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1.
J Affect Disord ; 245: 1079-1088, 2019 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-30699850

RESUMO

BACKGROUND: Genetic and physiological studies have implicated the striatum in bipolar disorder (BD). Although Glycogen synthase kinase 3 beta (GSK3ß) has been suggested to play a role in the pathophysiology of BD since it is inhibited by lithium, it remains unknown how GSK3ß activity might be involved. Therefore we examined the functional roles of GSK3ß and one of its substrates, CRMP2, within the striatum. METHODS: Using CRISPR-Cas9 system, we specifically ablated GSK3ß in the striatal neurons in vivo and in vitro. Sholl analysis was performed for the structural studies of medium spiny neurons (MSNs) and amphetamine-induced hyperlocomotion was measured to investigate the effects of gene ablations on the mania-like symptom of BD. RESULTS: GSK3ß deficiency in cultured neurons and in neurons of adult mouse brain caused opposite patterns of neurite changes. Furthermore, specific knockout of GSK3ß in the MSNs of the indirect pathway significantly suppressed amphetamine-induced hyperlocomotion. We demonstrated that these phenotypes of GSK3ß ablation were mediated by CRMP2, a major substrate of GSK3ß. LIMITATIONS: Amphetamine-induced hyperlocomotion only partially recapitulate the symptoms of BD. It requires further study to examine whether abnormality in GSK3ß or CRMP2 is also involved in depression phase of BD. Additionally, we could not confirm whether the behavioral changes observed in GSK3ß-ablated mice were indeed caused by the cellular structural changes observed in the striatal neurons. CONCLUSION: Our results demonstrate that GSK3ß and its substrate CRMP2 critically regulate the neurite structure of MSNs and their functions specifically within the indirect pathway of the basal ganglia network play a critical role in manifesting mania-like behavior of BD. Moreover, our data also suggest lithium may exert its effect on BD through a GSK3ß-independent mechanism, in addition to the GSK3ß inhibition-mediated mechanism.


Assuntos
Transtorno Bipolar/patologia , Corpo Estriado/patologia , Dendritos/ultraestrutura , Glicogênio Sintase Quinase 3 beta/deficiência , Locomoção/genética , Anfetamina/farmacologia , Animais , Transtorno Bipolar/tratamento farmacológico , Células Cultivadas , Corpo Estriado/metabolismo , Depressão , Modelos Animais de Doenças , Glicogênio Sintase Quinase 3 beta/fisiologia , Humanos , Lítio/farmacologia , Locomoção/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neostriado/patologia , Neurônios/citologia
2.
Neuron ; 101(2): 204-206, 2019 01 16.
Artigo em Inglês | MEDLINE | ID: mdl-30653934

RESUMO

Two papers in Cell uncover reciprocal crosstalk of local translation and mitochondria in neurons. Rangaraju et al. (2019) observe tethered compartments of stable mitochondria in dendrites that provide a local energy supply for mRNA translation at synapses. Cioni et al. (2019) report a novel association of axonal RNA granules with Rab7a-late endosomes that provides a platform for local translation supporting mitochondria.


Assuntos
Axônios/metabolismo , Dendritos/metabolismo , Mitocôndrias/fisiologia , Neurônios/citologia , Biossíntese de Proteínas/fisiologia , Animais , Transporte Axonal , Axônios/ultraestrutura , Plasticidade Celular , Dendritos/ultraestrutura , Neurônios/ultraestrutura , RNA Mensageiro/metabolismo
3.
Methods Mol Biol ; 1880: 243-256, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30610702

RESUMO

Autophagy is an essential degradative pathway that maintains neuronal homeostasis and prevents axon degeneration. However, the mechanisms of autophagy in neurons are only beginning to be understood. To address this fundamental gap in knowledge, we have established several key methodologies for live-cell imaging and quantitative analysis of autophagy in primary hippocampal neurons. Using these methods, we have defined compartment-specific dynamics of autophagy in real-time under basal versus stress conditions. For example, we have characterized autophagosome biogenesis in the distal axon and subsequent retrograde transport to the soma for degradation. Autophagosomes are also generated locally within the soma. In contrast to the axon, the majority of autophagosomes in dendrites are stationary, while some exhibit bidirectional movement. These studies establish an initial road map for autophagosome dynamics in each compartment of the neuron and set the stage for a more detailed understanding of neuronal autophagy in stress and disease.


