable proposition for the conversion mechanism is one in which conversion is affected by a motile neuron sensing in the near vicinity neurons that have already converted. Therefore, this model helps elucidate the function of Reelin during neuronal migration and cortical development.


Assuntos
Moléculas de Adesão Celular Neuronais/metabolismo , Córtex Cerebral/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Modelos Biológicos , Proteínas do Tecido Nervoso/metabolismo , Neurônios/metabolismo , Serina Endopeptidases/metabolismo , Algoritmos , Animais , Moléculas de Adesão Celular Neuronais/deficiência , Moléculas de Adesão Celular Neuronais/genética , Embrião de Mamíferos/metabolismo , Desenvolvimento Embrionário , Proteínas da Matriz Extracelular/deficiência , Proteínas da Matriz Extracelular/genética , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Camundongos , Camundongos Knockout , Proteínas do Tecido Nervoso/deficiência , Proteínas do Tecido Nervoso/genética , Fenótipo , Serina Endopeptidases/deficiência , Serina Endopeptidases/genética , Transdução de Sinais
5.
Conf Proc IEEE Eng Med Biol Soc ; 2014: 1525-8, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25570260

RESUMO

The cerebral cortex is folded into gyri and sulci in the brains of higher mammals. Quantitative study of the process by which the cortex folds during brain development is critical to a complete understanding of normal brain development and neuro-developmental disorders. In this work, we propose a new method by which to localise nonlinearities in the cortical folding process, and thereby identify regions of differential growth across the cortex. Our method is based on spherical harmonic (SPHARM) representation of the cortical surface. Linearity is assessed by comparison of each SPHARM reconstructed surface with an artificial surface constructed using a linear combination of SPHARM coefficients from data at adjoining developmental time points. The resultant quantification of cortical folding development is easy to interpret, and the method has low computational cost. We demonstrate application to a set of experimental MRI data of fetal sheep brains, across key developmental timepoints as the cortex first folds during development.


Assuntos
Córtex Cerebral/embriologia , Processamento de Imagem Assistida por Computador/métodos , Imagem por Ressonância Magnética/métodos , Algoritmos , Animais , Mapeamento Encefálico/métodos , Desenvolvimento Fetal , Modelos Lineares , Morfogênese , Ovinos , Software , Fatores de Tempo , Tronco
6.
Cereb Cortex ; 24(12): 3289-300, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-23897647

RESUMO

Ubiquitin ligases of the Nedd4 family are important for axon and dendrite development, but little is known about their adaptor, Nedd4 family-interacting protein 1 (Ndfip1), that is responsible for their enzymatic activation. To study the function of Ndfip1 in cortical development, we generated a conditional knock-out (conditional KO) in neurons. The Ndfip1 conditional KO mice were viable; however, cortical neurons in the adult brain exhibited atrophic characteristics, including stunted dendritic arbors, blebbing of dendrites, and fewer dendritic spines. In electron micrographs, these neurons appeared shrunken with compacted somata and involutions of the nuclear membrane. In culture, Ndfip1 KO neurons exhibited exuberant sprouting suggesting loss of developmental control. Biochemical analysis of postsynaptic density (PSD) fractions from Ndfip1 KO cortical and hippocampal neurons showed that the postsynaptic proteins (Arc and PSD-95) were reduced compared with wild-type controls. In addition, the PI3 kinase/Akt signaling pathway was altered. These results indicate that Ndfip1, through its Nedd4 effectors, is important for the development of dendrites and dendritic spines in the cortex.


