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1.
Physiol Rev ; 99(4): 1877-2013, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31460832

RESUMO

The importance of the gut-brain axis in maintaining homeostasis has long been appreciated. However, the past 15 yr have seen the emergence of the microbiota (the trillions of microorganisms within and on our bodies) as one of the key regulators of gut-brain function and has led to the appreciation of the importance of a distinct microbiota-gut-brain axis. This axis is gaining ever more traction in fields investigating the biological and physiological basis of psychiatric, neurodevelopmental, age-related, and neurodegenerative disorders. The microbiota and the brain communicate with each other via various routes including the immune system, tryptophan metabolism, the vagus nerve and the enteric nervous system, involving microbial metabolites such as short-chain fatty acids, branched chain amino acids, and peptidoglycans. Many factors can influence microbiota composition in early life, including infection, mode of birth delivery, use of antibiotic medications, the nature of nutritional provision, environmental stressors, and host genetics. At the other extreme of life, microbial diversity diminishes with aging. Stress, in particular, can significantly impact the microbiota-gut-brain axis at all stages of life. Much recent work has implicated the gut microbiota in many conditions including autism, anxiety, obesity, schizophrenia, Parkinson's disease, and Alzheimer's disease. Animal models have been paramount in linking the regulation of fundamental neural processes, such as neurogenesis and myelination, to microbiome activation of microglia. Moreover, translational human studies are ongoing and will greatly enhance the field. Future studies will focus on understanding the mechanisms underlying the microbiota-gut-brain axis and attempt to elucidate microbial-based intervention and therapeutic strategies for neuropsychiatric disorders.


Assuntos
Bactérias/metabolismo , Encefalopatias/microbiologia , Encéfalo/microbiologia , Microbioma Gastrointestinal , Intestinos/microbiologia , Fatores Etários , Envelhecimento , Animais , Bactérias/imunologia , Bactérias/patogenicidade , Comportamento , Encéfalo/imunologia , Encéfalo/metabolismo , Encéfalo/fisiopatologia , Encefalopatias/metabolismo , Encefalopatias/fisiopatologia , Encefalopatias/psicologia , Disbiose , Sistema Nervoso Entérico/metabolismo , Sistema Nervoso Entérico/microbiologia , Sistema Nervoso Entérico/fisiopatologia , Interações Hospedeiro-Patógeno , Humanos , Intestinos/imunologia , Neuroimunomodulação , Plasticidade Neuronal , Fatores de Risco
2.
Gastroenterology ; 157(1): 179-192.e2, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30930024

RESUMO

BACKGROUND & AIMS: Reduced gastrointestinal (GI) motility is a feature of disorders associated with intestinal dysbiosis and loss of beneficial microbes. It is not clear how consumption of beneficial commensal microbes, marketed as probiotics, affects the enteric nervous system (ENS). We studied the effects of the widely used probiotic and the commensal Lactobacillus rhamnosus GG (LGG) on ENS and GI motility in mice. METHODS: Conventional and germ free C57B6 mice were gavaged with LGG and intestinal tissues were collected; changes in the enteric neuronal subtypes were assessed by real-time polymerase chain reaction, immunoblots, and immunostaining. Production of reactive oxygen species (ROS) in the jejunal myenteric plexi and phosphorylation (p) of mitogen-activated protein kinase 1 (MAPK1) in the enteric ganglia were assessed by immunoblots and immunostaining. Fluorescence in situ hybridization was performed on jejunal cryosections with probes to detect formyl peptide receptor 1 (FPR1). GI motility in conventional mice was assessed after daily gavage of LGG for 1 week. RESULTS: Feeding of LGG to mice stimulated myenteric production of ROS, increased levels of phosphorylated MAPK1, and increased expression of choline acetyl transferase by neurons (P < .001). These effects were not observed in mice given N-acetyl cysteine (a ROS inhibitor) or LGGΩSpaC (an adhesion-mutant strain of LGG) or FPR1-knockout mice. Gavage of mice with LGG for 1 week significantly increased stool frequency, reduced total GI transit time, and increased contractions of ileal circular muscle strips in ex vivo experiments (P < .05). CONCLUSIONS: Using mouse models, we found that LGG-mediated signaling in the ENS requires bacterial adhesion, redox mechanisms, and FPR1. This pathway might be activated to increase GI motility in patients.