Assuntos
Autofagossomos/ultraestrutura , Autofagia , Microscopia de Fluorescência/métodos , Neurônios/citologia , Imagem Óptica/métodos , Animais , Axônios/ultraestrutura , Técnicas de Cultura de Células/métodos , Células Cultivadas , Dendritos/ultraestrutura , Hipocampo/citologia , Camundongos , Camundongos Transgênicos , Microscopia Confocal/métodos , Neuroglia/citologia , Neurônios/ultraestrutura
4.
Arthropod Struct Dev ; 47(5): 482-497, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30120986

RESUMO

In many acoustic insects, mate finding and mate choice are primarily based on acoustic signals. In several species with high-intensity calling songs, such as the studied katydid Mecopoda sp., males exhibit an increase in their thoracic temperature during singing, which is linearly correlated with the amount of energy invested in song production. If this increased body temperature is used by females as an additional cue to assess the male's quality during mate choice, as has been recently hypothesized ("hot-male" hypothesis), thermosensory structures would be required to evaluate this cue. In the present study, therefore, we investigated the ultrastructure and physiology of thermosensitive sensilla coeloconica on the antennal flagella of Mecopoda sp. using a combination of electron microscopy and electrophysiological recording techniques. We could identify three distinct types of sensilla coeloconica based on differences in the number and branching pattern of their dendrites. Physiological recordings revealed the innervation by antagonistically responding thermoreceptors (cold and warm) and bimodal hygro-/thermoreceptors (moist or dry) in various combinations. Our findings indicate that Mecopoda sp. females are capable of detecting a singing male from distances of at least several centimetres solely by assessing thermal cues.


Assuntos
Ortópteros/ultraestrutura , Sensilas/fisiologia , Animais , Antenas de Artrópodes/fisiologia , Antenas de Artrópodes/ultraestrutura , Dendritos/ultraestrutura , Eletrofisiologia , Feminino , Umidade , Masculino , Microscopia Eletrônica de Transmissão , Ortópteros/fisiologia , Sensilas/ultraestrutura , Temperatura Ambiente , Termorreceptores/fisiologia
5.
Neurochem Res ; 43(8): 1587-1598, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29948727

RESUMO

Arachidonic acid and docosahexaenoic acid (DHA) released by the action of phospholipases A2 (PLA2) on membrane phospholipids may be metabolized by lipoxygenases to the anti-inflammatory mediators lipoxin A4 (LXA4) and resolvin D1 (RvD1), and these can bind to a common receptor, formyl-peptide receptor 2 (FPR2). The contribution of this receptor to axonal or dendritic outgrowth is unknown. The present study was carried out to elucidate the distribution of FPR2 in the rat CNS and its role in outgrowth of neuronal processes. FPR2 mRNA expression was greatest in the brainstem, followed by the spinal cord, thalamus/hypothalamus, cerebral neocortex, hippocampus, cerebellum and striatum. The brainstem and spinal cord also contained high levels of FPR2 protein. The cerebral neocortex was moderately immunolabelled for FPR2, with staining mostly present as puncta in the neuropil. Dentate granule neurons and their axons (mossy fibres) in the hippocampus were very densely labelled. The cerebellar cortex was lightly stained, but the deep cerebellar nuclei, inferior olivary nucleus, vestibular nuclei, spinal trigeminal nucleus and dorsal horn of the spinal cord were densely labelled. Electron microscopy of the prefrontal cortex showed FPR2 immunolabel mostly in immature axon terminals or 'pre-terminals', that did not form synapses with dendrites. Treatment of primary hippocampal neurons with the FPR2 inhibitors, PBP10 or WRW4, resulted in reduced lengths of axons and dendrites. The CNS distribution of FPR2 suggests important functions in learning and memory, balance and nociception. This might be due to an effect of FPR2 in mediating arachidonic acid/LXA4 or DHA/RvD1-induced axonal or dendritic outgrowth.


Assuntos
Axônios/metabolismo , Encéfalo/metabolismo , Dendritos/metabolismo , Receptores de Lipoxinas/biossíntese , Medula Espinal/metabolismo , Animais , Axônios/química , Axônios/ultraestrutura , Encéfalo/ultraestrutura , Química Encefálica/fisiologia , Sobrevivência Celular/fisiologia , Sistema Nervoso Central/química , Sistema Nervoso Central/metabolismo , Sistema Nervoso Central/ultraestrutura , Dendritos/química , Dendritos/ultraestrutura , Masculino , Ratos , Ratos Wistar , Receptores de Lipoxinas/análise , Medula Espinal/química , Medula Espinal/ultraestrutura
6.
Biochem Biophys Res Commun ; 503(1): 123-130, 2018 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-29864422

RESUMO

Dendrite morphogenesis is a complex but well-orchestrated process. Various studies reported the involvement of alteration in dendrite morphology in different brain disorders, including neuropsychiatric disorders. Initially, ßB2-crystallin (gene symbol: Crybb2/CRYBB2) has been described as a structural protein of the ocular lens. Mutations of the corresponding gene, Crybb2, lead to cataract. Recent studies in mice suggested that mutations in Crybb2 cause alterations in hippocampal morphology and function, albeit its function in hippocampal neuron development remained elusive. In the current study, we found that Crybb2 contributes to dendritogenesis in vitro and in vivo. Furthermore, screening of previous data on differential expression-arrays, we found Tmsb4X up-regulated in Crybb2 mutants mouse brain. Additionally, Tmsb4X was co-expressed with Crybb2 at actin-enriched cell ruffles. Over-expression of Tmsb4X in cultured hippocampal neurons inhibited dendritogenesis, which phenocopied Crybb2 knock-down. The current study uncovers a new function of Crybb2 in brain development, especially in dendritogenesis, and the possible interplay partner Tmsb4X involved in this process.


