RESUMO
Fear discrimination is critical for survival, while fear generalization is effective for avoiding dangerous situations. Overgeneralized fear is a typical symptom of anxiety disorders, including generalized anxiety disorder and posttraumatic stress disorder (PTSD). Previous research demonstrated that fear discrimination learning is mediated by prefrontal mechanisms. While the prelimbic (PL) and infralimbic (IL) subdivisions of the medial prefrontal cortex (mPFC) are recognized for their excitatory and inhibitory effects on the fear circuit, respectively, the mechanisms driving fear discrimination are unidentified. To obtain insight into the mechanisms underlying context-specific fear discrimination, we investigated prefrontal neuronal ensembles representing distinct experiences associated with learning to disambiguate between dangerous and similar, but not identical, harmless stimuli. Here, we show distinct quantitative activation differences in response to conditioned and generalized fear experiences, as well as modulation of the neuronal ensembles associated with successful acquisition of context-safety contingencies. These findings suggest that prefrontal neuronal ensembles patterns code functional context-danger and context-safety relationships. The PL subdivision of the mPFC monitors context-danger associations to conditioned fear, whereas differential conditioning sparks additional ensembles associated with the inhibition of generalized fear in both the PL and IL subdivisions of the mPFC. Our data suggest that fear discrimination learning is associated with the modulation of prefrontal subpopulations in a subregion- and experience-specific fashion, and the learning of appropriate responses to conditioned and initially generalized fear experiences is driven by gradual updating and rebalancing of the prefrontal memory representations.
Assuntos
Condicionamento Clássico/fisiologia , Discriminação Psicológica/fisiologia , Medo/fisiologia , Neurônios/fisiologia , Córtex Pré-Frontal/citologia , Complexo Relacionado com a AIDS/genética , Complexo Relacionado com a AIDS/metabolismo , Análise de Variância , Animais , Doxiciclina/farmacologia , Reação de Congelamento Cataléptica/fisiologia , Expressão Gênica/efeitos dos fármacos , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Histonas/genética , Histonas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fosfopiruvato Hidratase/metabolismo , Fatores de TempoRESUMO
Perinatal neurodevelopment involves extensive formation of neural connections and onset of activity-dependent gene expression for synaptic function and plasticity. Exposure to psychostimulants at this stage imposes significant risks for developing cognitive and affective disorders later in life. However, how developmental exposure to psychostimulants may induce long-lasting molecular changes relevant to neural circuit function remains incompletely understood. In this study, we investigated the impacts of psychostimulant amphetamine on the activity-dependent induction of synaptic adaptor molecule Arc in the frontal cortex of neonatal mice. We found that transient exposure to amphetamine not only amplifies activity-dependent Arc mRNA expression immediately, but also potentiates subsequent induction of Arc mRNA in the absence of amphetamine. This priming effect is associated with a rapid and persistent increase in histone mono-methylation (H3K4me1), a marker for transcriptionally permissive chromatin, at the Arc locus, but not any long-lasting change in the phosphorylation of upstream transcription factor CREB. Furthermore, the increase in H3K4me1 at the Arc locus requires dopamine receptor signaling, and the priming of Arc expression correlates with the dopaminergic innervation pattern in the frontal cortex. Together, our results demonstrate that developmental exposure to psychostimulant amphetamine induces long-lasting chromatin changes and primes activity-dependent Arc gene induction. These findings reveal the molecular targets of psychostimulant during perinatal development that may contribute to long-term psychiatric risks.
Assuntos
Complexo Relacionado com a AIDS/genética , Estimulantes do Sistema Nervoso Central/farmacologia , Lobo Frontal/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Complexo Relacionado com a AIDS/metabolismo , Anfetamina/farmacologia , Análise de Variância , Animais , Benzazepinas/farmacologia , Proteína de Ligação a CREB/metabolismo , Imunoprecipitação da Cromatina , Dopamina/metabolismo , Estimulação Elétrica , Lobo Frontal/crescimento & desenvolvimento , Regulação da Expressão Gênica no Desenvolvimento/genética , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Fatores de TempoRESUMO
Negative affective aspects of opiate abstinence contribute to the persistence of substance abuse. Importantly, interconnected brain areas involved in aversive motivational processes, such as the ventral tegmental area (VTA) and medial prefrontal cortex (mPFC), become activated when animals are confined to withdrawal-paired environments. In the present study, place aversion was elicited in sham and adrenalectomized (ADX) animals by conditioned naloxone-precipitated drug withdrawal following exposure to chronic morphine. qPCR was employed to detect the expression of brain derived neurotrophic factor (Bdnf) and the immediate early genes (IEG) early growth response 1 (Egr-1) and activity-regulated cytoskeletal-associated protein (Arc) mRNAs in the VTA and mPFC at different time points of the conditioned place aversion (CPA) paradigm: after the conditioning phase and after the test phase. Sham + morphine rats exhibited robust CPA, which was impaired in ADX + morphine animals. Egr-1 and Arc were induced in the VTA and mPFC after morphine-withdrawal conditioning phase. Furthermore, Bdnf expression was enhanced in the VTA during the test phase. Bdnf induction seemed to be glucocorticoid-dependent, given that was correlated with HPA axis function and was not observed in morphine-dependent ADX animals. In addition, BDNF regulation and function was opposite in the VTA and mPFC during aversive-withdrawal memory retrieval. Our results suggest that IEGs and BDNF in these brain regions may play key roles in mediating the negative motivational component of opiate withdrawal.