Assuntos
Motilidade Gastrointestinal/fisiologia , Trânsito Gastrointestinal/fisiologia , Íleo/metabolismo , Jejuno/metabolismo , Lactobacillus rhamnosus , Plexo Mientérico/metabolismo , Neurônios/metabolismo , Probióticos , Espécies Reativas de Oxigênio/metabolismo , Acetilcisteína/farmacologia , Animais , Antioxidantes/farmacologia , Colina O-Acetiltransferase/metabolismo , Sistema Nervoso Entérico/citologia , Sistema Nervoso Entérico/metabolismo , Motilidade Gastrointestinal/efeitos dos fármacos , Trânsito Gastrointestinal/efeitos dos fármacos , Vida Livre de Germes , Íleo/efeitos dos fármacos , Íleo/inervação , Hibridização in Situ Fluorescente , Jejuno/efeitos dos fármacos , Jejuno/inervação , Camundongos , Camundongos Knockout , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Contração Muscular/efeitos dos fármacos , Plexo Mientérico/citologia , Neurônios/efeitos dos fármacos , Fosforilação , Reação em Cadeia da Polimerase em Tempo Real , Receptores de Formil Peptídeo/genética
3.
Drugs ; 79(8): 797-810, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30982161

RESUMO

Parkinson's disease (PD), the second most common neurodegenerative movement disorder, is characterized by progressive motor and non-motor symptoms [1]. Despite treatment with pharmacologic and surgical therapies, the disease will continue to relentlessly advance. Hence, there is a great deal of interest in potential disease-modifying therapies with the hope that the neurodegenerative process can be slowed or halted. The purpose of this review is to highlight the role toxic α-synuclein (α-syn) plays in PD pathogenesis and critically review the relevant literature about therapeutic modalities targeting α-syn. Toxic α-syn plays a key role in PD pathogenesis, disrupting important cellular functions, and, thus, targeting α-syn is a reasonable disease-modifying strategy. Current approaches under investigation include decreasing α-syn production with RNA interference (RNAi), inhibiting α-syn aggregation, promoting intracellular degradation of α-syn aggregates (via enhancing autophagy and enhancing lysosomal degradation), and promoting extracellular degradation of α-syn via active and passive immunization.


Assuntos
Doença de Parkinson , alfa-Sinucleína/metabolismo , Autofagia , Sistema Nervoso Entérico/metabolismo , Sistema Nervoso Entérico/patologia , Regulação da Expressão Gênica , Humanos , Terapia de Alvo Molecular/métodos , Mutação , Doença de Parkinson/genética , Doença de Parkinson/metabolismo , Doença de Parkinson/patologia , Doença de Parkinson/terapia , Agregados Proteicos , Receptores Adrenérgicos beta/metabolismo , alfa-Sinucleína/antagonistas & inibidores , alfa-Sinucleína/genética
4.
Int J Mol Sci ; 20(7)2019 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-30987291

RESUMO

Diabetic autonomic peripheral neuropathy (PN) involves a broad spectrum of organs. One of them is the gastrointestinal (GI) tract. The molecular mechanisms underlying the pathogenesis of digestive complications are not yet fully understood. Digestion is controlled by the central nervous system (CNS) and the enteric nervous system (ENS) within the wall of the GI tract. Enteric neurons exert regulatory effects due to the many biologically active substances secreted and released by enteric nervous system (ENS) structures. These include nitric oxide (NO), produced by the neural nitric oxide synthase enzyme (nNOS). It is a very important inhibitory factor, necessary for smooth muscle relaxation. Moreover, it was noted that nitrergic innervation can undergo adaptive changes during pathological processes. Additionally, nitrergic neurons function may be regulated through the synthesis of other active neuropeptides. Therefore, in the present study, using the immunofluorescence technique, we first examined the influence of hyperglycemia on the NOS- containing neurons in the porcine small intestine and secondly the co-localization of nNOS with vasoactive intestinal polypeptide (VIP), galanin (GAL) and substance P (SP) in all plexuses studied. Following chronic hyperglycaemia, we observed a reduction in the number of the NOS-positive neurons in all intestinal segments studied, as well as an increased in investigated substances in nNOS positive neurons. This observation confirmed that diabetic hyperglycaemia can cause changes in the neurochemical characteristics of enteric neurons, which can lead to numerous disturbances in gastrointestinal tract functions. Moreover, can be the basis of an elaboration of these peptides analogues utilized as therapeutic agents in the treatment of GI complications.


Assuntos
Intestino Delgado/citologia , Intestino Delgado/metabolismo , Neurônios/metabolismo , Óxido Nítrico Sintase Tipo I/metabolismo , Animais , Sistema Nervoso Central/citologia , Sistema Nervoso Central/metabolismo , Sistema Nervoso Entérico/citologia , Sistema Nervoso Entérico/metabolismo , Feminino , Galanina/metabolismo , Hiperglicemia/metabolismo , Substância P/metabolismo , Suínos , Peptídeo Intestinal Vasoativo/metabolismo
5.
Genesis ; 57(5): e23292, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30884088

RESUMO

Medullary thyroid carcinoma (MTC) develops from hyperplasia of thyroid C cells and represents one of the major causes of thyroid cancer mortality. Mutations in the cysteine-rich domain (CRD) of the RET gene are the most prevalent genetic cause of MTC. The current consensus holds that such cysteine mutations cause ligand-independent dimerization and constitutive activation of RET. However, given the number of the CRD mutations left uncharacterized, our understanding of the pathogenetic mechanisms by which CRD mutations lead to MTC remains incomplete. We report here that RET(C618F), a mutation identified in MTC patients, displays moderately high basal activity and requires the ligand for its full activation. To assess the biological significance of RET(C618F) in organogenesis, we generated a knock-in mouse line conditionally expressing RET(C618F) cDNA by the Ret promoter. The RET(C618F) allele can be made to be Ret-null and express mCherry by Cre-loxP recombination, which allows the assessment of the biological influence of RET(C618F) in vivo. Mice expressing RET(C618F) display mild C cell hyperplasia and increased numbers of enteric neurons, indicating that RET(C618F) confers gain-of-function phenotypes. This mouse line serves as a novel biological platform for investigating pathogenetic mechanisms involved in MTC and enteric hyperganglionosis.