Assuntos
Proteínas de Transporte/genética , Espinhas Dendríticas/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Proteínas de Membrana/genética , Neocórtex , Células Piramidais/diagnóstico por imagem , Animais , Animais Recém-Nascidos , Fracionamento Celular , Células Cultivadas , Proteína 4 Homóloga a Disks-Large , Embrião de Mamíferos , Regulação da Expressão Gênica no Desenvolvimento/genética , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Guanilato Quinases/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Proteínas de Membrana/deficiência , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neocórtex/citologia , Neocórtex/embriologia , Neocórtex/crescimento & desenvolvimento , Nestina/genética , Nestina/metabolismo , PTEN Fosfo-Hidrolase/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Transfecção , Ultrassonografia
7.
Cereb Cortex ; 24(11): 2835-47, 2014 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-23749873

RESUMO

Malformations of cortical development can arise when projection neurons generated in the germinal zones fail to migrate properly into the cortical plate. This process is critically dependent on the Reelin glycoprotein, which when absent leads to an inversion of cortical layers and blurring of borders. Reelin has other functions including supporting neuron migration and maintaining their trajectories; however, the precise role on glial fiber-dependent or -independent migration of neurons remains controversial. In this study, we wish to test the hypothesis that migrating cortical neurons at different levels of the cortical wall have differential responses to Reelin. We exposed neurons migrating across the cortical wall to exogenous Reelin and monitored their migratory behavior using time-lapse imaging. Our results show that, in the germinal zones, exogenous Reelin retarded neuron migration and altered their trajectories. This behavior is in contrast to the response of neurons located in the intermediate zone (IZ), possibly because Reelin receptors are not expressed in this zone. In the reeler cortex, Reelin receptors are expressed in the IZ and exposure to exogenous Reelin was able to rescue the migratory defect. These studies demonstrate that migrating neurons have nonequivalent responses to Reelin depending on their location within the cortical wall.


Assuntos
Moléculas de Adesão Celular Neuronais/farmacologia , Movimento Celular/efeitos dos fármacos , Córtex Cerebral/citologia , Proteínas da Matriz Extracelular/farmacologia , Proteínas do Tecido Nervoso/farmacologia , Serina Endopeptidases/farmacologia , Proteínas Adaptadoras de Transporte Vesicular/deficiência , Proteínas Adaptadoras de Transporte Vesicular/genética , Fatores Etários , Fosfatase Alcalina/genética , Fosfatase Alcalina/metabolismo , Análise de Variância , Animais , Linhagem Celular Transformada , Movimento Celular/genética , Eletroporação , Embrião de Mamíferos , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Técnicas In Vitro , Camundongos , Camundongos Mutantes Neurológicos , Microscopia Confocal , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Técnicas de Cultura de Órgãos , Transfecção
8.
Neurosignals ; 20(3): 168-89, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22572780

RESUMO

The modulation of cortical activity by GABAergic interneurons is required for normal brain function and is achieved through the immense level of heterogeneity within this neuronal population. Cortical interneurons share a common origin in the ventral telencephalon, yet during the maturation process diverse subtypes are generated that form the characteristic laminar arrangement observed in the adult brain. The long distance tangential and short-range radial migration into the cortical plate is regulated by a combination of intrinsic and extrinsic signalling mechanisms, and a great deal of progress has been made to understand these developmental events. In this review, we will summarize current findings regarding the molecular control of subtype specification and provide a detailed account of the migratory cues influencing interneuron migration and lamination. Furthermore, a dysfunctional GABAergic system is associated with a number of neurological and psychiatric conditions, and some of these may have a developmental aetiology with alterations in interneuron generation and migration. We will discuss the notion of additional sources of interneuron progenitors found in human and non-human primates and illustrate how the disruption of early developmental events can instigate a loss in GABAergic function.


Assuntos
Movimento Celular/fisiologia , Córtex Cerebral/citologia , Interneurônios/citologia , Animais , Córtex Cerebral/fisiologia , Interneurônios/fisiologia
9.
EMBO J ; 31(8): 1879-92, 2012 Apr 18.
Artigo em Inglês | MEDLINE | ID: mdl-22395070

RESUMO

Asymmetric cell division plays an indispensable role during corticogenesis for producing new neurons while maintaining a self-renewing pool of apical progenitors. The cellular and molecular determinants favouring asymmetric division are not completely understood. Here, we identify a novel mechanism for generating cellular asymmetry through the active transportation and local translation of Cyclin D2 mRNA in the basal process. This process is regulated by a unique cis-regulatory sequence found in the 3' untranslated region (3'UTR) of the mRNA. Unequal inheritance of Cyclin D2 protein to the basally positioned daughter cell with the basal process confers renewal of the apical progenitor after asymmetric division. Conversely, depletion of Cyclin D2 in the apically positioned daughter cell results in terminal neuronal differentiation. We demonstrate that Cyclin D2 is also expressed in the developing human cortex within similar domains, thus indicating that its role as a fate determinant is ancient and conserved.


