RESUMO
Clinical studies have reported that the psychedelic lysergic acid diethylamide (LSD) enhances empathy and social behavior (SB) in humans, but its mechanism of action remains elusive. Using a multidisciplinary approach including in vivo electrophysiology, optogenetics, behavioral paradigms, and molecular biology, the effects of LSD on SB and glutamatergic neurotransmission in the medial prefrontal cortex (mPFC) were studied in male mice. Acute LSD (30 µg/kg) injection failed to increase SB. However, repeated LSD (30 µg/kg, once a day, for 7 days) administration promotes SB, without eliciting antidepressant/anxiolytic-like effects. Optogenetic inhibition of mPFC excitatory neurons dramatically inhibits social interaction and nullifies the prosocial effect of LSD. LSD potentiates the α-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) and 5-HT2A, but not N-methyl-D-aspartate (NMDA) and 5-HT1A, synaptic responses in the mPFC and increases the phosphorylation of the serine-threonine protein kinases Akt and mTOR. In conditional knockout mice lacking Raptor (one of the structural components of the mTORC1 complex) in excitatory glutamatergic neurons (Raptorf/f:Camk2alpha-Cre), the prosocial effects of LSD and the potentiation of 5-HT2A/AMPA synaptic responses were nullified, demonstrating that LSD requires the integrity of mTORC1 in excitatory neurons to promote SB. Conversely, in knockout mice lacking Raptor in GABAergic neurons of the mPFC (Raptorf/f:Gad2-Cre), LSD promotes SB. These results indicate that LSD selectively enhances SB by potentiating mPFC excitatory transmission through 5-HT2A/AMPA receptors and mTOR signaling. The activation of 5-HT2A/AMPA/mTORC1 in the mPFC by psychedelic drugs should be explored for the treatment of mental diseases with SB impairments such as autism spectrum disorder and social anxiety disorder.
Assuntos
Comportamento Animal/efeitos dos fármacos , Dietilamida do Ácido Lisérgico/farmacologia , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Comportamento Social , Transmissão Sináptica/efeitos dos fármacos , Animais , Aprendizagem da Esquiva/efeitos dos fármacos , Masculino , Camundongos Endogâmicos C57BL , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Optogenética , Fosforilação/efeitos dos fármacos , Córtex Pré-Frontal/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Células Piramidais/efeitos dos fármacos , Células Piramidais/metabolismo , Receptores de AMPA/agonistas , Receptores de AMPA/metabolismo , Receptores de N-Metil-D-Aspartato/agonistas , Receptores de N-Metil-D-Aspartato/metabolismo , Receptores de Serotonina/metabolismo , Sinapses/efeitos dos fármacos , Sinapses/metabolismo , Serina-Treonina Quinases TOR/metabolismoRESUMO
Translational control plays a key role in regulation of neuronal activity and behavior. Deletion of the translational repressor 4E-BP2 in mice alters excitatory and inhibitory synaptic functions, engendering autistic-like behaviors. The contribution of 4E-BP2-dependent translational control in excitatory and inhibitory neurons and astrocytic cells to these behaviors remains unknown. To investigate this, we generated cell-type-specific conditional 4E-BP2 knockout mice and tested them for the salient features of autism, including repetitive stereotyped behaviors (self-grooming and marble burying), sociability (3-chamber social and direct social interaction tests), and communication (ultrasonic vocalizations in pups). We found that deletion of 4E-BP2 in GABAergic inhibitory neurons, defined by Gad2, resulted in impairments in social interaction and vocal communication. In contrast, deletion of 4E-BP2 in forebrain glutamatergic excitatory neurons, defined by Camk2a, or in astrocytes, defined by Gfap, failed to cause autistic-like behavioral abnormalities. Taken together, we provide evidence for an inhibitory-cell-specific role of 4E-BP2 in engendering autism-related behaviors.