Assuntos
Dendritos/genética , Timosina/genética , Cadeia B de beta-Cristalina/genética , Actinas/metabolismo , Animais , Células Cultivadas , Dendritos/metabolismo , Dendritos/ultraestrutura , Técnicas de Silenciamento de Genes , Hipocampo/citologia , Hipocampo/metabolismo , Camundongos , Camundongos Mutantes , Mutação , Neurogênese/genética , Neurogênese/fisiologia , Neurônios/citologia , Neurônios/metabolismo , RNA Interferente Pequeno/genética , Timosina/metabolismo , Regulação para Cima , Cadeia B de beta-Cristalina/antagonistas & inibidores , Cadeia B de beta-Cristalina/metabolismo
7.
Synapse ; 72(6): e22024, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29323756

RESUMO

Myelin mutant taiep rats show a progressive demyelination in the central nervous system due to an abnormal accumulation of microtubules in the cytoplasm and the processes on their oligodendrocytes. Demyelination is associated with electrophysiological alterations and the mutant had a progressive astrocytosis. The illness is associated with change in cytokine levels and in the expression of different nitric oxide synthase and concomitantly lipoperoxidation in several areas of the brain. However, until now there has been no detailed anatomical analysis of neurons in this mutant. The aim of this study was to analyze the dendritic morphology in the hippocampus using Golgi-Cox staining and spatial memory through Morris water maze test in young adult (3 months old) taiep rats and compare them with normal Sprague-Dawley. Our results showed that taiep rats have altered dendritic tree morphology in pyramidal neurons in the CA1 field of the hippocampus, but not in the CA3 region. These morphological changes did not produce a concomitant deficit in spatial memory acquisition or recall at this early stage of the disease. Our results suggest that impairment of dendritic morphology in the CA1 field of the hippocampus is a landmark of the pathology of this progressive multiple sclerosis model.


Assuntos
Doenças Autoimunes Desmielinizantes do Sistema Nervoso Central/patologia , Dendritos/patologia , Hipocampo/patologia , Células Piramidais/patologia , Análise de Variância , Animais , Doenças Autoimunes Desmielinizantes do Sistema Nervoso Central/genética , Doenças Autoimunes Desmielinizantes do Sistema Nervoso Central/fisiopatologia , Dendritos/ultraestrutura , Modelos Animais de Doenças , Masculino , Aprendizagem em Labirinto , Transtornos da Memória/fisiopatologia , Proteínas Associadas aos Microtúbulos/genética , Mutação/genética , Células Piramidais/ultraestrutura , Ratos , Ratos Mutantes , Ratos Sprague-Dawley , Coloração pela Prata , Aprendizagem Espacial/fisiologia
8.
Methods Mol Biol ; 1695: 161-170, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29190026

RESUMO

Labeling of cellular structures is of fundamental importance in the investigation of diseases of the central nervous system. Biolistic labeling of retinal ganglion cells (RGCs) allows visualization of dendritic and synaptic structures of RGCs in retinal explants from animal models of experimental glaucoma. This technique sparsely labels RGCs, and, due to the stochastic nature of the particle delivery, all RGC types can be potentially observed in the labeled tissue. Quantification of dendritic and synaptic properties permits examination of the specific alterations to RGC morphology at different stages of degeneration, such as dendritic shrinkage and excitatory synapse loss.


Assuntos
Biolística/métodos , Glaucoma/metabolismo , Células Ganglionares da Retina/citologia , Animais , Dendritos/metabolismo , Dendritos/ultraestrutura , Modelos Animais de Doenças , Glaucoma/diagnóstico por imagem , Camundongos , Células Ganglionares da Retina/metabolismo , Sinapses/metabolismo , Sinapses/ultraestrutura
9.
J Comp Neurol ; 526(3): 496-536, 2018 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-29088505

RESUMO

Gigantopyramidal neurons, referred to as Betz cells in primates, are characterized by large somata and extensive basilar dendrites. Although there have been morphological descriptions and drawings of gigantopyramidal neurons in a limited number of species, quantitative investigations have typically been limited to measures of soma size. The current study thus employed two separate analytical approaches: a morphological investigation using the Golgi technique to provide qualitative and quantitative somatodendritic measures of gigantopyramidal neurons across 19 mammalian species from 7 orders; and unbiased stereology to compare the soma volume of layer V pyramidal and gigantopyramidal neurons in primary motor cortex between 11 carnivore and 9 primate species. Of the 617 neurons traced in the morphological analysis, 181 were gigantopyramidal neurons, with deep (primarily layer V) pyramidal (n = 203) and superficial (primarily layer III) pyramidal (n = 233) neurons quantified for comparative purposes. Qualitatively, dendritic morphology varied considerably across species, with some (sub)orders (e.g., artiodactyls, perissodactyls, feliforms) exhibiting bifurcating, V-shaped apical dendrites. Basilar dendrites exhibited idiosyncratic geometry across and within taxonomic groups. Quantitatively, most dendritic measures were significantly greater in gigantopyramidal neurons than in superficial and deep pyramidal neurons. Cluster analyses revealed that most taxonomic groups could be discriminated based on somatodendritic morphology for both superficial and gigantopyramidal neurons. Finally, in agreement with Brodmann, gigantopyramidal neurons in both the morphological and stereological analyses were larger in feliforms (especially in the Panthera species) than in other (sub)orders, possibly due to specializations in muscle fiber composition and musculoskeletal systems.