Assuntos
Complexo Relacionado com a AIDS/metabolismo , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Encéfalo/metabolismo , Proteína 1 de Resposta de Crescimento Precoce/metabolismo , Glucocorticoides/metabolismo , Síndrome de Abstinência a Substâncias/patologia , Complexo Relacionado com a AIDS/genética , Adrenalectomia , Animais , Aprendizagem da Esquiva/efeitos dos fármacos , Fator Neurotrófico Derivado do Encéfalo/genética , Modelos Animais de Doenças , Proteína 1 de Resposta de Crescimento Precoce/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Glucocorticoides/farmacologia , Masculino , Morfina/efeitos adversos , Dependência de Morfina/complicações , Naloxona/uso terapêutico , Antagonistas de Entorpecentes/uso terapêutico , Entorpecentes/efeitos adversos , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Síndrome de Abstinência a Substâncias/tratamento farmacológico , Síndrome de Abstinência a Substâncias/etiologia , Síndrome de Abstinência a Substâncias/metabolismoRESUMO
Recent genetic tools have allowed researchers to visualize and manipulate memory traces (i.e., engrams) in small brain regions. However, the ultimate goal is to visualize memory traces across the entire brain in order to better understand how memories are stored in neural networks and how multiple memories may coexist. Intact tissue clearing and imaging is a new and rapidly growing area of focus that could accomplish this task. Here, we utilized the leading protocols for whole-brain clearing and applied them to the ArcCreERT2 mice, a murine line that allows for the indelible labeling of memory traces. We found that CLARITY and PACT greatly distorted the tissue, and iDISCO quenched enhanced yellow fluorescent protein (EYFP) fluorescence and hindered immunolabeling. Alternative clearing solutions, such as tert-Butanol, circumvented these harmful effects, but still did not permit whole-brain immunolabeling. CUBIC and CUBIC with Reagent-1A produced improved antibody penetration and preserved EYFP fluorescence, but also did not allow for whole-brain memory trace visualization. Modification of CUBIC with Reagent-1A resulted in EYFP fluorescence preservation and immunolabeling of the immediate early gene (IEG) Arc in deep brain areas; however, optimized memory trace labeling still required tissue slicing into mm-thick tissue sections. In summary, our data show that CUBIC with Reagent-1A* is the ideal method for reproducible clearing and immunolabeling for the visualization of memory traces in mm-thick tissue sections from ArcCreERT2 mice.
Assuntos
Complexo Relacionado com a AIDS/metabolismo , Encéfalo/metabolismo , Memória/fisiologia , Complexo Relacionado com a AIDS/genética , Animais , Encéfalo/anatomia & histologia , Channelrhodopsins/genética , Channelrhodopsins/metabolismo , Condicionamento Operante , Antagonistas de Estrogênios/farmacologia , Medo , Imuno-Histoquímica , Indicadores e Reagentes/farmacologia , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Camundongos , Camundongos Transgênicos , Microscopia Confocal , Receptores de Estrogênio/genética , Receptores de Estrogênio/metabolismo , Tamoxifeno/análogos & derivados , Tamoxifeno/farmacologiaRESUMO
AIM: Multifactors contribute to the development of postoperative cognitive dysfunction (POCD), of which the most important mechanism is neuroinflammation. Prostaglandin E2 (PGE2) is a key neuroinflammatory molecule and could modulate hippocampal synaptic transmission and plasticity. This study was designed to investigate whether PGE2 and its receptors signaling pathway were involved in the pathophysiology of POCD. METHODS: Sixteen-month old male C57BL/6J mice were exposed to laparotomy. Cognitive function was evaluated by fear conditioning test. The levels of PGE2 and its 4 distinct receptors (EP1-4) were assessed by biochemical analysis. Pharmacological or genetic methods were further applied to investigate the role of the specific PGE2 receptors. RESULTS: Here, we found that the transcription and translation level of the EP3 receptor in hippocampus increased remarkably, but not EP1, EP2, or EP4. Immunofluorescence results showed EP3 positive cells in the hippocampal CA1 region were mainly neurons. Furthermore, pharmacological blocking or genetic suppression of EP3 could alleviate surgery-induced hippocampus-dependent memory deficits and rescued the expression of plasticity-related proteins, including cAMP response element-binding protein (CREB), activity-regulated cytoskeletal-associated protein (Arc), and brain-derived neurotrophic factor (BDNF) in hippocampus. CONCLUSION: This study showed that PGE2-EP3 signaling pathway was involved in the progression of POCD and identified EP3 receptor as a promising treatment target.