Assuntos
Transtorno Autístico/metabolismo , Comportamento Animal , Fatores de Iniciação em Eucariotos/deficiência , Neurônios GABAérgicos/metabolismo , Interneurônios/metabolismo , Biossíntese de Proteínas , Animais , Astrócitos/metabolismo , Astrócitos/patologia , Transtorno Autístico/genética , Transtorno Autístico/patologia , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/genética , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Neurônios GABAérgicos/patologia , Proteína Glial Fibrilar Ácida/genética , Proteína Glial Fibrilar Ácida/metabolismo , Glutamato Descarboxilase/genética , Glutamato Descarboxilase/metabolismo , Interneurônios/patologia , Camundongos , Camundongos KnockoutRESUMO
Mammalian target of rapamycin (mTOR) is a serine-threonine kinase involved in almost every aspect of mammalian cell function. This kinase was initially believed to control protein translation in response to amino acids and trophic factors, and this function has become a canonical role for mTOR. However, mTOR can form two separate protein complexes (mTORCs). Recent advances clearly demonstrate that both mTORCs can respond to various stimuli and change myriad cellular processes. Therefore, our current view of the cellular roles of TORCs has rapidly expanded and cannot be fully explained without appreciating recent findings about the new modes of mTOR regulation and identification of non-canonical effectors of mTOR that contribute to transcription, cytoskeleton dynamics, and membrane trafficking. This review discusses the molecular details of these newly discovered non-canonical functions that allow mTORCs to control the cellular environment at multiple levels. This article is part of a Special Issue entitled: Inhibitors of Protein Kinases (2012).
Assuntos
Fenômenos Fisiológicos Celulares/fisiologia , Biossíntese de Proteínas , Transdução de Sinais/fisiologia , Serina-Treonina Quinases TOR/metabolismo , Animais , Núcleo Celular/metabolismo , Microambiente Celular/fisiologia , Humanos , Lisossomos/metabolismo , Modelos BiológicosRESUMO
The consolidation of learned information into long-lasting memories requires the strengthening of synaptic connections through de novo protein synthesis. Translation initiation factors play a cardinal role in gating the production of new proteins thereby regulating memory formation. Both positive and negative regulators of translation play a critical role in learning and memory consolidation. The eukaryotic initiation factor 4E (eIF4E) homologous protein (4EHP, encoded by the gene Eif4e2) is a pivotal negative regulator of translation but its role in learning and memory is unknown. To address this gap in knowledge, we generated excitatory (glutamatergic: CaMKIIα-positive) and inhibitory (GABAergic: GAD65-positive) conditional knockout mice for 4EHP, which were analyzed in various behavioral memory tasks. Knockout of 4EHP in Camk2a-expressing neurons (4EHP-cKOexc) did not impact long-term memory in either contextual fear conditioning or Morris water maze tasks. Similarly, long-term contextual fear memory was not altered in Gad2-directed 4EHP knockout mice (4EHP-cKOinh). However, when subjected to a short-term T-maze working memory task, both mouse models exhibited impaired cognition. We therefore tested the hypothesis that de novo protein synthesis plays a direct role in working memory. We discovered that phosphorylation of ribosomal protein S6, a measure of mTORC1 activity, is dramatically reduced in the CA1 hippocampus of 4EHP-cKOexc mice. Consistently, genetic reduction of mTORC1 activity in either excitatory or inhibitory neurons was sufficient to impair working memory. Taken together, these findings indicate that translational control by 4EHP and mTORC1 in both excitatory and inhibitory neurons are necessary for working memory.
Assuntos
Fator de Iniciação 4E em Eucariotos , Aprendizagem , Memória de Curto Prazo , Animais , Camundongos , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Camundongos Knockout , Proteínas de Ligação ao Cap de RNA/metabolismo , Fator de Iniciação 4E em Eucariotos/metabolismoRESUMO
Epileptogenesis is a process triggered by initial environmental or genetic factors that result in epilepsy and may continue during disease progression. Important parts of this process include changes in transcriptome and the pathological rewiring of neuronal circuits that involves changes in neuronal morphology. Mammalian/mechanistic target of rapamycin (mTOR) is upregulated by proconvulsive drugs, e.g., kainic acid, and is needed for progression of epileptogenesis, but molecular aspects of its contribution are not fully understood. Since mTOR can modulate transcription, we tested if rapamycin, an mTOR complex 1 inhibitor, affects kainic acid-evoked transcriptome changes. Using microarray technology, we showed that rapamycin inhibits the kainic acid-induced expression of multiple functionally heterogeneous genes. We further focused on engulfment and cell motility 1 (Elmo1), which is a modulator of actin dynamics and therefore could contribute to pathological rewiring of neuronal circuits during epileptogenesis. We showed that prolonged overexpression of Elmo1 in cultured hippocampal neurons increased axonal growth, decreased dendritic spine density, and affected their shape. In conclusion, data presented herein show that increased mTORC1 activity in response to kainic acid has no global effect on gene expression. Instead, our findings suggest that mTORC1 inhibition may affect development of epilepsy, by modulating expression of specific subset of genes, including Elmo1, and point to a potential role for Elmo1 in morphological changes that accompany epileptogenesis.
