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1.
Biomed Res ; 40(5): 179-188, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31597903

RESUMO

Single prenatal exposure to valproic acid (VPA) in rodents is a widely used preclinical model of autism spectrum disorder (ASD). Continuous prenatal VPA exposure has been recently proposed as a new ASD model that closely captures the neuropathological features of ASD, including increases in cerebral cortex volume and the number of cortical upper layer neurons. We investigated the influence of prenatal VPA exposure on the behavior of adult offspring of pregnant dams that received intraperitoneal injections of VPA twice on one day during the genesis of cortical upper layer neurons. Mice exposed to VPA at E14 (E14-VPA) showed typical behavior abnormalities including reduced social interaction, hyperactivity, and poor maze learning due to attention deficit/impulsivity relative to healthy controls. Histological analysis revealed that E14-VPA mice had significantly increased neuronal density and impaired neural activity in the prefrontal cortex, but not the somatosensory area, which is likely linked to the observed abnormalities in social behavior. These results suggest that this VPA exposure method is a good model for gaining new insights into the underlying neuropathology of ASD.


Assuntos
Transtorno do Deficit de Atenção com Hiperatividade/etiologia , Comportamento Animal/efeitos dos fármacos , Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/embriologia , Exposição Materna/efeitos adversos , Organogênese/efeitos dos fármacos , Comportamento Social , Ácido Valproico/efeitos adversos , Animais , Feminino , Aprendizagem em Labirinto/efeitos dos fármacos , Camundongos , Córtex Pré-Frontal/efeitos dos fármacos , Córtex Pré-Frontal/embriologia , Gravidez , Efeitos Tardios da Exposição Pré-Natal
2.
Nat Commun ; 10(1): 3946, 2019 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-31477701

RESUMO

Cerebral cortex expansion is a hallmark of mammalian brain evolution; yet, how increased neurogenesis is coordinated with structural and functional development remains largely unclear. The T-box protein TBR2/EOMES is preferentially enriched in intermediate progenitors and supports cortical neurogenesis expansion. Here we show that TBR2 regulates fine-scale spatial and circuit organization of excitatory neurons in addition to enhancing neurogenesis in the mouse cortex. TBR2 removal leads to a significant reduction in neuronal, but not glial, output of individual radial glial progenitors as revealed by mosaic analysis with double markers. Moreover, in the absence of TBR2, clonally related excitatory neurons become more laterally dispersed and their preferential synapse development is impaired. Interestingly, TBR2 directly regulates the expression of Protocadherin 19 (PCDH19), and simultaneous PCDH19 expression rescues neurogenesis and neuronal organization defects caused by TBR2 removal. Together, these results suggest that TBR2 coordinates neurogenesis expansion and precise microcircuit assembly via PCDH19 in the mammalian cortex.


Assuntos
Caderinas/genética , Córtex Cerebral/metabolismo , Neurogênese/genética , Neurônios/metabolismo , Proteínas com Domínio T/genética , Animais , Caderinas/metabolismo , Córtex Cerebral/citologia , Córtex Cerebral/embriologia , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica no Desenvolvimento , Células HEK293 , Humanos , Camundongos Knockout , Camundongos Transgênicos , Interferência de RNA , Sinapses/metabolismo , Proteínas com Domínio T/metabolismo
3.
Biol Pharm Bull ; 42(9): 1545-1553, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31474714

RESUMO

The aim of the present study is to construct and characterize a novel three-dimensional culture system for mouse neurons using the functional polymer, FP001. Stereoscopically extended neurites were found in primary mouse cortical neurons cultured in the FP001-containing medium. Neurons cultured with FP001 were distributed throughout the medium of the observation range whereas neurons cultured without FP001 were distributed only on the bottom of the dish. These results demonstrated that neurons can be three-dimensionally cultured using the FP001-containing medium. The mRNA expression of the glutamatergic neuronal marker vesicular glutamate transporter 1 in neurons cultured in the FP001-containing medium were higher than that in neurons cultured in the FP001-free medium. Expression of the matured neuronal marker, microtubule-associated protein 2 (MAP2) a,b, and the synapse formation marker, Synapsin I, in neurons cultured with FP001 was also higher than that in neurons cultured without FP001. The expression pattern of MAP2a,b in neurons cultured with FP001, but not that in neurons cultured without FP001, was similar to that in the embryonic cerebral cortex. Exposure to glutamate significantly increased 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction activity in neurons cultured with FP001 compared to that in neurons cultured without FP001. These results suggested that glutamatergic neurotransmission in neurons three-dimensionally cultured in the FP001-containing medium may be upregulated compared to neurons two-dimensionally cultured in the FP001-free medium. Thus, neurons with the properties close to those in the embryonic brain could be obtained by three-dimensionally culturing neurons using FP001, compared to two-dimensional culture with a conventional adhesion method.