Assuntos
Divisão Celular , Ciclina D2/biossíntese , Regulação da Expressão Gênica , Neurônios/fisiologia , Regiões 3' não Traduzidas , Humanos , Neurônios/citologia , RNA Mensageiro/metabolismo
10.
Cereb Cortex ; 21(5): 1018-27, 2011 May.
Artigo em Inglês | MEDLINE | ID: mdl-20847150

RESUMO

The Reelin signaling pathway is essential for proper cortical development, but it is unclear to whether Reelin function is primarily important for cortical layering or neuron migration. It has been proposed that Reelin is perhaps required only for somal translocation but not glial-dependent locomotion. This implies that the location of neurons responding to Reelin is restricted to the outer regions of the cortical plate (CP). To determine whether Reelin is required for migration outside of the CP, we used time-lapse imaging to track the behavior of cells undergoing locomotion in the germinal zones. We focused on the migratory activity in the ventricular/subventricular zones where the first transition of bipolar to multipolar migration occurs and where functional Reelin receptors are known to be expressed. Despite Reelin loss, neurons had no difficulty in undergoing radial migration and indeed displayed greater migratory speed. Additionally, compared with the wild-type, reeler neurons displayed altered trajectories with greater deviation from a radial path. These results suggest that Reelin loss has early consequences for migration in the germinal zones that are portrayed as defective radial trajectories and migratory speeds. Together, these abnormalities can give rise to the increased cell dispersion observed in the reeler cortex.


Assuntos
Moléculas de Adesão Celular Neuronais/deficiência , Movimento Celular/genética , Proteínas da Matriz Extracelular/deficiência , Neocórtex/citologia , Proteínas do Tecido Nervoso/deficiência , Neurônios/patologia , Serina Endopeptidases/deficiência , Animais , Moléculas de Adesão Celular Neuronais/genética , Proteínas da Matriz Extracelular/genética , Camundongos , Camundongos Mutantes Neurológicos , Neocórtex/metabolismo , Neocórtex/patologia , Proteínas do Tecido Nervoso/genética , Malformações do Sistema Nervoso/genética , Malformações do Sistema Nervoso/metabolismo , Malformações do Sistema Nervoso/fisiopatologia , Neurônios/citologia , Neurônios/fisiologia , Técnicas de Cultura de Órgãos , Serina Endopeptidases/genética , Transmissão Sináptica/genética
11.
Cereb Cortex ; 20(9): 2017-26, 2010 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-20053715

RESUMO

Reelin is an important protein that is indispensable for cortical lamination. In the absence of Reelin, cortical layers fail to form due to inappropriate neuron migration and positioning. The inversion of cortical layers is attributed to failure of neurons to migrate past earlier-generated neurons although how Reelin-insufficiency causes this is unclear. The issue is complicated by recent studies showing that very little Reelin is required for cortical layering. To test how variation in the number of Reelin-producing cells is linked to cortical lamination, we have employed Reelin(+/+) <--> Reelin(-/-) chimeras in which the number of Reelin-expressing neurons is adjusted. We found that the Reeler phenotype was rescued in chimeras with a large contribution of Reelin(+/+) neurons; conversely in chimeras with a weak contribution by Reelin(+/+) neurons, the mutant phenotype remained. However, increasing the number of Reelin(+/+) neurons beyond an unknown threshold resulted in partial rescue, with the formation of a correctly layered secondary cortex lying on top of an inverted mutant cortex. Therefore, the development of cortical layers in the correct order requires a minimal level of Reelin protein to be present although paradoxically, this is insufficient to prevent the simultaneous formation of inverted cortical layers in the same hemisphere.