Assuntos
Proteínas de Transporte/metabolismo , Hipocampo/citologia , Ácido Caínico/farmacologia , Complexos Multiproteicos/metabolismo , Neurônios/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Animais , Axônios/efeitos dos fármacos , Axônios/metabolismo , Análise por Conglomerados , Espinhas Dendríticas/efeitos dos fármacos , Espinhas Dendríticas/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/efeitos dos fármacos , Masculino , Alvo Mecanístico do Complexo 1 de Rapamicina , Neurônios/efeitos dos fármacos , Ratos Wistar , Sirolimo/farmacologia , Transcrição Gênica/efeitos dos fármacosRESUMO
Autophagosomes primarily mediate turnover of cytoplasmic proteins or organelles to provide nutrients and eliminate damaged proteins. In neurons, autophagosomes form in distal axons and are trafficked retrogradely to fuse with lysosomes in the soma. Although defective neuronal autophagy is associated with neurodegeneration, the function of neuronal autophagosomes remains incompletely understood. We show that in neurons, autophagosomes promote neuronal complexity and prevent neurodegeneration in vivo via retrograde transport of brain-derived neurotrophic factor (BDNF)-activated TrkB receptors. p150Glued/dynactin-dependent transport of TrkB-containing autophagosomes requires their association with the endocytic adaptor AP-2, an essential protein complex previously thought to function exclusively in clathrin-mediated endocytosis. These data highlight a novel non-canonical function of AP-2 in retrograde transport of BDNF/TrkB-containing autophagosomes in neurons and reveal a causative link between autophagy and BDNF/TrkB signalling.
Assuntos
Complexo 2 de Proteínas Adaptadoras/metabolismo , Encéfalo/patologia , Receptor trkB/metabolismo , Animais , Autofagossomos , Autofagia , Transporte Biológico , Encéfalo/metabolismo , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Complexo Dinactina/metabolismo , Endocitose , Camundongos , Camundongos Knockout , Proteínas Associadas aos Microtúbulos/metabolismo , Neurônios/metabolismo , Ligação Proteica , Ratos Wistar , Transdução de SinaisRESUMO
Fragile X syndrome (FXS) is the leading monogenic cause of autism spectrum disorders (ASD). Trinucleotide repeat expansions in FMR1 abolish FMRP expression, leading to hyperactivation of ERK and mTOR signaling upstream of mRNA translation. Here we show that metformin, the most widely used drug for type 2 diabetes, rescues core phenotypes in Fmr1-/y mice and selectively normalizes ERK signaling, eIF4E phosphorylation and the expression of MMP-9. Thus, metformin is a potential FXS therapeutic.
Assuntos
Comportamento Animal/efeitos dos fármacos , Fator de Iniciação 4E em Eucariotos/efeitos dos fármacos , Proteína do X Frágil da Deficiência Intelectual/genética , Síndrome do Cromossomo X Frágil/genética , Hipoglicemiantes/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Metaloproteinase 9 da Matriz/efeitos dos fármacos , Metformina/farmacologia , Comportamento Social , Animais , Modelos Animais de Doenças , Fator de Iniciação 4E em Eucariotos/metabolismo , Síndrome do Cromossomo X Frágil/metabolismo , Síndrome do Cromossomo X Frágil/fisiopatologia , Masculino , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Camundongos Knockout , Fosforilação/efeitos dos fármacos , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/metabolismo , Expansão das Repetições de TrinucleotídeosRESUMO
Neurogenesis is the process of neuron generation, which occurs not only during embryonic development but also in restricted niches postnatally. One such region is called the subventricular zone (SVZ), which gives rise to new neurons in the olfactory bulb (OB). Neurons that are born postnatally migrate through more complex territories and integrate into fully functional circuits. Therefore, differences in the differentiation of embryonic and postnatally born neurons may exist. Dendritogenesis is an important process for the proper formation of future neuronal circuits. Dendritogenesis in embryonic neurons cultured in vitro was shown to depend on the mammalian target of rapamycin (mTOR). Still unknown, however, is whether mTOR could regulate the dendritic arbor morphology of SVZ-derived postnatal OB neurons under physiological conditions in vivo. The present study used in vitro cultured and differentiated SVZ-derived neural progenitors and found that both mTOR complex 1 and mTOR complex 2 were required for the dendritogenesis of SVZ-derived neurons. Furthermore, using a combination of in vivo electroporation of neural stem cells in the SVZ and genetic and pharmacological inhibition of mTOR, it was found that mTOR was crucial for the growth of basal and apical dendrites in postnatally born OB neurons under physiological conditions and contributed to the stabilization of their basal dendrites. © 2016 Wiley Periodicals, Inc. Develop Neurobiol 76: 1308-1327, 2016.