Assuntos
Evolução Biológica , Córtex Motor/citologia , Células Piramidais/ultraestrutura , Animais , Contagem de Células , Dendritos/ultraestrutura , Feminino , Humanos , Masculino , Mamíferos/anatomia & histologia , Células Piramidais/classificação , Células Piramidais/citologia , Coloração pela Prata , Especificidade da Espécie
10.
Science ; 358(6363): 615-623, 2017 11 03.
Artigo em Inglês | MEDLINE | ID: mdl-29097543

RESUMO

Insects and mammals share similarities of neural organization underlying the perception of odors, taste, vision, sound, and gravity. We observed that insect somatosensation also corresponds to that of mammals. In Drosophila, the projections of all the somatosensory neuron types to the insect's equivalent of the spinal cord segregated into modality-specific layers comparable to those in mammals. Some sensory neurons innervate the ventral brain directly to form modality-specific and topological somatosensory maps. Ascending interneurons with dendrites in matching layers of the nerve cord send axons that converge to respective brain regions. Pathways arising from leg somatosensory neurons encode distinct qualities of leg movement information and play different roles in ground detection. Establishment of the ground pattern and genetic tools for neuronal manipulation should provide the basis for elucidating the mechanisms underlying somatosensation.


Assuntos
Drosophila melanogaster/fisiologia , Células Receptoras Sensoriais/fisiologia , Células Receptoras Sensoriais/ultraestrutura , Córtex Somatossensorial/citologia , Córtex Somatossensorial/fisiologia , Animais , Axônios/fisiologia , Axônios/ultraestrutura , Comportamento Animal/fisiologia , Dendritos/fisiologia , Dendritos/ultraestrutura , Interneurônios/citologia , Interneurônios/fisiologia , Locomoção , Atividade Motora , Vias Neurais/citologia , Vias Neurais/fisiologia , Medula Espinal/fisiologia , Sinapses/fisiologia , Sinapses/ultraestrutura
11.
J Neurosci ; 37(50): 12153-12166, 2017 Dec 13.
Artigo em Inglês | MEDLINE | ID: mdl-29118107

RESUMO

Combinatorial expansion by the cerebellar granule cell layer (GCL) is fundamental to theories of cerebellar contributions to motor control and learning. Granule cells (GrCs) sample approximately four mossy fiber inputs and are thought to form a combinatorial code useful for pattern separation and learning. We constructed a spatially realistic model of the cerebellar GCL and examined how GCL architecture contributes to GrC combinatorial diversity. We found that GrC combinatorial diversity saturates quickly as mossy fiber input diversity increases, and that this saturation is in part a consequence of short dendrites, which limit access to diverse inputs and favor dense sampling of local inputs. This local sampling also produced GrCs that were combinatorially redundant, even when input diversity was extremely high. In addition, we found that mossy fiber clustering, which is a common anatomical pattern, also led to increased redundancy of GrC input combinations. We related this redundancy to hypothesized roles of temporal expansion of GrC information encoding in service of learned timing, and we show that GCL architecture produces GrC populations that support both temporal and combinatorial expansion. Finally, we used novel anatomical measurements from mice of either sex to inform modeling of sparse and filopodia-bearing mossy fibers, finding that these circuit features uniquely contribute to enhancing GrC diversification and redundancy. Our results complement information theoretic studies of granule layer structure and provide insight into the contributions of granule layer anatomical features to afferent mixing.SIGNIFICANCE STATEMENT Cerebellar granule cells are among the simplest neurons, with tiny somata and, on average, just four dendrites. These characteristics, along with their dense organization, inspired influential theoretical work on the granule cell layer as a combinatorial expander, where each granule cell represents a unique combination of inputs. Despite the centrality of these theories to cerebellar physiology, the degree of expansion supported by anatomically realistic patterns of inputs is unknown. Using modeling and anatomy, we show that realistic input patterns constrain combinatorial diversity by producing redundant combinations, which nevertheless could support temporal diversification of like combinations, suitable for learned timing. Our study suggests a neural substrate for producing high levels of both combinatorial and temporal diversity in the granule cell layer.