Assuntos
Técnicas de Cultura de Células/métodos , Córtex Cerebral/citologia , Meios de Cultura/química , Neurônios/citologia , Polissacarídeos Bacterianos/química , Animais , Córtex Cerebral/embriologia , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos ICR , Proteínas Associadas aos Microtúbulos/metabolismo , Neuritos/metabolismo , Neurônios/metabolismo , Sinapsinas/metabolismo
4.
Med Hypotheses ; 131: 109309, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31443781

RESUMO

Stroke and traumatic brain injury (TBI) are significant clinical problems characterized by high rate of mortality and long-lasting disabilities, and an unmet need for new treatments. Current experimental stroke and TBI research are evolving to focus more on understanding the brain's self-protective mechanisms to meet the critical need of developing new therapies for these disorders. In this hypothesis-based manuscript, I provide several lines of evidence that peptidase neurolysin (Nln) is one of the brain's potent, self-protective mechanisms promoting preservation and recovery of the brain after acute injury. Based on published experimental observations and ongoing studies in our laboratory, I posit that Nln is a compensatory and cerebroprotective mechanism in the post-stroke/TBI brain that functions to process a diverse group of extracellular neuropeptides and by that to reduce excitotoxicity, oxidative stress, edema formation, blood brain barrier hyper-permeability, and neuroinflammation. If this hypothesis is correct, Nln could potentially serve as a single therapeutic target to modulate the function of multiple targets, the involved neuropeptide systems, critically involved in various mechanisms of brain injury and cerebroprotection/restoration. Such multi-pathway target would be highly desired for pharmacotherapy of stroke and TBI, because targeting one pathophysiological pathway has proven to be ineffective for such complex disorders.


Assuntos
Lesões Encefálicas Traumáticas/metabolismo , Metaloendopeptidases/fisiologia , Proteínas do Tecido Nervoso/fisiologia , Neuropeptídeos/metabolismo , Acidente Vascular Cerebral/metabolismo , Doença Aguda , Animais , Lesões Encefálicas Traumáticas/fisiopatologia , Morte Celular , Hipóxia Celular , Células Cultivadas , Córtex Cerebral/embriologia , Córtex Cerebral/metabolismo , Terapia Genética , Humanos , Metaloendopeptidases/biossíntese , Metaloendopeptidases/genética , Camundongos , Proteínas do Tecido Nervoso/biossíntese , Proteínas do Tecido Nervoso/genética , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Neurônios/patologia , Neurotoxinas/farmacologia , Bulbo Olfatório/metabolismo , Estresse Oxidativo , Oxigênio/farmacologia , Ratos , Proteínas Recombinantes/metabolismo , Sistema Renina-Angiotensina/fisiologia , Acidente Vascular Cerebral/fisiopatologia , Acidente Vascular Cerebral/terapia , Regulação para Cima
5.
Int J Dev Neurosci ; 76: 72-79, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31299388

RESUMO

The aim of this study was to evaluate the in vivo impacts of maternal n-3 polyunsaturated fatty acids (PUFAs) deficiency during pregnancy on the proliferation of neural progenitor cells (NPCs) in the developing cerebral cortex of fetal rats. Our results showed that about 5 weeks of maternal dietary n-3 PUFAs deprivation resulted in a substantial n-3 PUFA deficiency in fetal rat cerebral cortex. Importantly, by two survival schemes and two quantitative methods, we found that maternal intake of n-3 PUFAs deficient diet during the gestation significantly inhibited the proliferation of NPCs in fetal rat cerebral cortex. Moreover, the decreased cortical NPCs proliferation induced by nutritional n-3 PUFAs restriction did not originate from the increased NPCs apoptosis. Finally, our observations indicated that the down-regulation of cyclin E protein might be involved in the inhibitory effects of maternal n-3 PUFAs deficient diet on the proliferation of cortical NPCs. These findings highlight the importance of maternal intake of appropriate n-3 PUFAs and deepen our understanding of the exact effects of n-3 PUFAs on mammalian brain development.


Assuntos
Córtex Cerebral/embriologia , Ácidos Graxos Ômega-3/deficiência , Células-Tronco Neurais , Animais , Apoptose , Proliferação de Células , Córtex Cerebral/citologia , Ciclinas/biossíntese , Ciclinas/genética , Dieta , Ácidos Graxos/metabolismo , Feminino , Feto/metabolismo , Masculino , Fenômenos Fisiológicos da Nutrição Materna , Gravidez , Ratos , Ratos Wistar
6.
Yi Chuan ; 41(6): 524-533, 2019 Jun 20.
Artigo em Chinês | MEDLINE | ID: mdl-31257200

RESUMO

Normal development of the cerebral cortex is a basis for the formation and function of mammalian brains. During this process, the radial migration of cortical neurons, as well as the axon projection into specific layers, are the most important steps regulated by some transcription factors, but the underlying molecular mechanisms are still obscure. BMAL1 (brain and muscle Arnt-like protein 1) is a newly identified transcription factor that plays important roles in the circadian rhythms. It was recently found to regulate the proliferation of hippocampal neuronal progenitor/precursor cells (NPCs), implicating Bmal1 in the brain development. Here we employed both RT-RCR and real-time PCR to explore the expression pattern of the Bmal1 gene in the developing brain. We found BMAl1 is enriched in the brain cortex during the perinatal stages and peaked in P3 mouse brains. Combined with in utero electroporation and interference with RNAi, we found that reducing the expression level of Bmal1 in neurons, the radial migration of embryonic cortical neurons was largely delayed, in a gene dose-effect pattern. Moreover, reducing the level of Bmal1 expression in mouse brains, the axonal projection in the corpus callosum was also disrupted from ipsilateral to the lateral cerebral hemisphere. These findings indicate that BMAL1 is essential for the radial migration of neurons in the cerebral cortex and the axonal projection of the corpus callosum, providing insights into the molecular mechanisms of cerebral cortex development.