Assuntos
Padronização Corporal/genética , Moléculas de Adesão Celular Neuronais/biossíntese , Moléculas de Adesão Celular Neuronais/deficiência , Córtex Cerebral/anormalidades , Córtex Cerebral/crescimento & desenvolvimento , Proteínas da Matriz Extracelular/biossíntese , Proteínas da Matriz Extracelular/deficiência , Proteínas do Tecido Nervoso/biossíntese , Proteínas do Tecido Nervoso/deficiência , Neurônios/metabolismo , Serina Endopeptidases/biossíntese , Serina Endopeptidases/deficiência , Quimeras de Transplante/genética , Animais , Animais Recém-Nascidos , Moléculas de Adesão Celular Neuronais/genética , Movimento Celular/genética , Córtex Cerebral/metabolismo , Proteínas da Matriz Extracelular/genética , Feminino , Masculino , Camundongos , Camundongos Endogâmicos , Camundongos Knockout , Camundongos Transgênicos , Proteínas do Tecido Nervoso/genética , Malformações do Sistema Nervoso/genética , Malformações do Sistema Nervoso/metabolismo , Neurogênese/genética , Neurônios/patologia , Serina Endopeptidases/genética , Quimeras de Transplante/crescimento & desenvolvimento , Quimeras de Transplante/metabolismo
12.
Biophys J ; 97(3): 699-709, 2009 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-19651028

RESUMO

Migration is a dynamic process in which a cell searches the environment and translates acquired information into somal advancement. In particular, interneuron migration during development is accomplished by two distinct processes: the extension of neurites tipped with growth cones; and nucleus translocation, termed nucleokinesis. The primary purpose of our study is to investigate neurite branching and nucleokinesis using high-resolution time-lapse confocal microscopy and computational modeling. We demonstrate that nucleokinesis is accurately modeled by a spring-dashpot system and that neurite branching is independent of the nucleokinesis event, and displays the dynamics of a stochastic birth-death process. This is in contrast to traditional biological descriptions, which suggest a closer relationship between the two migratory mechanisms. Our models are validated on independent data sets acquired using two different imaging protocols, and are shown to be robust to alterations in guidance cues and cellular migratory mechanisms, through treatment with brain-derived neurotrophic factor, neurotrophin-4, and blebbistatin. We postulate that the stochastic branch dynamics exhibited by interneurons undergoing guidance-directed migration permit efficient exploration of the environment.


Assuntos
Movimento Celular , Interneurônios/fisiologia , Algoritmos , Animais , Encéfalo/crescimento & desenvolvimento , Encéfalo/fisiologia , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Simulação por Computador , Bases de Dados Factuais , Proteínas de Fluorescência Verde/genética , Compostos Heterocíclicos de 4 ou mais Anéis/metabolismo , Humanos , Camundongos , Camundongos Transgênicos , Microscopia Confocal/métodos , Modelos Neurológicos , Fatores de Crescimento Neural/metabolismo , Neuritos/fisiologia , Processos Estocásticos , Fatores de Tempo , Técnicas de Cultura de Tecidos
13.
Med Image Anal ; 13(6): 920-30, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19181561

RESUMO

Understanding the biomechanical mechanisms by which the cerebral cortex folds is a fundamental problem in neuroscience. Current mathematical models of cortical folding do not include three dimensional geometry or measurement of cortical growth in developing brains extracted from experimental data. We present two biomechanical models of cortical folding which integrate 3D geometry and information taken from MRI scans of fetal sheep brains at a number of key developmental stages. The first model utilises diffusion tensor imaging (DTI) measurements of white matter fibre orientation in the fetal sheep brains as a cue to the tension forces that may regulate folding. In the second model, tangential cortical growth is modelled by osmotic expansion of the tissue and regulated by inhomogeneous white matter rigidity as a biomechanism of cortical folding. This is based on quantitative analysis of cortical growth and inhomogeneous white matter anisotropy measured from the MRI data. We demonstrate that structural and diffusion tensor MRI can be combined with finite element modelling and an explicit growth mechanism of the cortex to create biologically meaningful models of the cortical folding process common to higher order mammals.


