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1.
J Neurosci ; 34(11): 4090-8, 2014 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-24623787

RESUMO

TrkA is a tyrosine kinase receptor required for development and survival of the peripheral nervous system. In the adult, TrkA and its ligand NGF are peripheral pain mediators, particularly in inflammatory pain states. However, how TrkA regulates the function of nociceptive neurons and whether its activity levels may lead to sensory abnormalities is still unclear. Here we report the characterization of a 3 aa (KFG) domain that negatively regulates TrkA level and function in response to NGF. Deletion of this domain in mouse causes a reduction of TrkA ubiquitination leading to an increase in TrkA protein levels and activity. The number of dorsal root ganglia neurons is not affected by the mutation. However, mutant mice have enhanced thermal sensitivity and inflammatory pain. Together, these data suggest that ubiquitination is a mechanism used in nociceptive neurons to regulate TrkA level and function. Our results may enhance our understanding of how ubiquitination affects TrkA activation following noxious thermal stimulation and inflammatory pain.


Assuntos
Fator de Crescimento Neural/metabolismo , Nociceptores/metabolismo , Receptor trkA/genética , Receptor trkA/metabolismo , Ubiquitinação/fisiologia , Animais , Linhagem Celular , Regulação para Baixo/fisiologia , Células-Tronco Embrionárias/citologia , Feminino , Gânglios Espinais/citologia , Células HEK293 , Humanos , Hiperalgesia/metabolismo , Hiperalgesia/fisiopatologia , Inflamação/metabolismo , Inflamação/fisiopatologia , Masculino , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Fator de Crescimento Neural/farmacologia , Estrutura Terciária de Proteína , Receptor trkA/química , Células Receptoras Sensoriais/efeitos dos fármacos , Células Receptoras Sensoriais/metabolismo
2.
Am J Physiol Cell Physiol ; 302(1): C141-53, 2012 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-21865582

RESUMO

Neurotrophin-dependent activation of the tyrosine kinase receptor trkB.FL modulates neuromuscular synapse maintenance and function; however, it is unclear what role the alternative splice variant, truncated trkB (trkB.T1), may have in the peripheral neuromuscular axis. We examined this question in trkB.T1 null mice and demonstrate that in vivo neuromuscular performance and nerve-evoked muscle tension are significantly increased. In vitro assays indicated that the gain-in-function in trkB.T1(-/-) animals resulted specifically from an increased muscle contractility, and increased electrically evoked calcium release. In the trkB.T1 null muscle, we identified an increase in Akt activation in resting muscle as well as a significant increase in trkB.FL and Akt activation in response to contractile activity. On the basis of these findings, we conclude that the trkB signaling pathway might represent a novel target for intervention across diseases characterized by deficits in neuromuscular function.


Assuntos
Contração Muscular/genética , Junção Neuromuscular/genética , Receptor trkB/deficiência , Receptor trkB/genética , Animais , Cálcio/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Atividade Motora/genética , Atividade Motora/fisiologia , Contração Muscular/fisiologia , Junção Neuromuscular/fisiologia , Receptor trkB/fisiologia
3.
Transl Psychiatry ; 12(1): 111, 2022 03 17.
Artigo em Inglês | MEDLINE | ID: mdl-35301275

RESUMO

Recent studies have suggested that the use of cognitive enhancers as adjuncts to exposure-based therapy in individuals suffering from post-traumatic stress disorder (PTSD) may be beneficial. Brain cholinergic signaling through basal forebrain projections to the hippocampus is an established pathway mediating fear response and cognitive flexibility. Here we employed a genetic strategy to enhance cholinergic activity through increased signaling of the NGF receptor TrkA. This strategy leads to increased levels of the marker of cholinergic activation, acetylcholine synthesizing enzyme choline acetyltransferase, in forebrain cholinergic regions and their projection areas such as the hippocampus. Mice with increased cholinergic activity do not display any neurobehavioral abnormalities except a selective attenuation of fear response and lower fear expression in extinction trials. Reduction in fear response is rescued by the GABA antagonist picrotoxin in mutant mice, and, in wild-type mice, is mimicked by the GABA agonist midazolam suggesting that GABA can modulate cholinergic functions on fear circuitries. Importantly, mutant mice also show a reduction in fear processing under stress conditions in a single prolonged stress (SPS) model of PTSD-like behavior, and augmentation of cholinergic signaling by the drug donepezil in wild-type mice promotes extinction learning in a similar SPS model of PTSD-like behavior. Donepezil is already in clinical use for the treatment of dementia suggesting a new translational application of this drug for improving exposure-based psychotherapy in PTSD patients.