Assuntos
Fatores de Transcrição ARNTL/fisiologia , Axônios , Movimento Celular , Córtex Cerebral/embriologia , Neurogênese , Animais , Feminino , Camundongos , Gravidez
7.
Nat Commun ; 10(1): 2396, 2019 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-31160561

RESUMO

Modern genetic studies indicate that human brain evolution is driven primarily by changes in gene regulation, which requires understanding the biological function of largely non-coding gene regulatory elements, many of which act in tissue specific manner. We leverage chromatin interaction profiles in human fetal and adult cortex to assign three classes of human-evolved elements to putative target genes. We find that human-evolved elements involving DNA sequence changes and those involving epigenetic changes are associated with human-specific gene regulation via effects on different classes of genes representing distinct biological pathways. However, both types of human-evolved elements converge on specific cell types and laminae involved in cerebral cortical expansion. Moreover, human evolved elements interact with neurodevelopmental disease risk genes, and genes with a high level of evolutionary constraint, highlighting a relationship between brain evolution and vulnerability to disorders affecting cognition and behavior. These results provide novel insights into gene regulatory mechanisms driving the evolution of human cognition and mechanisms of vulnerability to neuropsychiatric conditions.


Assuntos
Córtex Cerebral/embriologia , Epigênese Genética/genética , Evolução Molecular , Regulação da Expressão Gênica no Desenvolvimento/genética , Células-Tronco Neurais/metabolismo , Transtornos do Neurodesenvolvimento/genética , Encéfalo/embriologia , Encéfalo/metabolismo , Córtex Cerebral/metabolismo , Predisposição Genética para Doença , Humanos , Elementos Reguladores de Transcrição/genética
8.
Neurology ; 93(3): 114-123, 2019 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-31221716

RESUMO

De novo pathogenic variants in STXBP1 encoding syntaxin1-binding protein (STXBP1, also known as Munc18-1) lead to a range of early-onset neurocognitive conditions, most commonly early infantile epileptic encephalopathy type 4 (EIEE4, also called STXBP1 encephalopathy), a severe form of epilepsy associated with developmental delay/intellectual disability. Other neurologic features include autism spectrum disorder and movement disorders. The progression of neurologic symptoms has been reported in a few older affected individuals, with the appearance of extrapyramidal features, reminiscent of early onset parkinsonism. Understanding the pathologic process is critical to improving therapies, as currently available antiepileptic drugs have shown limited success in controlling seizures in EIEE4 and there is no precision medication approach for the other neurologic features of the disorder. Basic research shows that genetic knockout of STXBP1 or other presynaptic proteins of the exocytic machinery leads to widespread perinatal neurodegeneration. The mechanism that regulates this effect is under scrutiny but shares intriguing hallmarks with classical neurodegenerative diseases, albeit appearing early during brain development. Most critically, recent evidence has revealed that STXBP1 controls the self-replicating aggregation of α-synuclein, a presynaptic protein involved in various neurodegenerative diseases that are collectively known as synucleinopathies, including Parkinson disease. In this review, we examine the tantalizing link among STXBP1 function, EIEE, and the neurodegenerative synucleinopathies, and suggest that neural development in EIEE could be further affected by concurrent synucleinopathic mechanisms.


Assuntos
Proteínas Munc18/genética , Transtornos do Neurodesenvolvimento/genética , Espasmos Infantis/genética , /genética , Animais , Córtex Cerebral/embriologia , Exocitose/genética , Humanos , Chaperonas Moleculares , Proteínas Munc18/metabolismo , Transtornos do Neurodesenvolvimento/metabolismo , Transtornos do Neurodesenvolvimento/fisiopatologia , Crescimento Neuronal/genética , Espasmos Infantis/metabolismo , Espasmos Infantis/fisiopatologia , /fisiopatologia , alfa-Sinucleína/metabolismo
9.
Nutrients ; 11(5)2019 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-31121888

RESUMO

The pharmacological properties of Eleutherococcus senticosus leaf have not been clarified although it is taken as a food item. In this study, the effects of water extract of Eleutherococcus senticosus leaves on memory function were investigated in normal mice. Oral administration of the extract for 17 days significantly enhanced object recognition memory. Compounds absorbed in blood and the brain after oral administration of the leaf extract were detected by LC-MS/MS analyses. Primarily detected compounds in plasma and the cerebral cortex were ciwujianoside C3, eleutheroside M, ciwujianoside B, and ciwujianoside A1. Pure compounds except for ciwujianoside A1 were administered orally for 17 days to normal mice. Ciwujianoside C3, eleutheroside M, and ciwujianoside B significantly enhanced object recognition memory. These results demonstrated that oral administration of the leaf extract of E. senticosus enhances memory function, and that active ingredients in the extract, such as ciwujianoside C3, eleutheroside M, and ciwujianoside B, were able to penetrate and work in the brain. Those three compounds as well as the leaf extract had dendrite extension activity against primary cultured cortical neurons. The effect might relate to memory enhancement.