Assuntos
Córtex Cerebelar/citologia , Conectoma , Dendritos/fisiologia , Modelos Neurológicos , Fibras Nervosas/fisiologia , Pseudópodes/fisiologia , Vias Aferentes/fisiologia , Vias Aferentes/ultraestrutura , Animais , Proteínas de Bactérias/análise , Simulação por Computador , Conectoma/métodos , Dendritos/ultraestrutura , Dependovirus , Feminino , Genes Reporter , Vetores Genéticos , Proteínas Luminescentes/análise , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fibras Nervosas/ultraestrutura , Pseudópodes/ultraestrutura , Sinapses/fisiologia
12.
J Neurosci ; 37(50): 12141-12152, 2017 Dec 13.
Artigo em Inglês | MEDLINE | ID: mdl-29114075

RESUMO

In higher mammals, orientation tuning of neurons is organized into a quasi-periodic pattern in the primary visual cortex. Our previous model studies suggested that the topography of cortical orientation maps may originate from moiré interference of ON and OFF retinal ganglion cell (RGC) mosaics, but did not account for how the consistent spatial period of maps could be achieved. Here we address this issue with two crucial findings on the development of RGC mosaics: first, homotypic local repulsion between RGCs can develop a long-range hexagonal periodicity. Second, heterotypic interaction restrains the alignment of ON and OFF mosaics, and generates a periodic interference pattern map with consistent spatial frequency. To validate our model, we quantitatively analyzed the RGC mosaics in cat data, and confirmed that the observed retinal mosaics showed evidence of heterotypic interactions, contrary to the previous view that ON and OFF mosaics are developed independently.SIGNIFICANCE STATEMENT Orientation map is one of the most studied functional maps in the brain, but it has remained unanswered how the consistent spatial periodicity of maps could be developed. In the current study, we address this issue with our developmental model for the retinal origin of orientation map. We showed that local repulsive interactions between retinal ganglion cells (RGCs) can develop a hexagonal periodicity in the RGC mosaics and restrict the alignment between ON and OFF mosaics, so that they generate a periodic pattern with consistent spatial frequency for both the RGC mosaics and the cortical orientation maps. Our results demonstrate that the organization of functional maps in visual cortex, including its structural consistency, may be constrained by a retinal blueprint.


Assuntos
Simulação por Computador , Conectoma , Modelos Neurológicos , Percepção de Movimento/fisiologia , Células Ganglionares da Retina/citologia , Córtex Visual/fisiologia , Vias Aferentes/fisiologia , Vias Aferentes/ultraestrutura , Animais , Gatos , Comunicação Celular , Dendritos/fisiologia , Dendritos/ultraestrutura , Corpos Geniculados/fisiologia , Corpos Geniculados/ultraestrutura , Mamíferos/anatomia & histologia , Estimulação Luminosa , Células Ganglionares da Retina/fisiologia , Células Ganglionares da Retina/efeitos da radiação , Núcleos Talâmicos/fisiologia , Núcleos Talâmicos/ultraestrutura , Vias Visuais/fisiologia , Vias Visuais/ultraestrutura
13.
Cereb Cortex ; 27(11): 5353-5368, 2017 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-28968722

RESUMO

Neocortical vasoactive intestinal polypeptide (VIP) expressing cells are a diverse subpopulation of GABAergic interneurons issuing distinct axonal projections. They are known to inhibit other types of interneurons as well as excitatory principal neurons and possess a disinhibitory net effect in cortical circuits. In order to elucidate their targeting specificity, the output connectivity of VIP interneurons was studied at the subcellular level in barrel cortex of interneuron-specific Cre-driver mice, using pre- and postembedding electron microscopy. Systematically sampling VIP boutons across all layers, we found a substantial proportion of the innervated subcellular structures were dendrites (80%), with somata (13%), and spines (7%) being much less targeted. In layer VI, a high proportion of axosomatic synapses was found (39%). GABA-immunopositive ratio was quantified among the targets using statistically validated thresholds: only 37% of the dendrites, 7% of the spines, and 26% of the somata showed above-threshold immunogold labeling. For the main target structure "dendrite", a higher proportion of GABAergic subcellular profiles existed in deep than in superficial layers. In conclusion, VIP interneurons innervate non-GABAergic excitatory neurons and interneurons at their subcellular domains with layer-dependent specificity. This suggests a diverse output of VIP interneurons, which predicts multiple functionality in cortical circuitry beyond disinhibition.


Assuntos
Neurônios/metabolismo , Neurônios/ultraestrutura , Terminações Pré-Sinápticas/metabolismo , Córtex Somatossensorial/metabolismo , Córtex Somatossensorial/ultraestrutura , Peptídeo Intestinal Vasoativo/metabolismo , Animais , Dendritos/metabolismo , Dendritos/ultraestrutura , Imuno-Histoquímica , Masculino , Camundongos Transgênicos , Microscopia Imunoeletrônica , Microtomia , Vias Neurais/metabolismo , Vias Neurais/ultraestrutura , Terminações Pré-Sinápticas/ultraestrutura , Ácido gama-Aminobutírico/metabolismo
14.
Cell Rep ; 21(1): 97-109, 2017 Oct 03.
Artigo em Inglês | MEDLINE | ID: mdl-28978487

RESUMO

Mutations in the Golgi SNARE (SNAP [soluble NSF attachment protein] receptor) protein Membrin (encoded by the GOSR2 gene) cause progressive myoclonus epilepsy (PME). Membrin is a ubiquitous and essential protein mediating ER-to-Golgi membrane fusion. Thus, it is unclear how mutations in Membrin result in a disorder restricted to the nervous system. Here, we use a multi-layered strategy to elucidate the consequences of Membrin mutations from protein to neuron. We show that the pathogenic mutations cause partial reductions in SNARE-mediated membrane fusion. Importantly, these alterations were sufficient to profoundly impair dendritic growth in Drosophila models of GOSR2-PME. Furthermore, we show that Membrin mutations cause fragmentation of the presynaptic cytoskeleton coupled with transsynaptic instability and hyperactive neurotransmission. Our study highlights how dendritic growth is vulnerable even to subtle secretory pathway deficits, uncovers a role for Membrin in synaptic function, and provides a comprehensive explanatory basis for genotype-phenotype relationships in GOSR2-PME.