Assuntos
Carcinoma Neuroendócrino/genética , Proteínas Proto-Oncogênicas c-ret/genética , Neoplasias da Glândula Tireoide/genética , Animais , Carcinoma Neuroendócrino/metabolismo , Linhagem Celular Tumoral , Sistema Nervoso Entérico/metabolismo , Sistema Nervoso Entérico/patologia , Técnicas de Introdução de Genes/métodos , Mutação em Linhagem Germinativa , Humanos , Hiperplasia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mutação , Doenças do Sistema Nervoso/genética , Doenças do Sistema Nervoso/metabolismo , Doenças do Sistema Nervoso/patologia , Proteínas Proto-Oncogênicas c-ret/biossíntese , Proteínas Proto-Oncogênicas c-ret/metabolismo , Hiperplasia do Timo/genética , Hiperplasia do Timo/metabolismo , Glândula Tireoide/metabolismo , Glândula Tireoide/patologia , Neoplasias da Glândula Tireoide/metabolismo
6.
Nat Protoc ; 14(4): 1261-1279, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30911172

RESUMO

The enteric nervous system (ENS) represents a vast network of neuronal and glial cell types that develops entirely from migratory neural crest (NC) progenitor cells. Considerable improvements in the understanding of the molecular mechanisms underlying NC induction and regional specification have recently led to the development of a robust method to re-create the process in vitro using human pluripotent stem cells (hPSCs). Directing the fate of hPSCs toward the enteric NC (ENC) results in an accessible and scalable in vitro model of ENS development. The application of hPSC-derived enteric neural lineages provides a powerful platform for ENS-related disease modeling and drug discovery. Here we present a detailed protocol for the induction of a regionally specific NC intermediate that occurs over the course of a 15-d interval and is an effective source for the in vitro derivation of functional enteric neurons (ENs) from hPSCs. Additionally, we introduce a new and improved protocol that we have developed to optimize the protocol for future applications in regenerative medicine, in which components of undefined activity have been replaced with fully defined culture conditions. This protocol provides access to a broad range of human ENS lineages within a 30-d period.


Assuntos
Técnicas de Cultura de Células , Sistema Nervoso Entérico/citologia , Intestino Delgado/citologia , Crista Neural/citologia , Neurônios/citologia , Células-Tronco Pluripotentes/citologia , Antígenos CD/genética , Antígenos CD/metabolismo , Biomarcadores/metabolismo , Diferenciação Celular , Linhagem da Célula/fisiologia , Sistema Nervoso Entérico/metabolismo , Expressão Gênica , Genes Reporter , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Intestino Delgado/metabolismo , Crista Neural/metabolismo , Neurônios/metabolismo , Fator de Transcrição PAX3/genética , Fator de Transcrição PAX3/metabolismo , Células-Tronco Pluripotentes/metabolismo , Medicina Regenerativa/métodos
7.
Mol Autism ; 10: 3, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30733854

RESUMO

Background and aims: Autism spectrum disorder (ASD) is currently estimated to affect more than 1% of the world population. For people with ASD, gastrointestinal (GI) distress is a commonly reported but a poorly understood co-occurring symptom. Here, we investigate the physiological basis for GI distress in ASD by studying gut function in a zebrafish model of Phelan-McDermid syndrome (PMS), a condition caused by mutations in the SHANK3 gene. Methods: To generate a zebrafish model of PMS, we used CRISPR/Cas9 to introduce clinically related C-terminal frameshift mutations in shank3a and shank3b zebrafish paralogues (shank3abΔC). Because PMS is caused by SHANK3 haploinsufficiency, we assessed the digestive tract (DT) structure and function in zebrafish shank3abΔC +/- heterozygotes. Human SHANK3 mRNA was then used to rescue DT phenotypes in larval zebrafish. Results: Significantly slower rates of DT peristaltic contractions (p < 0.001) with correspondingly prolonged passage time (p < 0.004) occurred in shank3abΔC +/- mutants. Rescue injections of mRNA encoding the longest human SHANK3 isoform into shank3abΔC +/- mutants produced larvae with intestinal bulb emptying similar to wild type (WT), but still deficits in posterior intestinal motility. Serotonin-positive enteroendocrine cells (EECs) were significantly reduced in both shank3abΔC +/- and shank3abΔC -/- mutants (p < 0.05) while enteric neuron counts and overall structure of the DT epithelium, including goblet cell number, were unaffected in shank3abΔC +/- larvae. Conclusions: Our data and rescue experiments support mutations in SHANK3 as causal for GI transit and motility abnormalities. Reductions in serotonin-positive EECs and serotonin-filled ENS boutons suggest an endocrine/neural component to this dysmotility. This is the first study to date demonstrating DT dysmotility in a zebrafish single gene mutant model of ASD.