Assuntos
Encéfalo/efeitos dos fármacos , Eleutherococcus/química , Memória/efeitos dos fármacos , Extratos Vegetais/farmacologia , Folhas de Planta/química , Animais , Barreira Hematoencefálica/metabolismo , Encéfalo/metabolismo , Células Cultivadas , Córtex Cerebral/química , Córtex Cerebral/embriologia , Córtex Cerebral/ultraestrutura , Dendritos/efeitos dos fármacos , Dendritos/fisiologia , Glicosídeos/análise , Glicosídeos/farmacocinética , Glicosídeos/farmacologia , Masculino , Camundongos , Extratos Vegetais/análise , Extratos Vegetais/farmacocinética , Saponinas/análise , Saponinas/farmacocinética , Saponinas/farmacologia
10.
Dev Cell ; 49(4): 632-642.e7, 2019 05 20.
Artigo em Inglês | MEDLINE | ID: mdl-31112699

RESUMO

While it is now appreciated that certain long noncoding RNAs (lncRNAs) have important functions in cell biology, relatively few have been shown to regulate development in vivo, particularly with genetic strategies that establish cis versus trans mechanisms. Pnky is a nuclear-enriched lncRNA that is transcribed divergently from the neighboring proneural transcription factor Pou3f2. Here, we show that conditional deletion of Pnky from the developing cortex regulates the production of projection neurons from neural stem cells (NSCs) in a cell-autonomous manner, altering postnatal cortical lamination. Surprisingly, Pou3f2 expression is not disrupted by deletion of the entire Pnky gene. Moreover, expression of Pnky from a BAC transgene rescues the differential gene expression and increased neurogenesis of Pnky-knockout NSCs, as well as the developmental phenotypes of Pnky-deletion in vivo. Thus, despite being transcribed divergently from a key developmental transcription factor, the lncRNA Pnky regulates development in trans.


Assuntos
Córtex Cerebral/embriologia , Células-Tronco Neurais/metabolismo , RNA Longo não Codificante/genética , Animais , Encéfalo/metabolismo , Córtex Cerebral/metabolismo , Feminino , Interneurônios/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas do Tecido Nervoso/genética , Neurogênese/genética , Neurônios/metabolismo , Fatores do Domínio POU/genética , RNA Longo não Codificante/metabolismo , Transativadores/genética , Transativadores/metabolismo , Fatores de Transcrição/metabolismo
11.
Brain Nerve ; 71(4): 415-421, 2019 Apr.
Artigo em Japonês | MEDLINE | ID: mdl-30988231

RESUMO

The cerebral cortex is constructed through massive cell-production and accompanying cellular movements, which should be efficient and safe under spatiotemporal limitations. Efficiency and safety are also needed in the "production logistics" of manufacturing companies, and the "crowd dynamics" pertaining to people or vehicles. Investigating the growth of the embryonic cerebral cortex with an insight into such a system-level management of collective flows has recently revealed interesting cellular strategies to combat the spatiotemporal limitations. First, two sibling cells enter a narrow outflow space to overcome a bottleneck via a sequential departure similar to staggered commuting. This is achieved by asymmetric inheritance from the mother cell to one of the sibling cells with respect to cellular structure, and is analogous to a priority boarding pass. Second, newly generated sibling cells passively propel due to a mechanical force from the surrounding space. This space, densely filled with neighboring cells' processes, is elastic and assists in an indirect mother-to-daughter energy transfer. The mother cell has stored mechanical energy during its lateral/centrifugal expansion stage of mitosis while the surrounding space centripetally recoils to push the daughter cells. Third, young neurons act as a mechanical barrier to the movement of the mother cells thereby ensuring their normal proliferation.


Assuntos
Córtex Cerebral/embriologia , Neurônios/citologia , Movimento Celular , Proliferação de Células , Humanos
12.
Brain Dev ; 41(7): 567-576, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30954358

RESUMO

BACKGROUNDS: Metabotropic glutamate receptors, besides ionotropic receptors, mediate the complicated effect of glutamate on neurogenesis. Previous studies showed that metabotropic glutamate receptor 4 (mGluR4) regulated the proliferation and differentiation of neural stem/progenitor cells in vitro. However, little is known about the expression pattern of mGluR4 on prenatal central nervous system in vivo, especially the human being. METHODS: The normal brain tissues of human fetus were collected and divided into 4 groups according to the gestational age: 9-11 W, 14-16 W, 22-24 W and 32-36 W. Then the expression of mGluR4 was evaluated at mRNA and protein levels by means of PCR or immunohistochemistry method, respectively. The type of cell expressing mGluR4 was further investigated using double-labeling immunofluorescence. RESULTS: RT-PCR showed that the mRNA of mGluR4 could be detected in frontal lobe from 9 W to 32 W and real-time PCR quantificationally demonstrated the mRNA increased with development. Similarly, immnoreactivity was found in all layers of frontal lobe, VZ/SVZ. The intensity scores analysis showed that the staining became stronger and the range extended gradually with development. The double-labeling immunofluorescence showed that mGluR4 was present in neural stem/progenitor cells (nestin-positive cells after 9 W), young neurons (DCX-positive cells after 9 W), mature neurons (NeuN-positive cells in cortex after 32 W), as well as typical astrocytes (GFAP-positive cells in medulla after 32 W). CONCLUSION: These results supply an important evidence that mGluR4 is expressed in prenatal human cerebrum, and main kinds of cells related to neurogenesis are involved in its expression.