Assuntos
Transtornos Cognitivos/etiologia , Transtornos Cognitivos/patologia , Dinoprostona/metabolismo , Regulação da Expressão Gênica/fisiologia , Hipocampo/metabolismo , Laparotomia/efeitos adversos , Detecção de Sinal Psicológico/fisiologia , Complexo Relacionado com a AIDS/genética , Complexo Relacionado com a AIDS/metabolismo , Envelhecimento , Animais , Fator Neurotrófico Derivado do Encéfalo/genética , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Proteína de Ligação a CREB/genética , Proteína de Ligação a CREB/metabolismo , Condicionamento Psicológico , Comportamento Exploratório , Medo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Complicações Pós-Operatórias/patologia , Complicações Pós-Operatórias/fisiopatologia , RNA Mensageiro/metabolismo , Receptores de Prostaglandina E Subtipo EP3/metabolismo , Transdução GenéticaRESUMO
BACKGROUND: As rehabilitation strategies advance as therapeutic interventions, the modality and onset of rehabilitation after traumatic brain injury (TBI) are critical to optimize treatment. Our laboratory has detected and characterized a late-onset, long-lasting sensory hypersensitivity to whisker stimulation in diffuse brain-injured rats; a deficit that is comparable to visual or auditory sensory hypersensitivity in humans with an acquired brain injury. OBJECTIVE: We hypothesize that the modality and onset of rehabilitation therapies will differentially influence sensory hypersensitivity in response to the Whisker Nuisance Task (WNT) as well as WNT-induced corticosterone (CORT) stress response in diffuse brain-injured rats and shams. METHODS: After midline fluid percussion brain injury (FPI) or sham surgery, rats were assigned to one of four rehabilitative interventions: (1) whisker sensory deprivation during week one or (2) week two or (3) whisker stimulation during week one or (4) week two. At 28 days following FPI and sham procedures, sensory hypersensitivity was assessed using the WNT. Plasma CORT was evaluated immediately following the WNT (aggravated levels) and prior to the pre-determined endpoint 24 hours later (non-aggravated levels). RESULTS: Deprivation therapy during week two elicited significantly greater sensory hypersensitivity to the WNT compared to week one (pâ<â0.05), and aggravated CORT levels in FPI rats were significantly lower than sham levels. Stimulation therapy during week one resulted in low levels of sensory hypersensitivity to the WNT, similar to deprivation therapy and naïve controls, however, non-aggravated CORT levels in FPI rats were significantly higher than sham. CONCLUSION: These data indicate that modality and onset of sensory rehabilitation can differentially influence FPI and sham rats, having a lasting impact on behavioral and stress responses to the WNT, emphasizing the necessity for continued evaluation of modality and onset of rehabilitation after TBI.
Assuntos
Lesões Encefálicas Traumáticas/fisiopatologia , Lesões Encefálicas Traumáticas/reabilitação , Condicionamento Físico Animal/métodos , Navegação Espacial/fisiologia , Vibrissas/inervação , Complexo Relacionado com a AIDS/genética , Complexo Relacionado com a AIDS/metabolismo , Análise de Variância , Animais , Lesões Encefálicas Traumáticas/sangue , Corticosterona/sangue , Modelos Animais de Doenças , Regulação da Expressão Gênica/fisiologia , Masculino , Estimulação Física/métodos , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Privação Sensorial/fisiologiaRESUMO
The activity-regulated cytoskeleton associated protein Arc is strongly and quickly upregulated by neuronal activity, synaptic potentiation and learning. Arc entry in the synapse is followed by the endocytosis of glutamatergic AMPA receptors (AMPARs), and its nuclear accumulation has been shown in vitro to result in a small decline in the transcription of the GluA1 subunit of AMPARs. Since these effects result in a decline in synaptic strength, we asked whether a change in Arc dynamics may temporally correlate with sleep-dependent GluA1 down-regulation. We measured the ratio of nuclear to cytoplasmic Arc expression (Arc Nuc/Cyto) in the cerebral cortex of EGFP-Arc transgenic mice that were awake most of the night and then perfused immediately before lights on (W mice), or were awake most of the night and then allowed to sleep (S mice) or sleep deprived (SD mice) for the first 2 h of the light phase. In primary motor cortex (M1), neurons with high levels of nuclear Arc (High Arc cells) were present in all mice, but in these cells Arc Nuc/Cyto was higher in S mice than in W mice and, importantly, ~15% higher in S mice than in SD mice collected at the same time of day, ruling out circadian effects. Greater Arc Nuc/Cyto with sleep was observed in the superficial layers of M1, but not in the deep layers. In High Arc cells, Arc Nuc/Cyto was also ~15%-30% higher in S mice than in W and SD mice in the superficial layers of primary somatosensory cortex (S1) and cingulate cortex area 1 (Cg1). In High Arc Cells of Cg1, Arc Nuc/Cyto and cytoplasmic levels of GluA1 immunoreactivities in the soma were also negatively correlated, independent of behavioral state. Thus, Arc moves to the nucleus during both sleep and wake, but its nuclear to cytoplasmic ratio increases with sleep in the superficial layers of several cortical areas. It remains to be determined whether the relative increase in nuclear Arc contributes significantly to the overall decline in the strength of excitatory synapses that occurs during sleep. Similarly, it remains to be determined whether the entry of Arc into specific synapses is gated by sleep.
Assuntos
Complexo Relacionado com a AIDS/metabolismo , Núcleo Celular/metabolismo , Córtex Cerebral/citologia , Citoplasma/metabolismo , Neurônios/ultraestrutura , Sono/fisiologia , Complexo Relacionado com a AIDS/genética , Animais , Carbocianinas/metabolismo , Feminino , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Masculino , Camundongos , Camundongos Transgênicos , Microscopia Confocal , Neurônios/metabolismo , Receptores de AMPA/metabolismo , Vigília/fisiologiaRESUMO
Rodents require a minimal time period to explore a context prior to footshock to display plateau-level context fear at test. To investigate whether this rapid fear plateau reflects complete memory formation within that short time-frame, we used the immediate-early gene product Arc as an indicator of hippocampal context memory formation-related activity. We found that hippocampal Arc expression continued to increase well past the minimal time required for plateau-level fear. This raises the possibility that context fear conditioning occurs more rapidly than complete memory formation. Thus, animals may be able to condition robustly to both complete and incomplete contextual representations.