Assuntos
Transtorno Autístico/genética , Motilidade Gastrointestinal , Proteínas do Tecido Nervoso/genética , Proteínas de Peixe-Zebra/genética , Animais , Transtorno Autístico/fisiopatologia , Sistema Nervoso Entérico/citologia , Sistema Nervoso Entérico/metabolismo , Células Enteroendócrinas/metabolismo , Mucosa Intestinal/metabolismo , Intestinos/citologia , Intestinos/crescimento & desenvolvimento , Intestinos/fisiologia , Mutação , Neurônios/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Serotonina/metabolismo , Peixe-Zebra
8.
Methods Mol Biol ; 1950: 407-415, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30783988

RESUMO

The enteric nervous system of the lower gastrointestinal tract comprises intrinsic neural circuits as well as extrinsic afferent and efferent innervation. The development of strategies for neuronal gene transfer has created new opportunities for functional analysis, circuit mapping, and neuromodulation in the enteric nervous system. Studies of AAV-mediated gene transfer to enteric neurons and dorsal root ganglion neurons (DRG) have provided proofs-of-concept for the utility of AAV vectors for genetic manipulations of the intrinsic and extrinsic components of the enteric nervous system. Here we describe a method for AAV-mediated gene transfer to enteric neurons of the descending colon as well as colon-innervating DRG neurons by injection within the intestinal wall (intracolonic injection).


Assuntos
Dependovirus/genética , Sistema Nervoso Entérico/metabolismo , Técnicas de Transferência de Genes , Vetores Genéticos/genética , Animais , Feminino , Expressão Gênica , Genes Reporter , Vetores Genéticos/administração & dosagem , Masculino , Camundongos , Transdução Genética , Transgenes
9.
PLoS One ; 14(2): e0212856, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30794676

RESUMO

Antibiotic use during adolescence may result in dysbiosis-induced neuronal vulnerability both in the enteric nervous system (ENS) and central nervous system (CNS) contributing to the onset of chronic gastrointestinal disorders, such as irritable bowel syndrome (IBS), showing significant psychiatric comorbidity. Intestinal microbiota alterations during adolescence influence the expression of molecular factors involved in neuronal development in both the ENS and CNS. In this study, we have evaluated the expression of brain-derived neurotrophic factor (BDNF) and its high-affinity receptor tropomyosin-related kinase B (TrkB) in juvenile mice ENS and CNS, after a 2-week antibiotic (ABX) treatment. In both mucosa and mucosa-deprived whole-wall small intestine segments of ABX-treated animals, BDNF and TrKB mRNA and protein levels significantly increased. In longitudinal muscle-myenteric plexus preparations of ABX-treated mice the percentage of myenteric neurons staining for BDNF and TrkB was significantly higher than in controls. After ABX treatment, a consistent population of BDNF- and TrkB-immunoreactive neurons costained with SP and CGRP, suggesting up-regulation of BDNF signaling in both motor and sensory myenteric neurons. BDNF and TrkB protein levels were downregulated in the hippocampus and remained unchanged in the prefrontal cortex of ABX-treated animals. Immunostaining for BDNF and TrkB decreased in the hippocampus CA3 and dentate gyrus subregions, respectively, and remained unchanged in the prefrontal cortex. These data suggest that dysbiosis differentially influences the expression of BDNF-TrkB in the juvenile mice ENS and CNS. Such changes may potentially contribute later to the development of functional gut disorders, such as IBS, showing psychiatric comorbidity.


Assuntos
Antibacterianos/efeitos adversos , Fator Neurotrófico Derivado do Encéfalo/biossíntese , Encéfalo/metabolismo , Disbiose/metabolismo , Sistema Nervoso Entérico/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Glicoproteínas de Membrana/biossíntese , Proteínas Tirosina Quinases/biossíntese , Animais , Antibacterianos/farmacologia , Encéfalo/patologia , Disbiose/induzido quimicamente , Disbiose/patologia , Sistema Nervoso Entérico/patologia , Síndrome do Intestino Irritável/induzido quimicamente , Síndrome do Intestino Irritável/metabolismo , Síndrome do Intestino Irritável/patologia , Camundongos , Neurônios/metabolismo , Neurônios/patologia , Transdução de Sinais/efeitos dos fármacos
10.
Dev Biol ; 446(1): 22-33, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30448439