Assuntos
Dendritos/metabolismo , Mutação , Epilepsias Mioclônicas Progressivas/genética , Proteínas Qb-SNARE/genética , Via Secretória/genética , Sinapses/metabolismo , Animais , Dendritos/ultraestrutura , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Feminino , Fibroblastos/citologia , Fibroblastos/metabolismo , Expressão Gênica , Estudos de Associação Genética , Complexo de Golgi/metabolismo , Humanos , Masculino , Fusão de Membrana , Epilepsias Mioclônicas Progressivas/metabolismo , Epilepsias Mioclônicas Progressivas/patologia , Fenótipo , Cultura Primária de Células , Proteínas Qb-SNARE/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Sinapses/patologia , Adulto Jovem
15.
J Neurosci ; 37(41): 9889-9900, 2017 Oct 11.
Artigo em Inglês | MEDLINE | ID: mdl-28899920

RESUMO

Neurotransmission plays an essential role in neural circuit formation in the central nervous system (CNS). Although neurotransmission has been recently clarified as a key modulator of retinal circuit development, the roles of individual synaptic transmissions are not yet fully understood. In the current study, we investigated the role of neurotransmission from photoreceptor cells to ON bipolar cells in development using mutant mouse lines of both sexes in which this transmission is abrogated. We found that deletion of the ON bipolar cation channel TRPM1 results in the abnormal contraction of rod bipolar terminals and a decreased number of their synaptic connections with amacrine cells. In contrast, these histological alterations were not caused by a disruption of total glutamate transmission due to loss of the ON bipolar glutamate receptor mGluR6 or the photoreceptor glutamate transporter VGluT1. In addition, TRPM1 deficiency led to the reduction of total dendritic length, branch numbers, and cell body size in AII amacrine cells. Activated Goα, known to close the TRPM1 channel, interacted with TRPM1 and induced the contraction of rod bipolar terminals. Furthermore, overexpression of Channelrhodopsin-2 partially rescued rod bipolar cell development in the TRPM1-/- retina, whereas the rescue effect by a constitutively closed form of TRPM1 was lower than that by the native form. Our results suggest that TRPM1 channel opening is essential for rod bipolar pathway establishment in development.SIGNIFICANCE STATEMENT Neurotransmission has been recognized recently as a key modulator of retinal circuit development in the CNS. However, the roles of individual synaptic transmissions are not yet fully understood. In the current study, we focused on neurotransmission between rod photoreceptor cells and rod bipolar cells in the retina. We used genetically modified mouse models which abrogate each step of neurotransmission: presynaptic glutamate release, postsynaptic glutamate reception, or transduction channel function. We found that the TRPM1 transduction channel is required for the development of rod bipolar cells and their synaptic formation with subsequent neurons, independently of glutamate transmission. This study advances our understanding of neurotransmission-mediated retinal circuit refinement.


Assuntos
Células Amácrinas/fisiologia , Retina/crescimento & desenvolvimento , Células Bipolares da Retina/fisiologia , Células Fotorreceptoras Retinianas Bastonetes/fisiologia , Canais de Cátion TRPM/fisiologia , Vias Visuais/crescimento & desenvolvimento , Vias Visuais/fisiologia , Animais , Channelrhodopsins , Dendritos/fisiologia , Dendritos/ultraestrutura , Feminino , Ácido Glutâmico/fisiologia , Técnicas In Vitro , Masculino , Camundongos , Camundongos Knockout , Técnicas de Patch-Clamp , Retina/citologia , Transmissão Sináptica/fisiologia , Canais de Cátion TRPM/genética , Proteína Vesicular 1 de Transporte de Glutamato/biossíntese , Proteína Vesicular 1 de Transporte de Glutamato/genética
16.
J Comp Neurol ; 525(18): 3951-3961, 2017 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-28857161

RESUMO

Functional deficits in sensory systems are commonly noted in neurodevelopmental disorders, such as the Rett syndrome (RTT). Defects in methyl CpG binding protein gene (MECP2) largely accounts for RTT. Manipulations of the Mecp2 gene in mice provide useful models to probe into various aspects of brain development associated with the RTT. In this study, we focused on the somatosensory cortical phenotype in the Bird mouse model of RTT. We used voltage-sensitive dye imaging to evaluate whisker sensory evoked activity in the barrel cortex of mice. We coupled this functional assay with morphological analyses in postnatal mice and investigated the dendritic differentiation of barrel neurons and individual thalamocortical axon (TCA) arbors that synapse with them. We show that in Mecp2-deficient male mice, whisker-evoked activity is roughly topographic but weak in the barrel cortex. At the morphological level, we find that TCA arbors fail to develop into discrete, concentrated patches in barrel hollows, and the complexity of the dendritic branches in layer IV spiny stellate neurons is reduced. Collectively, our results indicate significant structural and functional impairments in the barrel cortex of the Bird mouse line, a popular animal model for the RTT. Such structural and functional anomalies in the primary somatosensory cortex may underlie orofacial tactile sensitivity issues and sensorimotor stereotypies characteristic of RTT.