Assuntos
Medo , Hipocampo/metabolismo , Memória/fisiologia , Complexo Relacionado com a AIDS/genética , Complexo Relacionado com a AIDS/metabolismo , Animais , Condicionamento Clássico , Eletrochoque/efeitos adversos , Reação de Congelamento Cataléptica/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , RNA Mensageiro/metabolismo , Fatores de TempoRESUMO
The activity-regulated gene Arc/Arg3.1 encodes a postsynaptic protein crucially involved in glutamatergic synaptic plasticity. Genetic mutations in Arc pathway and altered Arc expression in human frontal cortex have been associated with schizophrenia. Although Arc expression has been reported to vary with age, what mechanisms regulate Arc mRNA levels in frontal cortex during postnatal development remains unclear. Using quantitative mRNA analysis of mouse frontal cortical tissues, we mapped the developmental profiles of Arc expression and found that its mRNA levels are sharply amplified near the end of the second postnatal week, when mouse pups open their eyes for the first time after birth. Surprisingly, electrical stimulation of the frontal cortex before eye-opening is not sufficient to drive the amplification of Arc mRNA. Instead, this amplification needs both electrical stimulation and dopamine D1-type receptor (D1R) activation. Furthermore, visual stimuli-driven amplification of Arc mRNA is also dependent on D1R activation and dopamine neurons located in the ventral midbrain. These results indicate that dopamine is required to drive activity-dependent amplification of Arc mRNA in the developing postnatal frontal cortex and suggest that joint electrical and dopaminergic activation is essential to establish the normal expression pattern of a schizophrenia-associated gene during frontal cortical development.
Assuntos
Complexo Relacionado com a AIDS/genética , Dopamina/metabolismo , Lobo Frontal/crescimento & desenvolvimento , Lobo Frontal/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , RNA Mensageiro/metabolismo , Complexo Relacionado com a AIDS/metabolismo , Adrenérgicos/farmacologia , Fatores Etários , Anfetamina/farmacologia , Animais , Animais Recém-Nascidos , Antioxidantes/farmacologia , Ácido Ascórbico/farmacologia , Benzazepinas/farmacologia , Dopaminérgicos/farmacologia , Neurônios Dopaminérgicos/efeitos dos fármacos , Eletrochoque/métodos , Camundongos , Camundongos Endogâmicos C57BL , Análise em Microsséries , Oxidopamina/farmacologia , Receptores de Dopamina D1/metabolismo , Área Tegmentar Ventral/citologiaRESUMO
There is limited knowledge regarding how the brain controls the timing of meals. Similarly, there is a large gap in our understanding of how top-down cognitive processes, such as memory influence energy intake. We hypothesize that dorsal hippocampal (dHC) neurons, which are critical for episodic memory, form a memory of a meal and inhibit meal onset during the postprandial period. In support, we showed previously that reversible inactivation of these neurons during the period following a sucrose meal accelerates the onset of the next meal. If dHC neurons form a memory of a meal, then consumption should induce synaptic plasticity in dHC neurons. To test this, we determined (1) whether a sucrose meal increases the expression of the synaptic plasticity marker activity-regulated cytoskeleton-associated protein (Arc) in dHC CA1 neurons, (2) whether previous experience with sucrose influences sucrose-induced Arc expression, and (3) whether the orosensory stimulation produced by the noncaloric sweetener saccharin is sufficient to induce Arc expression. Male Sprague-Dawley rats were trained to consume a sweetened solution at a scheduled time daily. On the experimental day, they were given a solution for 7 min, euthanized, and then fluorescence in situ hybridization procedures were used to measure meal-induced Arc mRNA. Compared to caged control rats, Arc expression was significantly higher in rats that consumed sucrose or saccharin. Interestingly, rats given additional experience with sucrose had less Arc expression than rats with less sucrose experience, even though both groups consumed similar amounts on the experimental day. Thus, this study is the first to suggest that orosensory stimulation produced by consuming a sweetened solution and possibly the hedonic value of that sweet stimulation induces synaptic plasticity in dHC CA1 neurons in an experience-dependent manner. Collectively, these findings are consistent with our hypothesis that dHC neurons form a memory of a meal.
Assuntos
Complexo Relacionado com a AIDS/metabolismo , Região CA1 Hipocampal/citologia , Neurônios/efeitos dos fármacos , Edulcorantes/farmacologia , Paladar/fisiologia , Complexo Relacionado com a AIDS/genética , Vias Aferentes/efeitos dos fármacos , Vias Aferentes/fisiologia , Análise de Variância , Animais , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/fisiologia , Masculino , Neurônios/fisiologia , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Sacarina/farmacologia , Sacarose/farmacologia , Paladar/efeitos dos fármacos , Ensino , Fatores de TempoRESUMO
Synaptic plasticity is a property of neurons that can be induced by conditioning electrical stimulation (CS) of afferent fibers in the spinal cord. This is a widely studied property of spinal cord and hippocampal neurons. CS has been shown to trigger enhanced expression of immediate early gene proteins (IEGPs), with peak increases observed 2 hour post stimulation. Chronic morphine treatment has been shown to promoteinduce opioid-induced hyperalgesia, and also to increase CS-induced central sensitization in the dorsal horn. As IEGP expression may contribute to development of chronic pain states, we aimed to determine whether chronic morphine treatment affects the expression of IEGPs following sciatic nerve CS. Changes in expression of the IEGPs Arc, c-Fos or Zif268 were determined in cells of the lumbar dorsal horn of the spinal cord. Chronic Morphine pretreatment over 7 days led to a significant increase in the number of IEGP positive cells observed at both 2 h and 6 h after CS. The same pattern of immunoreactivity was obtained for all IEGPs, with peak increases occurring at 2 h post CS. In contrast, morphine treatment alone in sham operated animals had no effect on IEGP expression. We conclude that chronic morphine treatment enhances stimulus-induced expression of IEGPs in the lumbar dorsal horn. These data support the notion that morphine alters gene expression responses linked to nociceptive stimulation and plasticity.