Assuntos
Prosencéfalo Basal , Transtornos de Estresse Pós-Traumáticos , Animais , Prosencéfalo Basal/metabolismo , Colinérgicos/uso terapêutico , Extinção Psicológica/fisiologia , Medo/fisiologia , Humanos , Camundongos , Transtornos de Estresse Pós-Traumáticos/psicologia
4.
J Cell Biol ; 173(2): 291-9, 2006 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-16636148

RESUMO

Neurotrophins play an essential role in mammalian development. Most of their functions have been attributed to activation of the kinase-active Trk receptors and the p75 neurotrophin receptor. Truncated Trk receptor isoforms lacking the kinase domain are abundantly expressed during development and in the adult; however, their function and signaling capacity is largely unknown. We show that the neurotrophin-3 (NT3) TrkCT1-truncated receptor binds to the scaffold protein tamalin in a ligand-dependent manner. Moreover, NT3 initiation of this complex leads to activation of the Rac1 GTPase through adenosine diphosphate-ribosylation factor 6 (Arf6). At the cellular level, NT3 binding to TrkCT1-tamalin induces Arf6 translocation to the membrane, which in turn causes membrane ruffling and the formation of cellular protrusions. Thus, our data identify a new signaling pathway elicited by the kinase-deficient TrkCT1 receptor. Moreover, we establish NT3 as an upstream regulator of Arf6.


Assuntos
Fatores de Ribosilação do ADP/metabolismo , Proteínas de Transporte/metabolismo , Proteínas de Membrana/metabolismo , Receptor trkC/fisiologia , Transdução de Sinais , Proteínas rac1 de Ligação ao GTP/metabolismo , Fator 6 de Ribosilação do ADP , Linhagem Celular , Células Cultivadas , Humanos , Modelos Biológicos , Isoformas de Proteínas/fisiologia
5.
J Neurosci ; 29(3): 678-85, 2009 Jan 21.
Artigo em Inglês | MEDLINE | ID: mdl-19158294

RESUMO

Pathological or in vitro overexpression of the truncated TrkB (TrkB.T1) receptor inhibits signaling through the full-length TrkB (TrkB.FL) tyrosine kinase receptor. However, to date, the role of endogenous TrkB.T1 is still unknown. By studying mice lacking the truncated TrkB.T1 isoform but retaining normal spatiotemporal expression of TrkB.FL, we have analyzed TrkB.T1-specific physiological functions and its effect on endogenous TrkB kinase signaling in vivo. We found that TrkB.T1-deficient mice develop normally but show increased anxiety in association with morphological abnormalities in the length and complexity of neurites of neurons in the basolateral amygdala. However, no behavioral abnormalities were detected in hippocampal-dependent memory tasks, which correlated with lack of any obvious hippocampal morphological deficits or alterations in basal synaptic transmission and long-term potentiation. In vivo reduction of TrkB signaling by removal of one BDNF allele could be partially rescued by TrkB.T1 deletion, which was revealed by an amelioration of the enhanced aggression and weight gain associated with BDNF haploinsufficiency. Our results suggest that, at the physiological level, TrkB.T1 receptors are important regulators of TrkB.FL signaling in vivo. Moreover, TrkB.T1 selectively affects dendrite complexity of certain neuronal populations.


Assuntos
Encéfalo/anormalidades , Encéfalo/anatomia & histologia , Mutação , Neurônios/fisiologia , Receptor trkB/genética , Animais , Peso Corporal/genética , Encéfalo/ultraestrutura , Fator Neurotrófico Derivado do Encéfalo/genética , Condicionamento Psicológico/fisiologia , Comportamento Exploratório/fisiologia , Medo , Hipocampo/citologia , Técnicas In Vitro , Aprendizagem em Labirinto/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptor trkB/deficiência , Coloração pela Prata/métodos
6.
Nat Commun ; 11(1): 1950, 2020 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-32327658