RESUMO

The enteric nervous system is mostly derived from vagal neural crest (NC) cells adjacent to somites (s)1-7. We used in ovo focal fluorescent vital dyes and focal electroporation of fluorophore-encoding plasmids in quail embryos to investigate NC cell migration to the foregut initially and later throughout the entire gut. NC cells of different somite-level origins were largely separate until reaching the foregut at about QE2.5, when all routes converged. By QE3.5, NC cells of different somite-levels became mixed, although s1-s2 NC cells were mainly confined to rostral foregut. Mid-vagal NC-derived cells (s3 and s4 level) arrived earliest at the foregut, and occurred in greatest number. By QE6.5 ENS was present from foregut to hindgut. Mid-vagal NC-derived cells occurred in greatest numbers from foregut to distal hindgut. NC-derived cells of s2, s5, and s6 levels were fewer and were widely distributed but were never observed in the distal hindgut. Rostro-vagal (s1) and caudo-vagal (s7) levels were few and restricted to the foregut. Single somite levels of quail neural tube/NC from s1 to s8 were combined with chick aneural ChE4.5 midgut and hindgut and the ensemble was grown on the chorio-allantoic membrane for 6 days. This tests ENS-forming competence in the absence of intra-segmental competition between NC cells, of differential influences of segmental paraxial tissues, and of positional advantage. All vagal NC-levels, but not s8 level, furnished enteric plexuses in the recipient gut, but the density of both ENS cells in total and neurons was highest from mid-vagal level donors, as was the length colonised. We conclude that the fate and competence for ENS formation of vagal NC sub-levels is not uniform over the vagal level but is biased to favour mid-vagal levels. Overviewing this and prior studies suggests the vagal region is, as in its traditional sense, a natural unit but with complex sub-divisions.


Assuntos
Sistema Nervoso Entérico/embriologia , Crista Neural/embriologia , Somitos/embriologia , Nervo Vago/embriologia , Animais , Padronização Corporal , Diferenciação Celular , Movimento Celular , Embrião de Galinha , Galinhas , Coturnix , Sistema Digestório/citologia , Sistema Digestório/embriologia , Sistema Digestório/metabolismo , Sistema Nervoso Entérico/citologia , Sistema Nervoso Entérico/metabolismo , Intestinos/citologia , Intestinos/embriologia , Intestinos/inervação , Crista Neural/citologia , Crista Neural/metabolismo , Neurônios/citologia , Neurônios/metabolismo , Somitos/citologia , Somitos/metabolismo , Nervo Vago/citologia , Nervo Vago/metabolismo
11.
Dev Biol ; 446(1): 34-42, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30529057

RESUMO

Cells of the vagal neural crest (NC) form most of the enteric nervous system (ENS) by a colonising wave in the embryonic gut, with high cell proliferation and differentiation. Enteric neuropathies have an ENS deficit and cell replacement has been suggested as therapy. This would be performed post-natally, which raises the question of whether the ENS cell population retains its initial ENS-forming potential with age. We tested this on the avian model in organ culture in vitro (3 days) using recipient aneural chick midgut/hindgut combined with ENS-donor quail midgut or hindgut of ages QE5 to QE10. ENS cells from young donor tissues (≤ QE6) avidly colonised the aneural recipient, but this capacity dropped rapidly 2-3 days after the transit of the ENS cell wavefront. This loss in capability was autonomous to the ENS population since a similar decline was observed in ENS cells isolated by HNK1 FACS. Using QE5, 6, 8 and 10 midgut donors and extending the time of assay to 8 days in chorio-allantoic membrane grafts did not produce 'catch up' colonisation. NC-derived cells were counted in dissociated quail embryo gut and in transverse sections of chick embryo gut using NC, neuron and glial marker antibodies. This showed that the decline in ENS-forming ability correlated with a decrease in proportion of ENS cells lacking both neuronal and glial differentiation markers, but there were still large numbers of such cells even at stages with low colonisation ability. Moreover, ENS cells in small numbers from young donors were far superior in colonisation ability to larger numbers of apparently undifferentiated cells from older donors. This suggests that the decline of ENS-forming ability has both quantitative and qualitative aspects. In this case, ENS cells for cell therapies should aim to replicate the embryonic ENS stage rather than using post-natal ENS stem/progenitor cells.


Assuntos
Sistema Digestório/embriologia , Sistema Nervoso Entérico/embriologia , Intestino Delgado/embriologia , Crista Neural/embriologia , Animais , Diferenciação Celular , Movimento Celular , Células Cultivadas , Embrião de Galinha , Galinhas , Membrana Corioalantoide/transplante , Coturnix , Sistema Digestório/citologia , Sistema Digestório/metabolismo , Sistema Nervoso Entérico/citologia , Sistema Nervoso Entérico/metabolismo , Intestino Delgado/citologia , Intestino Delgado/inervação , Crista Neural/citologia , Crista Neural/metabolismo , Neuroglia/citologia , Neuroglia/metabolismo , Neurônios/citologia , Neurônios/metabolismo , Técnicas de Cultura de Órgãos
12.
Dev Biol ; 445(2): 256-270, 2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30472119

RESUMO

The enteric nervous system is thought to originate solely from the neural crest. Transgenic lineage tracing revealed a novel population of clonal pancreatic duodenal homeobox-1 (Pdx1)-Cre lineage progenitor cells in the tunica muscularis of the gut that produced pancreatic descendants as well as neurons upon differentiation in vitro. Additionally, an in vivo subpopulation of endoderm lineage enteric neurons, but not glial cells, was seen especially in the proximal gut. Analysis of early transgenic embryos revealed Pdx1-Cre progeny (as well as Sox-17-Cre and Foxa2-Cre progeny) migrating from the developing pancreas and duodenum at E11.5 and contributing to the enteric nervous system. These results show that the mammalian enteric nervous system arises from both the neural crest and the endoderm. Moreover, in adult mice there are separate Wnt1-Cre neural crest stem cells and Pdx1-Cre pancreatic progenitors within the muscle layer of the gut.