Assuntos
Encéfalo/embriologia , Lobo Frontal/embriologia , Receptores de Glutamato Metabotrópico/metabolismo , Encéfalo/metabolismo , Diferenciação Celular/fisiologia , Sistema Nervoso Central/citologia , Sistema Nervoso Central/embriologia , Sistema Nervoso Central/metabolismo , Córtex Cerebral/citologia , Córtex Cerebral/embriologia , Córtex Cerebral/metabolismo , Feminino , Desenvolvimento Fetal/genética , Lobo Frontal/citologia , Lobo Frontal/metabolismo , Ácido Glutâmico/metabolismo , Humanos , Imuno-Histoquímica , Masculino , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Neurogênese/fisiologia , Neurônios/metabolismo , Gravidez , Receptores de Glutamato Metabotrópico/genética
13.
J Mol Neurosci ; 68(4): 539-548, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30982164

RESUMO

Rap1 and N-cadherin regulate glia-independent translocation of cortical neurons. It remains unclear how Rap1 regulates N-cadherin-mediated neuronal migration. Here, we overexpressed Rap1gap in mouse brains (embryonic day 16) to inactivate Rap1, and observed that neurons did not migrate to the outer layer. We confirmed that Rap1 was involved in the regulation of late neurons in vivo. Rap1gap overexpression and Rap1 suppression in CHO cells decreased the expression of cytoskeletal proteins such as tubulin. Changes in the expression of cell morphology regulators, such as N-cadherin and ß-catenin, were also observed. Inhibition of N-cadherin in mouse brains prevented neuronal migration to the outer layer. The morphology of CHO cells was changed after overexpression of Rap1gap. We propose that Rap1 regulates the expression of N-cadherin during embryonic development, which affects ß-catenin expression. Beta-catenin in turn regulates cytoskeletal protein expression, ultimately affecting neuronal morphology and migration.


Assuntos
Caderinas/metabolismo , Movimento Celular , Neurônios/metabolismo , Proteínas rap1 de Ligação ao GTP/metabolismo , Animais , Células CHO , Córtex Cerebral/citologia , Córtex Cerebral/embriologia , Córtex Cerebral/metabolismo , Cricetinae , Cricetulus , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/fisiologia , Tubulina (Proteína)/genética , Tubulina (Proteína)/metabolismo , beta Catenina/genética , beta Catenina/metabolismo , Proteínas rap1 de Ligação ao GTP/genética
14.
Genes Dev ; 33(5-6): 365-376, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30808661

RESUMO

Synaptotagmin-11 (Syt11) is a Synaptotagmin isoform that lacks an apparent ability to bind calcium, phospholipids, or SNARE proteins. While human genetic studies have linked mutations in the Syt11 gene to schizophrenia and Parkinson's disease, the localization or physiological role of Syt11 remain unclear. We found that in neurons, Syt11 resides on abundant vesicles that differ from synaptic vesicles and resemble trafficking endosomes. These vesicles recycle via the plasma membrane in an activity-dependent manner, but their exocytosis is slow and desynchronized. Constitutive knockout mice lacking Syt11 died shortly after birth, suggesting Syt11-mediated membrane transport is required for survival. In contrast, selective ablation of Syt11 in excitatory forebrain neurons using a conditional knockout did not affect life span but impaired synaptic plasticity and memory. Syt11-deficient neurons displayed normal secretion of fast neurotransmitters and peptides but exhibited a reduction of long-term synaptic potentiation. Hence, Syt11 is an essential component of a neuronal vesicular trafficking pathway that differs from the well-characterized synaptic vesicle trafficking pathway but is also essential for life.


Assuntos
Plasticidade Neuronal/genética , Neurônios/fisiologia , Vesículas Sinápticas/metabolismo , Sinaptotagminas/genética , Sinaptotagminas/metabolismo , Animais , Córtex Cerebral/embriologia , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Técnicas de Introdução de Genes , Hipocampo/fisiopatologia , Memória/fisiologia , Camundongos , Camundongos Knockout , Neurotransmissores/metabolismo , Prosencéfalo/citologia , Prosencéfalo/fisiologia , Potenciais Sinápticos/genética , Transmissão Sináptica , Vesículas Sinápticas/genética , Sinaptotagminas/deficiência
15.
Neurosci Bull ; 35(3): 497-506, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30790215

RESUMO

Neuroligins (NLs) are postsynaptic cell-adhesion proteins that play important roles in synapse formation and the excitatory-inhibitory balance. They have been associated with autism in both human genetic and animal model studies, and affect synaptic connections and synaptic plasticity in several brain regions. Yet current research mainly focuses on pyramidal neurons, while the function of NLs in interneurons remains to be understood. To explore the functional difference among NLs in the subtype-specific synapse formation of both pyramidal neurons and interneurons, we performed viral-mediated shRNA knockdown of NLs in cultured rat cortical neurons and examined the synapses in the two major types of neurons. Our results showed that in both types of neurons, NL1 and NL3 were involved in excitatory synapse formation, and NL2 in GABAergic synapse formation. Interestingly, NL1 affected GABAergic synapse formation more specifically than NL3, and NL2 affected excitatory synapse density preferentially in pyramidal neurons. In summary, our results demonstrated that different NLs play distinct roles in regulating the development and balance of excitatory and inhibitory synapses in pyramidal neurons and interneurons.