Assuntos
Analgésicos Opioides/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas Imediatamente Precoces/metabolismo , Morfina/farmacologia , Nervo Isquiático/fisiologia , Corno Dorsal da Medula Espinal/efeitos dos fármacos , Complexo Relacionado com a AIDS/genética , Complexo Relacionado com a AIDS/metabolismo , Vias Aferentes/fisiologia , Análise de Variância , Animais , Proteína 1 de Resposta de Crescimento Precoce/genética , Proteína 1 de Resposta de Crescimento Precoce/metabolismo , Estimulação Elétrica , Feminino , Proteínas Imediatamente Precoces/genética , Proteínas Proto-Oncogênicas c-fos/metabolismo , Ratos , Ratos Sprague-Dawley , Corno Dorsal da Medula Espinal/metabolismo , Fatores de TempoRESUMO
Treatment of Parkinson's disease (PD) with dopamine replacement relieves symptoms of poverty of movement, but often causes drug-induced dyskinesias. Accumulating clinical and pre-clinical evidence suggests that the primary motor cortex (M1) is involved in the pathophysiology of PD and that modulating cortical activity may be a therapeutic target in PD and dyskinesia. However, surprisingly little is known about how M1 neurotransmitter tone or gene expression is altered in PD, dyskinesia or associated animal models. The present study utilized the rat unilateral 6-hydroxydopamine (6-OHDA) model of PD/dyskinesia to characterize structural and functional changes taking place in M1 monoamine innervation and gene expression. 6-OHDA caused dopamine pathology in M1, although the lesion was less severe than in the striatum. Rats with 6-OHDA lesions showed a PD motor impairment and developed dyskinesia when given L-DOPA or the D1 receptor agonist, SKF81297. M1 expression of two immediate-early genes (c-Fos and ARC) was strongly enhanced by either L-DOPA or SKF81297. At the same time, expression of genes specifically involved in glutamate and GABA signaling were either modestly affected or unchanged by lesion and/or treatment. We conclude that M1 neurotransmission and signal transduction in the rat 6-OHDA model of PD/dyskinesia mirror features of human PD, supporting the utility of the model to study M1 dysfunction in PD and the elucidation of novel pathophysiological mechanisms and therapeutic targets.
Assuntos
Complexo Relacionado com a AIDS/metabolismo , Discinesia Induzida por Medicamentos/patologia , Expressão Gênica/fisiologia , Córtex Motor/metabolismo , Proteínas Proto-Oncogênicas c-fos/metabolismo , Transmissão Sináptica/fisiologia , Complexo Relacionado com a AIDS/genética , Animais , Benzazepinas/efeitos adversos , Modelos Animais de Doenças , Dopaminérgicos/efeitos adversos , Discinesia Induzida por Medicamentos/etiologia , Lateralidade Funcional/efeitos dos fármacos , Expressão Gênica/efeitos dos fármacos , Levodopa/uso terapêutico , Masculino , Feixe Prosencefálico Mediano/lesões , Oxidopamina/toxicidade , Transtornos Parkinsonianos/induzido quimicamente , Transtornos Parkinsonianos/tratamento farmacológico , Proteínas Proto-Oncogênicas c-fos/genética , Ratos , Ratos Sprague-Dawley , Índice de Gravidade de Doença , Transmissão Sináptica/efeitos dos fármacosRESUMO
Antipsychotics may modulate the transcription of multiple gene programs, including those belonging to postsynaptic density (PSD) network, within cortical and subcortical brain regions. Understanding which brain region is activated progressively by increasing doses of antipsychotics and how their different receptor profiles may impact such an activation could be relevant to better correlate the mechanism of action of antipsychotics both with their efficacy and side effects. We analyzed the differential topography of PSD transcripts by incremental doses of two antipsychotics: haloperidol, the prototypical first generation antipsychotic with prevalent dopamine D2 receptors antagonism, and asenapine, a second generation antipsychotic characterized by multiple receptors occupancy. We investigated the expression of PSD genes involved in synaptic plasticity and previously demonstrated to be modulated by antipsychotics: Homer1a, and its related interacting constitutive genes Homer1b/c and PSD95, as well as Arc, C-fos and Zif-268, also known to be induced by antipsychotics administration. We found that increasing acute doses of haloperidol induced immediate-early genes (IEGs) expression in different striatal areas, which were progressively recruited by incremental doses with a dorsal-to-ventral gradient of expression. Conversely, increasing acute asenapine doses progressively de-recruited IEGs expression in cortical areas and increased striatal genes signal intensity. These effects were mirrored by a progressive reduction in locomotor animal activity by haloperidol, and an opposite increase by asenapine. Thus, we demonstrated for the first time that antipsychotics may progressively recruit PSD-related IEGs expression in cortical and subcortical areas when administered at incremental doses and these effects may reflect a fine-tuned dose-dependent modulation of the PSD.