Assuntos
Sistema Nervoso Entérico/embriologia , Animais , Linhagem da Célula/genética , Duodeno/embriologia , Duodeno/inervação , Duodeno/metabolismo , Endoderma/citologia , Endoderma/embriologia , Endoderma/metabolismo , Sistema Nervoso Entérico/citologia , Sistema Nervoso Entérico/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Proteínas HMGB/genética , Proteínas HMGB/metabolismo , Fator 3-beta Nuclear de Hepatócito/genética , Fator 3-beta Nuclear de Hepatócito/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Camundongos , Camundongos Transgênicos , Crista Neural/citologia , Crista Neural/embriologia , Crista Neural/metabolismo , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Pâncreas/embriologia , Pâncreas/inervação , Pâncreas/metabolismo , Fatores de Transcrição SOXF/genética , Fatores de Transcrição SOXF/metabolismo , Transativadores/genética , Transativadores/metabolismo , Proteína Wnt1/genética , Proteína Wnt1/metabolismo
13.
J Neurochem ; 148(6): 746-760, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30589944

RESUMO

The protein alpha-synuclein whose expression is strongly implicated in Parkinson's disease (PD) is not only expressed in the CNS but also in the enteric nervous system (ENS). The growing body of evidence suggesting that gastrointestinal inflammation is involved in the development of PD led us to investigate the effects of inflammation on alpha-synuclein expression in primary culture of rat ENS and in mice with dextran sulfate sodium-induced colitis. Using western blot and qPCR, we found that both lipopolysaccharide and a combination of tumor necrosis factor-α and interleukin 1-ß decreased the expression levels of alpha-synuclein in primary culture of rat ENS, an effect that was prevented in the presence of the p38 inhibitors SB203580 and BIRB 796. Lipopolysaccharide and tumor necrosis factor-α/interleukin 1-ß had no effect on alpha-synuclein expression in primary culture of rat CNS and in human erythroid leukemia cells. In mice, acute but not chronic dextran sulfate sodium-induced colitis was associated with a decreased expression of colonic alpha-synuclein. As a whole, our findings indicate that acute inflammatory insults down-regulate alpha-synuclein expression in the ENS via a p38 pathway. They provide new insights into the widely discussed concepts of alpha-synuclein expression and aggregation in the ENS in PD and raise issues about the possible role of gastrointestinal inflammation in the development of PD. OPEN SCIENCE BADGES: This article has received a badge for *Open Materials* because it provided all relevant information to reproduce the study in the manuscript. The complete Open Science Disclosure form for this article can be found at the end of the article. More information about the Open Practices badges can be found at https://cos.io/our-services/open-science-badges/.


Assuntos
Sistema Nervoso Entérico/metabolismo , Inflamação/metabolismo , Neurônios/metabolismo , alfa-Sinucleína/biossíntese , Animais , Regulação para Baixo , Sistema Nervoso Entérico/patologia , Humanos , Inflamação/patologia , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/patologia , Ratos , Ratos Sprague-Dawley , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
14.
Cell Tissue Res ; 375(3): 605-618, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30324494

RESUMO

Piezo channels play fundamental roles in many physiological processes. Their presence and functional role in the enteric nervous system is still not known. We hypothesize that they play a role in mechanotransduction in enteric neurons. Our aims are to quantify the presence of both Piezo1 and 2 in enteric neurons throughout the gastrointestinal tract using immunohistochemistry and analyze their function(s) using neuroimaging techniques and pharmacological investigations. In order to perform a systematic and comparative study, we performed our experiments in gastrointestinal tissue from guinea pigs, mice and humans. Piezo1 (20-70%) is expressed by both enteric neuronal cell bodies and fibers in the myenteric and submucosal plexi of all the species investigated. Generally, Piezo1 expressing somata are more numerous in the submucosal plexus (50-80%) than in the myenteric plexus (15-35%) apart from the stomach where Piezo1 is expressed in up to 60% of cell bodies. Myenteric Piezo1 neurons mainly (60-100%) but not exclusively, also express nitric oxide synthase, a minority express choline acetyltransferase. In the submucosal plexus, Piezo1 neurons co-express vasoactive intestinal peptide (40-90%). Conversely, expression of Piezo2 is extremely rare in the somata of enteric neurons and is present in few neurites. In functional experiments, 38-76% of the mechanosensitive neurons expressed Piezo1 channels. Statistical analysis showed a positive significant correlation between mechanosensitive and Piezo1 positive neurons. However, pharmacological experiments using an activator and an inhibitor of Piezo channels did not demonstrate changes in mechanotransduction. A major role of Piezo1 in the mechanosensitivity of enteric neurons can be excluded.