RESUMO

BDNF signaling in hypothalamic circuitries regulates mammalian food intake. However, whether BDNF exerts metabolic effects on peripheral organs is currently unknown. Here, we show that the BDNF receptor TrkB.T1 is expressed by pancreatic ß-cells where it regulates insulin release. Mice lacking TrkB.T1 show impaired glucose tolerance and insulin secretion. ß-cell BDNF-TrkB.T1 signaling triggers calcium release from intracellular stores, increasing glucose-induced insulin secretion. Additionally, BDNF is secreted by skeletal muscle and muscle-specific BDNF knockout phenocopies the ß-cell TrkB.T1 deletion metabolic impairments. The finding that BDNF is also secreted by differentiated human muscle cells and induces insulin secretion in human islets via TrkB.T1 identifies a new regulatory function of BDNF on metabolism that is independent of CNS activity. Our data suggest that muscle-derived BDNF may be a key factor mediating increased glucose metabolism in response to exercise, with implications for the treatment of diabetes and related metabolic diseases.


Assuntos
Fator Neurotrófico Derivado do Encéfalo/metabolismo , Secreção de Insulina , Células Secretoras de Insulina/metabolismo , Fibras Musculares Esqueléticas/metabolismo , Animais , Fator Neurotrófico Derivado do Encéfalo/genética , Cálcio/metabolismo , Células Cultivadas , Glucose/metabolismo , Intolerância à Glucose , Humanos , Ilhotas Pancreáticas/metabolismo , Masculino , Camundongos , Camundongos Knockout , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Receptor trkB/química , Receptor trkB/genética , Receptor trkB/metabolismo , Transdução de Sinais
7.
Elife ; 82019 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-31429825

RESUMO

Brain-derived neurotrophic factor (BDNF) is a potent modulator of brain synaptic plasticity. Signaling defects caused by dysregulation of its Ntrk2 (TrkB) kinase (TrkB.FL) and truncated receptors (TrkB.T1) have been linked to the pathophysiology of several neurological and neurodegenerative disorders. We found that upregulation of Rbfox1, an RNA binding protein associated with intellectual disability, epilepsy and autism, increases selectively hippocampal TrkB.T1 isoform expression. Physiologically, increased Rbfox1 impairs BDNF-dependent LTP which can be rescued by genetically restoring TrkB.T1 levels. RNA-seq analysis of hippocampi with upregulation of Rbfox1 in conjunction with the specific increase of TrkB.T1 isoform expression also shows that the genes affected by Rbfox1 gain of function are surprisingly different from those influenced by Rbfox1 deletion. These findings not only identify TrkB as a major target of Rbfox1 pathophysiology but also suggest that gain or loss of function of Rbfox1 regulate different genetic landscapes.


Assuntos
Fator Neurotrófico Derivado do Encéfalo/metabolismo , Hipocampo/fisiologia , Potenciação de Longa Duração , Glicoproteínas de Membrana/biossíntese , Proteínas Tirosina Quinases/biossíntese , Fatores de Processamento de RNA/biossíntese , Regulação para Cima , Animais , Perfilação da Expressão Gênica , Camundongos , Isoformas de Proteínas/biossíntese , Análise de Sequência de RNA
8.
Neurobiol Aging ; 46: 160-8, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27498053

RESUMO

Human immunodeficiency virus 1 and its envelope protein gp120 reduce synaptodendritic complexity. However, the mechanisms contributing to this pathological feature are still not understood. The proneurotrophin brain-derived neurotrophic factor promotes synaptic simplification through the activation of the p75 neurotrophin receptor (p75NTR). Here, we have used gp120 transgenic (gp120tg) mice to investigate whether p75NTR has a role in gp120-mediated neurotoxicity. Old (∼10 months) gp120tg mice exhibited an increase in proneurotrophin brain-derived neurotrophic factor levels in the hippocampus as well as a decrease in the number of dendritic spines when compared to age-matched wild type. These effects were not observed in 3- or 6-month-old mice. To test if the reduction in spine density and morphology is caused by the activation of p75NTR, we crossed gp120tg mice with p75NTR null mice. We found that deletion of only 1 copy of the p75NTR gene in gp120tg mice is sufficient to normalize the number of hippocampal spines, strongly suggesting that the neurotoxic effect of gp120 is mediated by p75NTR. These data indicate that p75NTR antagonists could provide an adjunct therapy against synaptic simplification caused by human immunodeficiency virus 1.