Assuntos
Moléculas de Adesão Celular Neuronais/fisiologia , Interneurônios/fisiologia , Proteínas de Membrana/fisiologia , Proteínas do Tecido Nervoso/fisiologia , Células Piramidais/fisiologia , Sinapses/fisiologia , Animais , Células Cultivadas , Córtex Cerebral/embriologia , Córtex Cerebral/fisiologia , Neurônios GABAérgicos/fisiologia , Isoformas de Proteínas/fisiologia , Ratos Sprague-Dawley
16.
J Neurosci ; 39(8): 1386-1404, 2019 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-30617207

RESUMO

Heparan sulfate (HS) is a cell surface and extracellular matrix carbohydrate extensively modified by differential sulfation. HS interacts physically with canonical fibroblast growth factor (FGF) proteins that signal through the extracellular signal regulated kinase (ERK)/mitogen activated protein kinase (MAPK) pathway. At the embryonic mouse telencephalic midline, FGF/ERK signaling drives astroglial precursor somal translocation from the ventricular zone of the corticoseptal boundary (CSB) to the induseum griseum (IG), producing a focus of Slit2-expressing astroglial guidepost cells essential for interhemispheric corpus callosum (CC) axon navigation. Here, we investigated the cell and molecular function of a specific form of HS sulfation, 2-O HS sulfation catalyzed by the enzyme Hs2st, in midline astroglial development and in regulating FGF protein levels and interaction with HS. Hs2st -/- embryos of either sex exhibit a grossly enlarged IG due to precocious astroglial translocation and conditional Hs2st mutagenesis and ex vivo culture experiments show that Hs2st is not required cell autonomously by CC axons or by the IG astroglial cell lineage, but rather acts non-cell autonomously to suppress the transmission of translocation signals to astroglial precursors. Rescue of the Hs2st -/- astroglial translocation phenotype by pharmacologically inhibiting FGF signaling shows that the normal role of Hs2st is to suppress FGF-mediated astroglial translocation. We demonstrate a selective action of Hs2st on FGF protein by showing that Hs2st (but not Hs6st1) normally suppresses the levels of Fgf17 protein in the CSB region in vivo and use a biochemical assay to show that Hs2st (but not Hs6st1) facilitates a physical interaction between the Fgf17 protein and HS.SIGNIFICANCE STATEMENT We report a novel non-cell-autonomous mechanism regulating cell signaling in developing brain. Using the developing mouse telencephalic midline as an exemplar, we show that the specific sulfation modification of the cell surface and extracellular carbohydrate heparan sulfate (HS) performed by Hs2st suppresses the supply of translocation signals to astroglial precursors by a non-cell-autonomous mechanism. We further show that Hs2st modification selectively facilitates a physical interaction between Fgf17 and HS and suppresses Fgf17 protein levels in vivo, strongly suggesting that Hs2st acts selectively on Fgf17 signaling. HS interacts with many signaling proteins potentially encoding numerous selective interactions important in development and disease, so this class of mechanism may apply more broadly to other biological systems.


Assuntos
Astrócitos/metabolismo , Heparitina Sulfato/metabolismo , Proteínas do Tecido Nervoso/fisiologia , Células-Tronco Neurais/metabolismo , Prosencéfalo/enzimologia , Sulfatos/metabolismo , Sulfotransferases/fisiologia , Animais , Biomarcadores , Linhagem da Célula , Movimento Celular , Córtex Cerebral/citologia , Córtex Cerebral/embriologia , Córtex Cerebral/metabolismo , Fatores de Crescimento de Fibroblastos/metabolismo , Proteínas de Homeodomínio/análise , Camundongos , N-Acetilglucosaminiltransferases/deficiência , N-Acetilglucosaminiltransferases/fisiologia , Prosencéfalo/citologia , Prosencéfalo/embriologia , Sulfotransferases/deficiência , Fatores de Transcrição/análise
17.
Neuron ; 101(4): 707-720.e5, 2019 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-30638744

RESUMO

RNA-binding proteins (RBPs) regulate genetic diversity, but the degree to which they do so in individual cell types in vivo is unknown. We developed NOVA2 cTag-crosslinking and immunoprecipitation (CLIP) to generate functional RBP-RNA maps from different neuronal populations in the mouse brain. Combining cell type datasets from Nova2-cTag and Nova2 conditional knockout mice revealed differential NOVA2 regulatory actions on alternative splicing (AS) on the same transcripts expressed in different neurons. This includes functional differences in transcripts expressed in cortical and cerebellar excitatory versus inhibitory neurons, where we find NOVA2 is required for, respectively, development of laminar structure, motor coordination, and synapse formation. We also find that NOVA2-regulated AS is coupled to NOVA2 regulation of intron retention in hundreds of transcripts, which can sequester the trans-acting splicing factor PTBP2. In summary, cTag-CLIP complements single-cell RNA sequencing (RNA-seq) studies by providing a means for understanding RNA regulation of functional cell diversity.