Assuntos
Células-Tronco Neurais/metabolismo , Neurogênese/fisiologia , Neurônios/metabolismo , Bulbo Olfatório/crescimento & desenvolvimento , Serina-Treonina Quinases TOR/metabolismo , Animais , Diferenciação Celular/fisiologia , Movimento Celular/fisiologia , Ventrículos Cerebrais/fisiologia , Camundongos , Células-Tronco Neurais/citologia , Neurônios/citologiaRESUMO
INTRODUCTION: Tuberous sclerosis complex (TSC) is a genetic disease resulting from mutation in TSC1 or TSC2 and subsequent hyperactivation of mammalian Target of Rapamycin (mTOR). Common TSC features include brain lesions, such as cortical tubers and subependymal giant cell astrocytomas (SEGAs). However, the current treatment with mTOR inhibitors has critical limitations. We aimed to identify new targets for TSC pharmacotherapy. RESULTS: The results of our shRNA screen point to glutamate-cysteine ligase catalytic subunit (GCLC), a key enzyme in glutathione synthesis, as a contributor to TSC-related phenotype. GCLC inhibition increased cellular stress and reduced mTOR hyperactivity in TSC2-depleted neurons and SEGA-derived cells. Moreover, patients' brain tubers showed elevated GCLC and stress markers expression. Finally, GCLC inhibition led to growth arrest and death of SEGA-derived cells. CONCLUSIONS: We describe GCLC as a part of redox adaptation in TSC, needed for overgrowth and survival of mutant cells, and provide a potential novel target for SEGA treatment.
Assuntos
Encéfalo/patologia , Glutamato-Cisteína Ligase/metabolismo , Neurônios/metabolismo , Esclerose Tuberosa/patologia , Adolescente , Animais , Butionina Sulfoximina/farmacologia , Células COS , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Criança , Chlorocebus aethiops , Inibidores Enzimáticos/farmacologia , Feminino , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Humanos , Imunossupressores/farmacologia , Masculino , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , RNA Interferente Pequeno/farmacologia , Sirolimo/farmacologia , Serina-Treonina Quinases TOR/metabolismo , Proteína 1 do Complexo Esclerose Tuberosa , Proteína 2 do Complexo Esclerose Tuberosa , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo , Adulto JovemRESUMO
Mammalian target of rapamycin (mTOR) is a protein kinase that senses nutrient availability, trophic factors support, cellular energy level, cellular stress, and neurotransmitters and adjusts cellular metabolism accordingly. Adequate mTOR activity is needed for development as well as proper physiology of mature neurons. Consequently, changes in mTOR activity are often observed in neuropathology. Recently, several groups reported that seizures increase mammalian target of rapamycin (mTOR) kinase activity, and such increased activity in genetic models can contribute to spontaneous seizures. However, the current knowledge about the spatiotemporal pattern of mTOR activation induced by proconvulsive agents is rather rudimentary. Also consequences of insufficient mTOR activity on a status epilepticus are poorly understood. Here, we systematically investigated these two issues. We showed that mTOR signaling was activated by kainic acid (KA)-induced status epilepticus through several brain areas, including the hippocampus and cortex as well as revealed two waves of mTOR activation: an early wave (2 h) that occurs in neurons and a late wave that predominantly occurs in astrocytes. Unexpectedly, we found that pretreatment with rapamycin, a potent mTOR inhibitor, gradually (i) sensitized animals to KA treatment and (ii) induced gross anatomical changes in the brain.