Assuntos
Sistema Nervoso Entérico/metabolismo , Mecanotransdução Celular , Proteínas de Membrana/metabolismo , Animais , Feminino , Cobaias , Humanos , Masculino , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Neurônios/metabolismo
15.
Cell Mol Gastroenterol Hepatol ; 7(3): 655-678, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30594740

RESUMO

BACKGROUND & AIMS: RET, the receptor for the glial cell line-derived neurotrophic factor (GDNF) family ligands, is the most frequently mutated gene in congenital aganglionic megacolon or Hirschsprung's disease (HSCR). The leading cause of mortality in HSCR is HSCR-associated enterocolitis (HAEC), which is characterized by altered mucin composition, mucin retention, bacterial adhesion to enterocytes, and epithelial damage, although the order of these events is obscure. In mice, loss of GDNF signaling leads to a severely underdeveloped enteric nervous system and neonatally fatal kidney agenesis, thereby precluding the use of these mice for modeling postnatal HSCR and HAEC. Our aim was to generate a postnatally viable mouse model for HSCR/HAEC and analyze HAEC etiology. METHODS: GDNF family receptor alpha-1 (GFRa1) hypomorphic mice were generated by placing a selectable marker gene in the sixth intron of the Gfra1 locus using gene targeting in mouse embryonic stem cells. RESULTS: We report that 70%-80% reduction in GDNF co-receptor GFRa1 expression levels in mice results in HSCR and HAEC, leading to death within the first 25 postnatal days. These mice mirror the disease progression and histopathologic findings in children with untreated HSCR/HAEC. CONCLUSIONS: In GFRa1 hypomorphic mice, HAEC proceeds from goblet cell dysplasia, with abnormal mucin production and retention, to epithelial damage. Microbial enterocyte adherence and tissue invasion are late events and therefore unlikely to be the primary cause of HAEC. These results suggest that goblet cells may be a potential target for preventative treatment and that reduced expression of GFRa1 may contribute to HSCR susceptibility.


Assuntos
Enterocolite/complicações , Enterocolite/metabolismo , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Doença de Hirschsprung/complicações , Doença de Hirschsprung/metabolismo , Animais , Proteínas Sanguíneas/metabolismo , Neurônios Colinérgicos/metabolismo , Colo/inervação , Colo/patologia , Citocinas/genética , Citocinas/metabolismo , Sistema Nervoso Entérico/metabolismo , Sistema Nervoso Entérico/patologia , Enterocolite/sangue , Genótipo , Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Células Caliciformes/patologia , Doença de Hirschsprung/sangue , Homozigoto , Hipertrofia , Mucosa Intestinal/patologia , Camundongos Endogâmicos C57BL , Mucinas/metabolismo , Células-Tronco Neurais/metabolismo , Proteínas Proto-Oncogênicas c-ret , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
16.
Cell ; 176(1-2): 85-97.e14, 2019 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-30580965

RESUMO

Animals must respond to the ingestion of food by generating adaptive behaviors, but the role of gut-brain signaling in behavioral regulation is poorly understood. Here, we identify conserved ion channels in an enteric serotonergic neuron that mediate its responses to food ingestion and decipher how these responses drive changes in foraging behavior. We show that the C. elegans serotonergic neuron NSM acts as an enteric sensory neuron that acutely detects food ingestion. We identify the novel and conserved acid-sensing ion channels (ASICs) DEL-7 and DEL-3 as NSM-enriched channels required for feeding-dependent NSM activity, which in turn drives slow locomotion while animals feed. Point mutations that alter the DEL-7 channel change NSM dynamics and associated behavioral dynamics of the organism. This study provides causal links between food ingestion, molecular and physiological properties of an enteric serotonergic neuron, and adaptive feeding behaviors, yielding a new view of how enteric neurons control behavior.


Assuntos
Canais Iônicos Sensíveis a Ácido/metabolismo , Sistema Nervoso Entérico/metabolismo , Comportamento Alimentar/fisiologia , Canais Iônicos Sensíveis a Ácido/fisiologia , Animais , Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/fisiologia , Proteínas de Caenorhabditis elegans/metabolismo , Sistema Nervoso Entérico/fisiologia , Alimentos , Canais Iônicos/metabolismo , Canais Iônicos/fisiologia , Locomoção , Neurônios/metabolismo , Células Receptoras Sensoriais/metabolismo , Neurônios Serotoninérgicos/metabolismo , Neurônios Serotoninérgicos/fisiologia , Serotonina , Transdução de Sinais
17.
J Parkinsons Dis ; 8(s1): S31-S39, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30584161

RESUMO

In the last two decades it has become clear that Parkinson's disease (PD) is associated with a plethora of gastrointestinal symptoms originating from functional and structural changes in the gut and its associated neural structures. This is of particular interest not only because such symptoms have a major impact on the quality of life of PD patients, but also since accumulating evidence suggests that in at least a subgroup of patients, these disturbances precede the motor symptoms and diagnosis of PD by years and may thus give important insights into the origin and pathogenesis of the disease. In this mini-review we attempt to concisely summarize the current knowledge after two decades of research on the gut-brain axis in PD. We focus on alpha-synuclein pathology, biomarkers, and the gut microbiota and envision the development and impact of these research areas for the two decades to come.