Assuntos
Proteína 2 de Ligação a Metil-CpG/deficiência , Síndrome de Rett/genética , Síndrome de Rett/patologia , Córtex Somatossensorial/patologia , Vias Aferentes/fisiologia , Animais , Carbocianinas/metabolismo , Dendritos/patologia , Dendritos/ultraestrutura , Modelos Animais de Doenças , Masculino , Proteína 2 de Ligação a Metil-CpG/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neurônios/metabolismo , Neurônios/ultraestrutura , Coloração pela Prata , Córtex Somatossensorial/citologia , Vibrissas/inervação , Imagens com Corantes Sensíveis à Voltagem
17.
J Cell Sci ; 130(21): 3663-3675, 2017 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-28935671

RESUMO

Central nervous system (CNS) axons lose their intrinsic ability to regenerate upon maturity, whereas peripheral nervous system (PNS) axons do not. A key difference between these neuronal types is their ability to transport integrins into axons. Integrins can mediate PNS regeneration, but are excluded from adult CNS axons along with their Rab11 carriers. We reasoned that exclusion of the contents of Rab11 vesicles including integrins might contribute to the intrinsic inability of CNS neurons to regenerate, and investigated this by performing laser axotomy. We identify a novel regulator of selective axon transport and regeneration, the ARF6 guanine-nucleotide-exchange factor (GEF) EFA6 (also known as PSD). EFA6 exerts its effects from a location within the axon initial segment (AIS). EFA6 does not localise at the AIS in dorsal root ganglion (DRG) axons, and in these neurons, ARF6 activation is counteracted by an ARF GTPase-activating protein (GAP), which is absent from the CNS, ACAP1. Depleting EFA6 from cortical neurons permits endosomal integrin transport and enhances regeneration, whereas overexpressing EFA6 prevents DRG regeneration. Our results demonstrate that ARF6 is an intrinsic regulator of regenerative capacity, implicating EFA6 as a focal molecule linking the AIS, signalling and transport.This article has an associated First Person interview with the first author of the paper.


Assuntos
Segmento Inicial do Axônio/metabolismo , Transporte Axonal/genética , Córtex Cerebral/metabolismo , Dendritos/metabolismo , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Cadeias alfa de Integrinas/metabolismo , Neurônios/metabolismo , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Animais , Segmento Inicial do Axônio/ultraestrutura , Córtex Cerebral/ultraestrutura , Dendritos/ultraestrutura , Embrião de Mamíferos , Proteínas Ativadoras de GTPase/genética , Proteínas Ativadoras de GTPase/metabolismo , Gânglios Espinais/metabolismo , Gânglios Espinais/ultraestrutura , Regulação da Expressão Gênica no Desenvolvimento , Fatores de Troca do Nucleotídeo Guanina/antagonistas & inibidores , Fatores de Troca do Nucleotídeo Guanina/genética , Cadeias alfa de Integrinas/genética , Masculino , Microtúbulos , Neurônios/ultraestrutura , Cultura Primária de Células , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais , Proteínas rab de Ligação ao GTP/genética , Proteínas rab de Ligação ao GTP/metabolismo
18.
Nature ; 549(7673): 469-475, 2017 09 28.
Artigo em Inglês | MEDLINE | ID: mdl-28959971

RESUMO

Research on neuronal connectivity in the cerebral cortex has focused on the existence and strength of synapses between neurons, and their location on the cell bodies and dendrites of postsynaptic neurons. The synaptic architecture of individual presynaptic axonal trees, however, remains largely unknown. Here we used dense reconstructions from three-dimensional electron microscopy in rats to study the synaptic organization of local presynaptic axons in layer 2 of the medial entorhinal cortex, the site of grid-like spatial representations. We observe path-length-dependent axonal synapse sorting, such that axons of excitatory neurons sequentially target inhibitory neurons followed by excitatory neurons. Connectivity analysis revealed a cellular feedforward inhibition circuit involving wide, myelinated inhibitory axons and dendritic synapse clustering. Simulations show that this high-precision circuit can control the propagation of synchronized activity in the medial entorhinal cortex, which is known for temporally precise discharges.