Assuntos
Processamento Alternativo , Antígenos de Neoplasias/genética , Cerebelo/embriologia , Córtex Cerebral/embriologia , Neurogênese , Neurônios/metabolismo , Proteínas de Ligação a RNA/genética , Animais , Antígenos de Neoplasias/metabolismo , Células Cultivadas , Cerebelo/citologia , Cerebelo/fisiologia , Córtex Cerebral/citologia , Potenciais Pós-Sinápticos Excitadores , Feminino , Potenciais Pós-Sinápticos Inibidores , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Neurônios/citologia , Neurônios/fisiologia , Proteína de Ligação a Regiões Ricas em Polipirimidinas/genética , Proteína de Ligação a Regiões Ricas em Polipirimidinas/metabolismo , Proteínas de Ligação a RNA/metabolismo
18.
J Neurosci ; 39(4): 678-691, 2019 01 23.
Artigo em Inglês | MEDLINE | ID: mdl-30504273

RESUMO

The actin cytoskeleton is crucial for neuronal migration in the mammalian developing cerebral cortex. The adaptor protein Drebrin-like (Dbnl) plays important roles in reorganization of the actin cytoskeleton, dendrite formation, and endocytosis by interacting with F-actin, cobl, and dynamin. Although Dbnl is known to be expressed in the brain, the functions of this molecule during brain development are largely unknown. In this study, to examine the roles of Dbnl in the developing cerebral cortex, we conducted experiments using mice of both sexes with knockdown of Dbnl, effected by in utero electroporation, in the migrating neurons of the embryonic cortex. Time-lapse imaging of the Dbnl-knockdown neurons revealed that the presence of Dbnl is a prerequisite for appropriate formation of processes in the multipolar neurons in the multipolar cell accumulation zone or the deep part of the subventricular zone, and for neuronal polarization and entry into the cortical plate. We found that Dbnl knockdown decreased the amount of N-cadherin protein expressed on the plasma membrane of the cortical neurons. The defect in neuronal migration caused by Dbnl knockdown was rescued by moderate overexpression of N-cadherin and αN-catenin or by transfection of the phospho-mimic form (Y337E, Y347E), but not the phospho-resistant form (Y337F, Y347F), of Dbnl. These results suggest that Dbnl controls neuronal migration, neuronal multipolar morphology, and cell polarity in the developing cerebral cortex via regulating N-cadherin expression.SIGNIFICANCE STATEMENT Disruption of neuronal migration can cause neuronal disorders, such as lissencephaly and subcortical band heterotopia. During cerebral cortical development, the actin cytoskeleton plays a key role in neuronal migration; however, the mechanisms of regulation of neuronal migration by the actin cytoskeleton still remain unclear. Herein, we report that the novel protein Dbnl, an actin-binding protein, controls multiple events during neuronal migration in the developing mouse cerebral cortex. We also showed that this regulation is mediated by phosphorylation of Dbnl at tyrosine residues 337 and 347 and αN-catenin/N-cadherin, suggesting that the Dbnl-αN-catenin/N-cadherin pathway is important for neuronal migration in the developing cortex.


Assuntos
Caderinas/biossíntese , Movimento Celular/fisiologia , Córtex Cerebral/crescimento & desenvolvimento , Córtex Cerebral/fisiologia , Proteínas dos Microfilamentos/fisiologia , Neurônios/fisiologia , Domínios de Homologia de src/fisiologia , Animais , Caderinas/genética , Membrana Celular/metabolismo , Córtex Cerebral/embriologia , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Técnicas de Silenciamento de Genes , Ventrículos Laterais/citologia , Ventrículos Laterais/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos ICR , Proteínas dos Microfilamentos/biossíntese , Proteínas dos Microfilamentos/genética , Neurônios/ultraestrutura , Gravidez , Domínios de Homologia de src/genética
19.
Ultrasound Obstet Gynecol ; 54(2): 199-206, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30381845

RESUMO

OBJECTIVE: To evaluate Sylvian fissure development by assessing Sylvian fissure angles in fetuses with malformation of cortical development (MCD). METHODS: This was a retrospective study of 22 fetuses with MCD. Cases with a stored three-dimensional (3D) brain volume acquired at 18 + 0 to 30 + 6 weeks of gestation at an ultrasound-based research clinic between January 2010 and December 2017 were identified through a database. Of the 22 fetuses, seven had an extracranial abnormality, such as cardiac, renal, gastrointestinal and/or digital anomalies, and five had a minor abnormality such as micrognathia, low-set ears and/or single umbilical artery. To confirm the final clinical diagnosis of brain abnormality, postmortem histological findings or prenatal or postnatal magnetic resonance images were used. For measurement of Sylvian fissure angle, an anterior coronal plane of the fetal brain on transvaginal 3D volume multiplanar imaging was visualized as a single image from the three orthogonal views. The right and left Sylvian fissure angles were measured between a horizontal reference line (0°) and a line drawn along the upper side of the respective Sylvian fissure. The Sylvian fissure angle on both sides was plotted on the graphs of the reference ranges for gestational age in weeks. RESULTS: In 21 (95.5%; 95% CI, 86.8-100.0%) of 22 fetuses with MCD, the Sylvian fissure angle on one or both sides was larger than the 90th percentile of the normal reference. There was one case with apparent focal MCD in the parietal lobe, but the Sylvian fissure angles were normal. A case with apparent unilateral cortical dysplasia and one with apparent unilateral schizencephaly had conspicuous discrepancies between the left and right Sylvian fissure angles. Abnormal genetic test results were obtained in six cases, including four cases with a mutation in a single gene. CONCLUSIONS: This study has shown that the Sylvian fissures, as defined by the Sylvian fissure angle, have delayed development in most MCD cases prior to the diagnosis of the condition. The Sylvian fissure angle may potentially be a strong indicator for the subsequent development of cortical malformation, before the time point at which the gyri and sulci become obvious on the fetal brain surface. Further research is required to validate these findings. © 2018 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of the International Society of Ultrasound in Obstetrics and Gynecology.