Assuntos
Antipsicóticos/farmacologia , Córtex Cerebral/efeitos dos fármacos , Corpo Estriado/efeitos dos fármacos , Densidade Pós-Sináptica/efeitos dos fármacos , Complexo Relacionado com a AIDS/genética , Complexo Relacionado com a AIDS/metabolismo , Análise de Variância , Animais , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Córtex Cerebral/metabolismo , Corpo Estriado/metabolismo , Proteína 4 Homóloga a Disks-Large , Relação Dose-Resposta a Droga , Proteína 1 de Resposta de Crescimento Precoce/genética , Proteína 1 de Resposta de Crescimento Precoce/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas de Arcabouço Homer , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Atividade Motora/efeitos dos fármacos , Proteínas Oncogênicas v-fos/genética , Proteínas Oncogênicas v-fos/metabolismo , Densidade Pós-Sináptica/metabolismo , Ligação Proteica/efeitos dos fármacos , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Estatísticas não ParamétricasRESUMO
Activation of microglia may facilitate age-related impairment in cognitive functions including hippocampal-dependent memory. Considerable evidence indicates that hippocampal-derived estrogen improves hippocampal-dependent learning and memory. We hypothesize that activated microglia may inhibit de novo hippocampal estrogen synthesis and in turn suppress hippocampal synaptic protein expression. The present study aimed to elucidate the role of lipopolysaccharide (LPS)-activated microglial HAPI cells on estrogen synthesis and expression of synaptic proteins using H19-7 hippocampal neurons with a neuron-microglia co-culture system. LPS induced expression of the microglial activation markers major histocompatibility complex II (MHC II), CD11b, and ionized calcium-binding adapter molecule 1 (Iba1). Prolonged LPS exposure also enhanced the secretion of interleukin (IL)-6 and nitric oxide (NO) from microglial HAPI cells. Exposure to either LPS-activated microglia or IL-6, significantly suppressed the expression of synaptic proteins and the secretion of de novo hippocampal estrogen in H19-7 hippocampal neurons. In addition, LPS-activated microglia also decreased the expression of estrogen receptors (ERα and ERß) in H19-7 hippocampal neurons. Our findings demonstrate a potential mechanism of microglia activation underlying the reduction in estrogen-mediated signaling on synaptic proteins in hippocampal neurons, which may be involved in hippocampal-dependent memory formation.
Assuntos
Regulação para Baixo/fisiologia , Estrogênios/metabolismo , Hipocampo/citologia , Microglia/metabolismo , Neurônios/metabolismo , Complexo Relacionado com a AIDS/genética , Complexo Relacionado com a AIDS/metabolismo , Análise de Variância , Linhagem Celular Transformada , Técnicas de Cocultura , Proteína 4 Homóloga a Disks-Large , Relação Dose-Resposta a Droga , Regulação para Baixo/efeitos dos fármacos , Ensaio de Imunoadsorção Enzimática , Estrogênios/genética , Guanilato Quinases/metabolismo , Humanos , Interleucina-10/genética , Interleucina-10/metabolismo , Interleucina-6/genética , Interleucina-6/metabolismo , Lipopolissacarídeos/farmacologia , Proteínas de Membrana/metabolismo , Microglia/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , RNA Mensageiro/metabolismo , Receptores de Estrogênio/genética , Receptores de Estrogênio/metabolismo , Sinaptofisina/genética , Sinaptofisina/metabolismo , Fatores de TempoRESUMO
The ability to visualize behaviourally evoked neural activity patterns across the rodent brain is essential for understanding the distributed brain networks mediating particular behaviours. However, current imaging methods are limited in their spatial resolution and/or ability to obtain brain-wide coverage of functional activity. Here, we describe a new automated method for obtaining cellular-level, whole-brain maps of behaviourally induced neural activity in the mouse. This method combines the use of transgenic immediate-early gene reporter mice to visualize neural activity; serial two-photon tomography to image the entire brain at cellular resolution; advanced image processing algorithms to count the activated neurons and align the datasets to the Allen Mouse Brain Atlas; and statistical analysis to identify the network of activated brain regions evoked by behaviour. We demonstrate the use of this approach to determine the whole-brain networks activated during the retrieval of fear memories. Consistent with previous studies, we identified a large network of amygdalar, hippocampal, and neocortical brain regions implicated in fear memory retrieval. Our proposed methods can thus be used to map cellular networks involved in the expression of normal behaviours as well as to investigate in depth circuit dysfunction in mouse models of neurobiological disease.