Assuntos
Axônios/fisiologia , Córtex Entorrinal/citologia , Córtex Entorrinal/fisiologia , Vias Neurais/citologia , Vias Neurais/fisiologia , Sinapses/fisiologia , Animais , Axônios/ultraestrutura , Conectoma , Dendritos/fisiologia , Dendritos/ultraestrutura , Córtex Entorrinal/ultraestrutura , Potenciais Pós-Sinápticos Excitadores , Imagem Tridimensional , Interneurônios/fisiologia , Masculino , Microscopia Eletrônica , Modelos Neurológicos , Inibição Neural/fisiologia , Vias Neurais/ultraestrutura , Ratos , Sinapses/ultraestrutura
19.
CNS Neurosci Ther ; 23(9): 748-758, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28786172

RESUMO

INTRODUCTION: Apolipoprotein E4 (APOE4) is a major genetic risk factor for late-onset sporadic Alzheimer disease. Emerging evidence demonstrates a hippocampus-associated learning and memory deficit in aged APOE4 human carriers and also in aged mice carrying human APOE4 gene. This suggests that either exogenous APOE4 or endogenous APOE4 alters the cognitive profile and hippocampal structure and function. However, little is known regarding how Apoe4 modulates hippocampal dendritic morphology, synaptic function, and neural network activity in young mice. AIM: In this study, we compared hippocampal dendritic and spine morphology and synaptic function of young (4 months) mice with transgenic expression of the human APOE4 and APOE3 genes. METHODS: Hippocampal dendritic and spine morphology and synaptic function were assessed by neuronal imaging and electrophysiological approaches. RESULTS: Morphology results showed that shortened dendritic length and reduced spine density occurred at hippocampal CA1 neurons in Apoe4 mice compared to Apoe3 mice. Electrophysiological results demonstrated that in the hippocampal CA3-CA1 synapses of young Apoe4 mice, basic synaptic transmission, and paired-pulse facilitation were enhanced but long-term potentiation and carbachol-induced hippocampal theta oscillations were impaired compared to young Apoe3 mice. However, both Apoe genotypes responded similarly to persistent stimulations (4, 10, and 40 Hz for 4 seconds). CONCLUSION: Our results suggest significant alterations in hippocampal dendritic structure and synaptic function in Apoe4 mice, even at an early age.


Assuntos
Apolipoproteína E4/genética , Hipocampo/citologia , Rede Nervosa/patologia , Neurônios/fisiologia , Sinapses/genética , Animais , Apolipoproteína E3/genética , Fenômenos Biofísicos , Dendritos/ultraestrutura , Espinhas Dendríticas/fisiologia , Modelos Animais de Doenças , Estimulação Elétrica , Potenciais Pós-Sinápticos Excitadores/genética , Hipocampo/fisiologia , Humanos , Técnicas In Vitro , Camundongos , Camundongos Transgênicos , Neurônios/efeitos dos fármacos , Neurônios/ultraestrutura , Estatísticas não Paramétricas , Sinapses/metabolismo , Vesículas Sinápticas/genética
20.
J Comp Neurol ; 525(17): 3653-3665, 2017 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-28758193

RESUMO

A small population of retinal ganglion cells expresses the photopigment melanopsin and function as autonomous photoreceptors. They encode global luminance levels critical for light-mediated non-image forming visual processes including circadian rhythms and the pupillary light reflex. There are five melanopsin ganglion cell subtypes (M1-M5). M1 and displaced M1 (M1d) cells have dendrites that ramify within the outermost layer of the inner plexiform layer. It was recently discovered that some melanopsin ganglion cells extend dendrites into the outer retina. Outer Retinal Dendrites (ORDs) either ramify within the outer plexiform layer (OPL) or the inner nuclear layer, and while present in the mature retina, are most abundant postnatally. Anatomical evidence for synaptic transmission between cone photoreceptor terminals and ORDs suggests a novel photoreceptor to ganglion cell connection in the mammalian retina. While it is known that the number of ORDs in the retina is developmentally regulated, little is known about the morphology, the cells from which they originate, or their spatial distribution throughout the retina. We analyzed the morphology of melanopsin-immunopositive ORDs in the OPL at different developmental time points in the mouse retina and identified five types of ORDs originating from either M1 or M1d cells. However, a pattern emerges within these: ORDs from M1d cells are generally longer and more highly branched than ORDs from conventional M1 cells. Additionally, we found ORDs asymmetrically distributed to the dorsal retina. This morphological analysis provides the first step in identifying a potential role for biplexiform melanopsin ganglion cell ORDs.


Assuntos
Dendritos/metabolismo , Retina/citologia , Células Ganglionares da Retina/citologia , Células Ganglionares da Retina/metabolismo , Opsinas de Bastonetes/metabolismo , Fatores Etários , Animais , Animais Recém-Nascidos , Antraquinonas/metabolismo , Contagem de Células , Dendritos/ultraestrutura , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Opsinas/metabolismo , Retina/crescimento & desenvolvimento , Células Fotorreceptoras Retinianas Cones/metabolismo , Células Ganglionares da Retina/classificação , Proteína Vesicular 1 de Transporte de Glutamato/metabolismo , Vias Visuais/crescimento & desenvolvimento , Vias Visuais/metabolismo
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