Assuntos
Córtex Cerebral/anormalidades , Córtex Cerebral/diagnóstico por imagem , Anormalidades Congênitas/diagnóstico por imagem , Malformações do Desenvolvimento Cortical/diagnóstico por imagem , Autopsia , Encéfalo/anatomia & histologia , Encéfalo/diagnóstico por imagem , Córtex Cerebral/embriologia , Anormalidades Congênitas/genética , Anormalidades Congênitas/patologia , Feminino , Desenvolvimento Fetal , Feto , Idade Gestacional , Humanos , Imageamento Tridimensional , Imagem por Ressonância Magnética/métodos , Malformações do Desenvolvimento Cortical/genética , Malformações do Desenvolvimento Cortical/patologia , Gravidez , Segundo Trimestre da Gravidez , Terceiro Trimestre da Gravidez , Valores de Referência , Estudos Retrospectivos , Ultrassonografia Doppler Transcraniana/métodos , Ultrassonografia Pré-Natal/métodos
20.
Ultrasound Obstet Gynecol ; 54(2): 190-198, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30381851

RESUMO

OBJECTIVE: To (1) evaluate the normal development of the Sylvian fissures in the anterior coronal view of the fetal brain at 18-30 weeks' gestation by transvaginal three-dimensional (3D) ultrasound, (2) develop reference ranges of measurements of the right and left Sylvian fissure angles during normal pregnancy at 18-30 weeks' gestation, and (3) examine intra- and interobserver repeatability of measurements of the right and left Sylvian fissure angles. METHODS: This was a prospective cross-sectional study of 422 women with a singleton pregnancy attending an ultrasound-based research clinic between March and December 2017. The entry criteria for the study were appropriately grown live fetus with no suspected structural and/or chromosomal defects between 18 + 0 and 30 + 6 weeks' gestation. Normal development of the Sylvian fissures was assessed in the anterior coronal plane of the fetal brain using transvaginal 3D volume multiplanar imaging. The coronal view was visualized as a single image from the three orthogonal views. Subsequently, the right and left Sylvian fissure angles were measured between a horizontal reference line (0°) and a line drawn along the upper side of the respective Sylvian fissure. Intra- and interobserver repeatability of the Sylvian fissure angle measurements was assessed by Bland-Altman plots. Reference equations were constructed for right and left Sylvian fissure angles for gestational age (GA) and head circumference (HC) using the Generalized Additive Models for Location Scale and Shape package. RESULTS: In the anterior coronal view of the fetal brain, an inward rotation of the upper portion of the Sylvian fissures was observed during the second and third trimesters of pregnancy. There was a significant negative polynomial association between the Sylvian fissure angles and GA and HC. Both Sylvian fissure angles crossed the reference line (zero), going from positive to negative, at around 25 weeks' gestation or at HC of 22 cm. Z-score difference between the smoothed percentiles of the right and left Sylvian fissure angles indicated that median, 10th and 90th smoothed percentiles were closest and almost the same for the GA-based references between 18 and 28 weeks and for the HC-based references between 14 and 24 cm. The intraclass correlation coefficient of the right and left Sylvian fissure angle measurements between the two sonographers was excellent at 0.993 (95% CI, 0.988-0.996) and 0.991 (95% CI, 0.985-0.995), respectively. On Bland-Altman analysis, the mean difference between the two sonographers in right Sylvian fissure angle measurement was 0.4° (95% CI, -10.2 to 10.1°) and in left Sylvian fissure angle it was 1.0° (95% CI, -9.6 to 11.6°). CONCLUSIONS: Assessment of the Sylvian fissure angles is highly reproducible. Sylvian fissure angle reference charts can serve as a screening tool for malformations of cortical development, guiding subsequent follow-up and referral for fetal brain magnetic resonance imaging and/or assessment by an expert neurosonologist. Copyright © 2018 ISUOG. Published by John Wiley & Sons Ltd.


Assuntos
Córtex Cerebral/anatomia & histologia , Córtex Cerebral/diagnóstico por imagem , Imageamento Tridimensional/instrumentação , Adulto , Córtex Cerebral/embriologia , Estudos Transversais , Feminino , Desenvolvimento Fetal/fisiologia , Idade Gestacional , Humanos , Variações Dependentes do Observador , Gravidez , Segundo Trimestre da Gravidez , Terceiro Trimestre da Gravidez , Estudos Prospectivos , Valores de Referência , Ultrassonografia , Ultrassonografia Pré-Natal/métodos
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