Assuntos
Mapeamento Encefálico , Encéfalo/fisiologia , Condicionamento Psicológico/fisiologia , Medo/fisiologia , Neurônios/fisiologia , Complexo Relacionado com a AIDS/genética , Complexo Relacionado com a AIDS/metabolismo , Animais , Benzofuranos , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Camundongos , Camundongos Transgênicos , Quinolinas , Estatísticas não ParamétricasRESUMO
In spite of the fact that cholesterol does not pass the blood-brain barrier, treatment of mice with dietary cholesterol causes significant effects on a number of genes in the brain and in addition a memory impairment. We have suggested that these effects are mediated by 27-hydroxycholesterol, which is able to pass the blood-brain barrier. To test this hypothesis we utilized Cyp27-/- mice lacking 27-hydroxycholesterol. The negative effect on memory observed after treatment of wildtype mice with dietary cholesterol was not observed in these mice. The cholesterol diet reduced the levels of the "memory protein" Arc (Activity Regulated Cytoskeleton associated protein) in the hippocampus of the wildtype mice but not in the hippocampus of the Cyp27-/- mice. The results are consistent with 27-hydroxycholesterol as the mediator of the negative effects of cholesterol on cognition.
Assuntos
Colesterol/efeitos adversos , Transtornos Cognitivos/etiologia , Transtornos Cognitivos/genética , Hidroxicolesteróis/uso terapêutico , Complexo Relacionado com a AIDS/genética , Complexo Relacionado com a AIDS/metabolismo , Animais , Peso Corporal/efeitos dos fármacos , Peso Corporal/genética , Colestanotriol 26-Mono-Oxigenase/deficiência , Colestanotriol 26-Mono-Oxigenase/genética , Ácido Cólico/administração & dosagem , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Masculino , Aprendizagem em Labirinto/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , RNA Mensageiro/metabolismo , Aprendizagem Espacial/efeitos dos fármacosRESUMO
The induction of hypoxia-inducible factors (HIFs) is essential for the adaptation of tumor cells to a low-oxygen environment. We found that the expression of the apoptosis inhibitor ARC (apoptosis repressor with a CARD domain) was induced by hypoxia in a variety of cancer cell types, and its induction is primarily HIF1 dependent. Chromatin immunoprecipitation (ChIP) and reporter assays also indicate that the ARC gene is regulated by direct binding of HIF1 to a hypoxia response element (HRE) located at bp -190 upstream of the transcription start site. HIFs play an essential role in the pathogenesis of renal cell carcinoma (RCC) under normoxic conditions, through the loss of the Von Hippel-Lindau gene (VHL). Accordingly, our results show that ARC is not expressed in normal renal tissue but is highly expressed in 65% of RCC tumors, which also express high levels of carbonic anhydrase IX (CAIX), a HIF1-dependent protein. Compared to controls, ARC-deficient RCCs exhibited decreased colony formation and increased apoptosis in vitro. In addition, loss of ARC resulted in a dramatic reduction of RCC tumor growth in SCID mice in vivo. Thus, HIF-mediated increased expression of ARC in RCC can explain how loss of VHL can promote survival early in tumor formation.
Assuntos
Apoptose/fisiologia , Carcinoma de Células Renais/metabolismo , Regulação Neoplásica da Expressão Gênica/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Neoplasias Renais/metabolismo , Proteínas Supressoras de Tumor/genética , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Complexo Relacionado com a AIDS/genética , Animais , Apoptose/genética , Carcinoma de Células Renais/genética , Hipóxia Celular/genética , Hipóxia Celular/fisiologia , Proliferação de Células , Sobrevivência Celular , Células Cultivadas , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Neoplasias Renais/genética , Neoplasias Renais/patologia , Camundongos , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Proteína Supressora de Tumor Von Hippel-Lindau/metabolismoRESUMO
Histone deactylase (HDAC) inhibitors show promise as therapeutics for neurodegenerative and psychiatric diseases. Increased expression of brain-derived neurotrophic factor (BDNF) has been associated with memory-enhancing and neuroprotective properties of these drugs, but the mechanism of BDNF induction is not well understood. Here, we compared the effects of a class I/IIb selective HDAC inhibitor SAHA, a class I selective inhibitor MS-275, a class II selective inhibitor MC1568 and a HDAC6 selective inhibitor tubacin on Bdnf mRNA expression in rat primary neurons. We show that inhibition of class II HDACs resulted in rapid upregulation of Bdnf mRNA levels, whereas class I HDAC inhibition produced a markedly delayed Bdnf induction. In contrast to relatively slow upregulation of Bdnf transcripts, histone acetylation at BDNF promoters I and IV was rapidly induced by SAHA. Bdnf induction by SAHA and MS-275 at 24 h was sensitive to protein synthesis inhibition, suggesting that delayed Bdnf induction by HDAC inhibitors is secondary to changed expression of its regulators. HDAC4 and HDAC5 repressed Bdnf promoter IV activity, supporting the role of class II HDACs in regulation of Bdnf expression. In addition, we show a critical role for the cAMP/Ca2+ response element (CRE) in induction of Bdnf promoter IV by MS-275, MC1568, SAHA and sodium valproate. In contrast, MEF2-binding CaRE1 element was not necessary for promoter IV induction by HDAC inhibition. Finally, we show that similarly to Bdnf, the studied HDAC inhibitors differentially induced expression of neuronal activity-regulated genes c-fos and Arc. Together, our findings implicate class II HDACs in transcriptional regulation of Bdnf and indicate that class II selective HDAC inhibitors may have potential as therapeutics for nervous system disorders.