Assuntos
Envelhecimento/metabolismo , Envelhecimento/patologia , Espinhas Dendríticas/patologia , Proteína gp120 do Envelope de HIV/toxicidade , Receptor de Fator de Crescimento Neural/fisiologia , Sinapses/patologia , Animais , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Espinhas Dendríticas/virologia , Infecções por HIV/complicações , Infecções por HIV/virologia , HIV-1 , Hipocampo/metabolismo , Hipocampo/patologia , Humanos , Masculino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Terapia de Alvo Molecular , Transtornos Neurocognitivos/etiologia , Transtornos Neurocognitivos/patologia , Transtornos Neurocognitivos/terapia , Receptor de Fator de Crescimento Neural/antagonistas & inibidores , Receptor de Fator de Crescimento Neural/metabolismo , Sinapses/virologia
9.
J Cell Biol ; 210(6): 1003-12, 2015 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-26347138

RESUMO

Brain-derived neurotrophic factor (BDNF) is critical for mammalian development and plasticity of neuronal circuitries affecting memory, mood, anxiety, pain sensitivity, and energy homeostasis. Here we report a novel unexpected role of BDNF in regulating the cardiac contraction force independent of the nervous system innervation. This function is mediated by the truncated TrkB.T1 receptor expressed in cardiomyocytes. Loss of TrkB.T1 in these cells impairs calcium signaling and causes cardiomyopathy. TrkB.T1 is activated by BDNF produced by cardiomyocytes, suggesting an autocrine/paracrine loop. These findings unveil a novel signaling mechanism in the heart that is activated by BDNF and provide evidence for a global role of this neurotrophin in the homeostasis of the organism by signaling through different TrkB receptor isoforms.


Assuntos
Fator Neurotrófico Derivado do Encéfalo/metabolismo , Cardiomiopatias/enzimologia , Glicoproteínas de Membrana/metabolismo , Força Muscular , Contração Miocárdica , Miócitos Cardíacos/enzimologia , Proteínas Tirosina Quinases/metabolismo , Animais , Comunicação Autócrina , Fator Neurotrófico Derivado do Encéfalo/genética , Fator Neurotrófico Derivado do Encéfalo/farmacologia , Sinalização do Cálcio , Cardiomiopatias/genética , Cardiomiopatias/patologia , Cardiomiopatias/fisiopatologia , Ativação Enzimática , Genótipo , Preparação de Coração Isolado , Masculino , Glicoproteínas de Membrana/deficiência , Glicoproteínas de Membrana/genética , Camundongos Knockout , Camundongos Transgênicos , Força Muscular/efeitos dos fármacos , Contração Miocárdica/efeitos dos fármacos , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/patologia , Comunicação Parácrina , Fenótipo , Proteínas Tirosina Quinases/deficiência , Proteínas Tirosina Quinases/genética , Transdução de Sinais , Fatores de Tempo
10.
PLoS One ; 7(6): e39946, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22761934

RESUMO

Brain Derived Neurotrophic Factor (BDNF) exerts strong pro-survival effects on developing and injured motoneurons. However, in clinical trials, BDNF has failed to benefit patients with amyotrophic lateral sclerosis (ALS). To date, the cause of this failure remains unclear. Motoneurons express the TrkB kinase receptor but also high levels of the truncated TrkB.T1 receptor isoform. Thus, we investigated whether the presence of this receptor may affect the response of diseased motoneurons to endogenous BDNF. We deleted TrkB.T1 in the hSOD1(G93A) ALS mouse model and evaluated the impact of this mutation on motoneuron death, muscle weakness and disease progression. We found that TrkB.T1 deletion significantly slowed the onset of motor neuron degeneration. Moreover, it delayed the development of muscle weakness by 33 days. Although the life span of the animals was not affected we observed an overall improvement in the neurological score at the late stage of the disease. To investigate the effectiveness of strategies aimed at bypassing the TrkB.T1 limit to BDNF signaling we treated SOD1 mutant mice with the adenosine A2A receptor agonist CGS21680, which can activate motoneuron TrkB receptor signaling independent of neurotrophins. We found that CGS21680 treatment slowed the onset of motor neuron degeneration and muscle weakness similarly to TrkB.T1 removal. Together, our data provide evidence that endogenous TrkB.T1 limits motoneuron responsiveness to BDNF in vivo and suggest that new strategies such as Trk receptor transactivation may be used for therapeutic intervention in ALS or other neurodegenerative disorders.