Assuntos
Córtex Cerebral/anatomia & histologia , Córtex Cerebral/crescimento & desenvolvimento , Modelos Biológicos , Morfogênese/fisiologia , Fibras Nervosas Mielinizadas/fisiologia , Fibras Nervosas Mielinizadas/ultraestrutura , Animais , Anisotropia , Córtex Cerebral/embriologia , Simulação por Computador , Módulo de Elasticidade/fisiologia , Ovinos , Estresse Mecânico
14.
Development ; 135(12): 2139-49, 2008 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-18506028

RESUMO

Neuronal migration is integral to the development of the cerebral cortex and higher brain function. Cortical neuron migration defects lead to mental disorders such as lissencephaly and epilepsy. Interaction of neurons with their extracellular environment regulates cortical neuron migration through cell surface receptors. However, it is unclear how the signals from extracellular matrix proteins are transduced intracellularly. We report here that mouse embryos lacking the Ras family guanine nucleotide exchange factor, C3G (Rapgef1, Grf2), exhibit a cortical neuron migration defect resulting in a failure to split the preplate into marginal zone and subplate and a failure to form a cortical plate. C3G-deficient cortical neurons fail to migrate. Instead, they arrest in a multipolar state and accumulate below the preplate. The basement membrane is disrupted and radial glial processes are disorganised and lack attachment in C3G-deficient brains. C3G is activated in response to reelin in cortical neurons, which, in turn, leads to activation of the small GTPase Rap1. In C3G-deficient cells, Rap1 GTP loading in response to reelin stimulation is reduced. In conclusion, the Ras family regulator C3G is essential for two aspects of cortex development, namely radial glial attachment and neuronal migration.


Assuntos
Córtex Cerebral/citologia , Córtex Cerebral/embriologia , Fator 2 de Liberação do Nucleotídeo Guanina/fisiologia , Neuroglia/fisiologia , Neurônios/fisiologia , Animais , Adesão Celular/fisiologia , Movimento Celular/fisiologia , Células Cultivadas , Cruzamentos Genéticos , Embrião de Mamíferos , Fator 2 de Liberação do Nucleotídeo Guanina/genética , Camundongos , Camundongos Endogâmicos CBA , Camundongos Endogâmicos , Mutação , Técnicas de Cultura de Órgãos , Células-Tronco/citologia
15.
Neuron ; 56(4): 621-39, 2007 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-18031681

RESUMO

Development of appropriate dendritic arbors is crucial for neuronal information transfer. We show, using seizure-related gene 6 (sez-6) null mutant mice, that Sez-6 is required for normal dendritic arborization of cortical neurons. Deep-layer pyramidal neurons in the somatosensory cortex of sez-6 null mice exhibit an excess of short dendrites, and cultured cortical neurons lacking Sez-6 display excessive neurite branching. Overexpression of individual Sez-6 isoforms in knockout neurons reveals opposing actions of membrane-bound and secreted Sez-6 proteins, with membrane-bound Sez-6 exerting an antibranching effect under both basal and depolarizing conditions. Layer V pyramidal neurons in knockout brain slices show reduced excitatory postsynaptic responses and a reduced dendritic spine density, reflected by diminished punctate staining for postsynaptic density 95 (PSD-95). In behavioral tests, the sez-6 null mice display specific exploratory, motor, and cognitive deficits. In conclusion, cell-surface protein complexes involving Sez-6 help to sculpt the dendritic arbor, in turn enhancing synaptic connectivity.