Assuntos
Fator Neurotrófico Derivado do Encéfalo/metabolismo , Córtex Cerebral/citologia , Regulação da Expressão Gênica/efeitos dos fármacos , Inibidores de Histona Desacetilases/farmacologia , Neurônios/efeitos dos fármacos , Complexo Relacionado com a AIDS/genética , Complexo Relacionado com a AIDS/metabolismo , Animais , Animais Recém-Nascidos , Benzamidas/farmacologia , Fator Neurotrófico Derivado do Encéfalo/genética , Células Cultivadas , Cicloeximida/farmacologia , Relação Dose-Resposta a Droga , Histona Desacetilases/genética , Histona Desacetilases/metabolismo , Humanos , Ácidos Hidroxâmicos/farmacologia , Proteínas Proto-Oncogênicas c-fos/genética , Proteínas Proto-Oncogênicas c-fos/metabolismo , Piridinas/farmacologia , Pirróis/farmacologia , Ratos , Ratos Sprague-Dawley , Fatores de TempoRESUMO
Prefrontal serotonin 2A receptors (5-HT2A Rs) have been linked to the pathogenesis and treatment of schizophrenia. Many antipsychotics fully occupy 5-HT2A R at clinical relevant doses, and activation of 5-HT2A receptors by lysergic acid diethylamide (LSD) and LSD-like drugs induces a schizophrenia-like psychosis in humans. Subchronic phencyclidine (PCP) administration is a well-established model for schizophrenia-like symptoms in rodents. The aim of the present study was to investigate whether subchronic PCP administration changes expression, binding, or functionality of cortical 5-HT2A Rs. As a measure of 5-HT2A R functionality, we used the 5-HT2A R agonist 2,5-dimethoxy-4-iodoamphetamine (DOI)-induced head-twitch response (HTR) and mRNA expression of the immediate-early genes (IEGs) activity-related cytoskeletal associated-protein (Arc), c-fos, and early growth response protein 2 (egr-2) in the frontal cortex. Mice were treated with PCP (10 mg/kg) or saline for 10 days, followed by a 5-day washout period. The PCP pretreatment increased the overall induction of HTR and frontal cortex IEG mRNA expression following a single challenge with DOI. These functional changes were not associated with changes in 5-HT2A R binding. Also, binding of the 5-HT1A R and the 5-HT transporter was unaffected. Finally, basal mRNA level of Arc was increased in the prefrontal cortex after subchronic PCP administration as revealed with in situ hybridization. Together these findings indicate that PCP administration produces changes in the brain that result in an increase in the absolute effect of DOI. Therefore, neurotransmission involving the 5-HT2A R could contribute to the behavioral deficits observed after PCP treatment. © 2013 Wiley Periodicals, Inc.
Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Fenciclidina/farmacologia , Córtex Pré-Frontal/efeitos dos fármacos , Receptores 5-HT2 de Serotonina/metabolismo , Esquizofrenia/induzido quimicamente , Esquizofrenia/patologia , Transdução de Sinais/efeitos dos fármacos , Complexo Relacionado com a AIDS/genética , Complexo Relacionado com a AIDS/metabolismo , Anfetaminas/farmacologia , Análise de Variância , Animais , Autorradiografia , Modelos Animais de Doenças , Regulação da Expressão Gênica/fisiologia , Genes Precoces/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fenciclidina/toxicidade , Córtex Pré-Frontal/metabolismo , Proteínas Proto-Oncogênicas c-fos/genética , Proteínas Proto-Oncogênicas c-fos/metabolismo , RNA Mensageiro/metabolismo , Agonistas do Receptor de Serotonina/farmacologia , Fatores de TempoRESUMO
A small number of rare, recurrent genomic copy number variants (CNVs) are known to substantially increase susceptibility to schizophrenia. As a consequence of the low fecundity in people with schizophrenia and other neurodevelopmental phenotypes to which these CNVs contribute, CNVs with large effects on risk are likely to be rapidly removed from the population by natural selection. Accordingly, such CNVs must frequently occur as recurrent de novo mutations. In a sample of 662 schizophrenia proband-parent trios, we found that rare de novo CNV mutations were significantly more frequent in cases (5.1% all cases, 5.5% family history negative) compared with 2.2% among 2623 controls, confirming the involvement of de novo CNVs in the pathogenesis of schizophrenia. Eight de novo CNVs occurred at four known schizophrenia loci (3q29, 15q11.2, 15q13.3 and 16p11.2). De novo CNVs of known pathogenic significance in other genomic disorders were also observed, including deletion at the TAR (thrombocytopenia absent radius) region on 1q21.1 and duplication at the WBS (Williams-Beuren syndrome) region at 7q11.23. Multiple de novos spanned genes encoding members of the DLG (discs large) family of membrane-associated guanylate kinases (MAGUKs) that are components of the postsynaptic density (PSD). Two de novos also affected EHMT1, a histone methyl transferase known to directly regulate DLG family members. Using a systems biology approach and merging novel CNV and proteomics data sets, systematic analysis of synaptic protein complexes showed that, compared with control CNVs, case de novos were significantly enriched for the PSD proteome (P=1.72 × 10â»6. This was largely explained by enrichment for members of the N-methyl-D-aspartate receptor (NMDAR) (P=4.24 × 10â»6) and neuronal activity-regulated cytoskeleton-associated protein (ARC) (P=3.78 × 10â»8) postsynaptic signalling complexes. In an analysis of 18 492 subjects (7907 cases and 10 585 controls), case CNVs were enriched for members of the NMDAR complex (P=0.0015) but not ARC (P=0.14). Our data indicate that defects in NMDAR postsynaptic signalling and, possibly, ARC complexes, which are known to be important in synaptic plasticity and cognition, play a significant role in the pathogenesis of schizophrenia.