Assuntos
Esclerose Lateral Amiotrófica/fisiopatologia , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Receptor trkB/fisiologia , Adenosina/análogos & derivados , Adenosina/farmacologia , Agonistas do Receptor A2 de Adenosina/farmacologia , Animais , Modelos Animais de Doenças , Progressão da Doença , Camundongos , Fenetilaminas/farmacologia , Receptor trkB/genética , Superóxido Dismutase/genética
11.
PLoS One ; 6(10): e27034, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-22066028

RESUMO

Duchenne and Becker muscular dystrophy patients often develop a cardiomyopathy for which the pathogenesis is still unknown. We have employed the murine animal model of Duchenne muscular dystrophy (mdx), which develops a cardiomyopathy that includes some characteristics of the human disease, to study the molecular basis of this pathology. Here we show that the mdx mouse heart has defects consistent with alteration in compounds that regulate energy homeostasis including a marked decrease in creatine-phosphate (PC). In addition, the mdx heart is more susceptible to anoxia than controls. Since the cardio-protective ATP sensitive potassium channel (K(ATP)) complex and PC have been shown to interact we investigated whether deficits in PC levels correlate with other molecular events including K(ATP) ion channel complex presence, its functionality and interaction with dystrophin. We found that this channel complex is present in the dystrophic cardiac cell membrane but its ability to sense a drop in the intracellular ATP concentration and consequently open is compromised by the absence of dystrophin. We further demonstrate that the creatine kinase muscle isoform (CKm) is displaced from the plasma membrane of the mdx cardiac cells. Considering that CKm is a determinant of K(ATP) channel complex function we hypothesize that dystrophin acts as a scaffolding protein organizing the K(ATP) channel complex and the enzymes necessary for its correct functioning. Therefore, the lack of proper functioning of the cardio-protective K(ATP) system in the mdx cardiomyocytes may be part of the mechanism contributing to development of cardiac disease in dystrophic patients.


Assuntos
Cardiotônicos/metabolismo , Distrofina/metabolismo , Canais KATP/metabolismo , Miócitos Cardíacos/metabolismo , Animais , Membrana Celular/enzimologia , Cromatografia Líquida de Alta Pressão , Creatina Quinase/metabolismo , Hipóxia/metabolismo , Imunoprecipitação , Técnicas In Vitro , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos mdx , Modelos Biológicos , Miocárdio/metabolismo , Miocárdio/patologia , Canais de Potássio Corretores do Fluxo de Internalização , Isoformas de Proteínas/metabolismo , Transporte Proteico
13.
Mol Cell ; 10(5): 1151-62, 2002 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-12453422

RESUMO

Passage of E. coli RNA polymerase through an intrinsic transcription terminator, which encodes an RNA hairpin followed by a stretch of uridine residues, results in quick dissociation of the elongation complex. We show that folding of the hairpin disrupts the three upstream base pairs of the 8 bp RNA:DNA hybrid, a major stability determinant in the complex. Shortening the weak rU:dA hybrid from 8 nt to 5 nt causes dissociation of the complex. During termination, the hairpin does not directly compete for base pairing with the 8 bp hybrid. Thus, melting of the hybrid seems to result from spatial restrictions in RNA polymerase that couple the hairpin formation with the disruption of the hybrid immediately downstream from the stem. Our results suggest that a similar mechanism disrupts elongation complexes of yeast RNA polymerase II in vitro.


Assuntos
DNA/química , Escherichia coli/enzimologia , Ácidos Nucleicos Heteroduplexes/metabolismo , RNA Polimerase II/metabolismo , RNA/química , Transcrição Gênica , Sequência de Bases , Modelos Genéticos , Dados de Sequência Molecular , Ácidos Nucleicos Heteroduplexes/química , Hibridização de Ácido Nucleico , Permanganato de Potássio/farmacologia , Ligação Proteica , RNA/metabolismo , Fatores de Tempo
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