Assuntos
Córtex Cerebral/anormalidades , Córtex Cerebral/citologia , Dendritos/ultraestrutura , Regulação da Expressão Gênica no Desenvolvimento/genética , Proteínas do Tecido Nervoso/genética , Células Piramidais/citologia , Animais , Diferenciação Celular/genética , Membrana Celular/genética , Membrana Celular/metabolismo , Células Cultivadas , Córtex Cerebral/metabolismo , Transtornos Cognitivos/genética , Transtornos Cognitivos/metabolismo , Transtornos Cognitivos/fisiopatologia , Dendritos/metabolismo , Espinhas Dendríticas/metabolismo , Espinhas Dendríticas/ultraestrutura , Proteína 4 Homóloga a Disks-Large , Potenciais Pós-Sinápticos Excitadores/genética , Feminino , Guanilato Quinases , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Malformações do Sistema Nervoso/genética , Malformações do Sistema Nervoso/metabolismo , Malformações do Sistema Nervoso/fisiopatologia , Vias Neurais/anormalidades , Vias Neurais/citologia , Vias Neurais/metabolismo , Técnicas de Cultura de Órgãos , Técnicas de Patch-Clamp , Células Piramidais/metabolismo , Transmissão Sináptica/genética
16.
Cereb Cortex ; 16 Suppl 1: i57-63, 2006 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-16766708

RESUMO

The interactions between migrating interneurons and their environment that lead to stereotypic migration pathways remain largely undefined. We have used time-lapse imaging to record the migratory responses of labeled interneurons to different regions of the migratory pathway in organotypic slice cultures. We tested the hypothesis that the length of the migratory pathway is not equally permissive for interneuron migration, with separate zones of inhibition and attraction. Three different experimental approaches were used to address this issue, including explant cocultures, cortical overlay cultures, and rotation of cortical slices. The results clearly identify the lateral region to be an attractive substrate for interneuron entry at embryonic day 12.5, whereas the medial region at this stage contains a zone of inhibition. This property of the medial neocortex is temporally regulated with switching from inhibition to attraction within 24 h. We suggest that this temporal regulation may provide a mechanism for gating the entry of interneurons into the hippocampus while ensuring that cortical interneurons are properly confined within the neocortical wall. In this manner, interneurons arising from common precursors and sharing common migratory pathways are able to populate different pallial structures.


Assuntos
Interneurônios/citologia , Neocórtex/citologia , Neocórtex/embriologia , Rede Nervosa/citologia , Rede Nervosa/embriologia , Organogênese/fisiologia , Envelhecimento/patologia , Envelhecimento/fisiologia , Animais , Padronização Corporal/fisiologia , Agregação Celular , Diferenciação Celular , Movimento Celular , Técnicas In Vitro , Interneurônios/fisiologia , Camundongos , Neocórtex/crescimento & desenvolvimento , Rede Nervosa/crescimento & desenvolvimento
17.
Mol Cell Biol ; 24(18): 8221-6, 2004 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-15340081

RESUMO

The neuregulins (NRGs) are a family of four structurally related growth factors that are expressed in the developing and adult brain. NRG-1 is essential for normal heart formation and has been implicated in the development and maintenance of both neurons and glia. NRG-2 was identified on the basis of its homology to NRG-1 and, like NRG-1, is expressed predominantly by neurons in the central nervous system. We have generated mice with the active domain of NRG-2 deleted in an effort to characterize the biological function of NRG-2 in vivo. In contrast to the NRG-1 knockout animals, NRG-2 knockouts have no apparent heart defects and survive embryogenesis. Mutant mice display early growth retardation and reduced reproductive capacity. No obvious histological differences were observed in the major sites of NRG-2 expression. Our results indicate that in vivo NRG-2 activity differs substantially from that of NRG-1 and that it is not essential for normal development in utero.


Assuntos
Fatores de Crescimento Neural/deficiência , Fatores de Crescimento Neural/genética , Animais , Sequência de Bases , Encéfalo/crescimento & desenvolvimento , DNA Complementar/genética , Feminino , Marcação de Genes , Genes erbB , Coração/crescimento & desenvolvimento , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fatores de Crescimento Neural/fisiologia , Proteínas do Tecido Nervoso/deficiência , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/fisiologia , Neuregulina-1 , Fenótipo
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