Assuntos
Sistema Nervoso Entérico/microbiologia , Microbioma Gastrointestinal/fisiologia , Doença de Parkinson/microbiologia , Biomarcadores , Encéfalo/metabolismo , Sistema Nervoso Entérico/metabolismo , Humanos , Doença de Parkinson/metabolismo , alfa-Sinucleína/metabolismo
18.
J Neurosci ; 38(44): 9346-9354, 2018 10 31.
Artigo em Inglês | MEDLINE | ID: mdl-30381426

RESUMO

The enteric nervous system (ENS) is a large, complex division of the peripheral nervous system that regulates many digestive, immune, hormonal, and metabolic functions. Recent advances have elucidated the dynamic nature of the mature ENS, as well as the complex, bidirectional interactions among enteric neurons, glia, and the many other cell types that are important for mediating gut behaviors. Here, we provide an overview of ENS development and maintenance, and focus on the latest insights gained from the use of novel model systems and live-imaging techniques. We discuss major advances in the understanding of enteric glia, and the functional interactions among enteric neurons, glia, and enteroendocrine cells, a large class of sensory epithelial cells. We conclude by highlighting recent work on muscularis macrophages, a group of immune cells that closely interact with the ENS in the gut wall, and the importance of neurological-immune system communication in digestive health and disease.


Assuntos
Encéfalo/metabolismo , Sistema Nervoso Entérico/metabolismo , Gastroenteropatias/metabolismo , Trato Gastrointestinal/metabolismo , Animais , Encéfalo/imunologia , Encéfalo/patologia , Sistema Nervoso Entérico/imunologia , Sistema Nervoso Entérico/patologia , Gastroenteropatias/imunologia , Gastroenteropatias/patologia , Trato Gastrointestinal/imunologia , Trato Gastrointestinal/patologia , Humanos , Neurobiologia
19.
Curr Opin Pharmacol ; 43: 145-149, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30340053

RESUMO

Pediatric gastrointestinal motility disorders represent a range of severe developmental or acquired conditions that disrupt enteric neuromuscular function. Current medical and surgical therapeutic options are very limited but recent advances have highlighted the possibility of improved or curative stem cell-based treatments. Not only has the ability to harvest, propagate and transplant human-derived enteric neural stem cells (ENSCs) been demonstrated but recent in vivo transplantation studies have confirmed that ENSCs are capable of engraftment within recipient intestine of animal models of enteric neuropathy and effecting functional rescue. Pluripotent stem cell-derived cells and pharmacological modulation of both endogenous and transplanted neural stem cells have further enhanced the exciting prospect of clinical application of such stem cell-based therapies in the near future.


Assuntos
Sistema Nervoso Entérico/cirurgia , Gastroenteropatias/cirurgia , Motilidade Gastrointestinal , Trato Gastrointestinal/cirurgia , Células-Tronco Neurais/transplante , Células-Tronco Pluripotentes/transplante , Transplante de Células-Tronco/métodos , Fatores Etários , Animais , Sistema Nervoso Entérico/metabolismo , Sistema Nervoso Entérico/fisiopatologia , Gastroenteropatias/diagnóstico , Gastroenteropatias/metabolismo , Gastroenteropatias/fisiopatologia , Trato Gastrointestinal/inervação , Trato Gastrointestinal/metabolismo , Humanos , Regeneração Nervosa , Células-Tronco Neurais/metabolismo , Fenótipo , Células-Tronco Pluripotentes/metabolismo , Recuperação de Função Fisiológica , Fatores de Risco , Índice de Gravidade de Doença , Transplante de Células-Tronco/efeitos adversos , Resultado do Tratamento
20.
Histochem Cell Biol ; 150(6): 703-709, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30250972

RESUMO

Recent advances in neurogastroenterology have extended and refined our knowledge on the roles monoamines play in physiology and pathophysiology of the gastrointestinal tract. The catecholamine noradrenaline, as the primary transmitter of postganglionic sympathetic neurons, orchestrates motility and secretory reflexes and controls arterial perfusion as well as immune functions. The catecholamine dopamine is produced by a subpopulation of enteric neurons which possibly use it as transmitter. Serotonin, largely produced by enterochromaffin cells and to a small extent by enteric neurons profoundly affects gut motility, enteric neuron development and is also involved in immunomodulation. However, its mode of action and the relative contribution of non-neuronal versus neuronal serotonin was recently subject to debate again. Histamine, although entirely of non-neuronal origin, is pivotal for gastrointestinal neuroimmunomodulation besides its paracrine effect in gastric HCl production.


Assuntos
Aminas/metabolismo , Sistema Nervoso Entérico/metabolismo , Catecolaminas/biossíntese , Catecolaminas/química , Humanos , Serotonina/biossíntese , Serotonina/química
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