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1.
Vet Res ; 52(1): 53, 2021 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-33823911

RESUMO

Trypanosoma cruzi is a zoonotic parasite endemic in the southern US and the Americas, which may frequently infect dogs, but limited information is available about infections in cats. We surveyed a convenience sample of 284 shelter cats from Southern Louisiana to evaluate T. cruzi infection using serological and PCR tests. Parasites from PCR positive cats were also genotyped by PCR and deep sequencing to assess their genetic diversity. We detected a seropositivity rate for T. cruzi of at least 7.3% (17/234), and 24.6% of cats (70/284) were PCR positive for the parasite. Seropositivity increased with cat age (R2 = 0.91, P = 0.011), corresponding to an incidence of 7.2% ± 1.3 per year, while PCR positivity decreased with age (R2 = 0.93, P = 0.007). Cats were predominantly infected with parasites from TcI and TcVI DTUs, and to a lesser extent from TcIV and TcV DTUs, in agreement with the circulation of these parasite DTUs in local transmission cycles. These results indicate that veterinarians should have a greater awareness of T. cruzi infection in pets and that it would be important to better evaluate the risk for spillover infections in humans.

2.
PLoS Negl Trop Dis ; 14(12): e0008932, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33332357

RESUMO

BACKGROUND: Chagas disease is a neglected zoonosis of growing concern in the southern US, caused by the parasite Trypanosoma cruzi. We genotyped parasites in a large cohort of PCR positive dogs to shed light on parasite transmission cycles and assess potential relationships between parasite diversity and serological test performance. METHODOLOGY/PRINCIPAL FINDINGS: We used a metabarcoding approach based on deep sequencing of T. cruzi mini-exon marker to assess parasite diversity. Phylogenetic analysis of 178 sequences from 40 dogs confirmed the presence of T. cruzi discrete typing unit (DTU) TcI and TcIV, as well as TcII, TcV and TcVI for the first time in US dogs. Infections with multiple DTUs occurred in 38% of the dogs. These data indicate a greater genetic diversity of T. cruzi than previously detected in the US. Comparison of T. cruzi sequence diversity indicated that highly similar T. cruzi strains from these DTUs circulate in hosts and vectors in Louisiana, indicating that they are involved in a shared T. cruzi parasite transmission cycle. However, TcIV and TcV were sampled more frequently in vectors, while TcII and TcVI were sampled more frequently in dogs. CONCLUSIONS/SIGNIFICANCE: These observations point to ecological host-fitting being a dominant mechanism involved in the diversification of T. cruzi-host associations. Dogs with negative, discordant or confirmed positive T. cruzi serology harbored TcI parasites with different mini-exon sequences, which strongly supports the hypothesis that parasite genetic diversity is a key factor affecting serological test performance. Thus, the identification of conserved parasite antigens should be a high priority for the improvement of current serological tests.


Assuntos
Doença de Chagas/veterinária , Éxons/genética , Variação Genética , Trypanosoma cruzi/genética , Animais , Doença de Chagas/epidemiologia , Doença de Chagas/parasitologia , Doença de Chagas/transmissão , Estudos de Coortes , Cães , Genótipo , Humanos , Louisiana/epidemiologia , Filogenia , Testes Sorológicos/veterinária , Trypanosoma cruzi/imunologia , Trypanosoma cruzi/fisiologia , Zoonoses
3.
Parasit Vectors ; 13(1): 577, 2020 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-33189151

RESUMO

BACKGROUND: Trypanosoma cruzi - the causative agent of Chagas disease - is known to circulate in commensal pests, but its occurrence in urban environments is not well understood. We addressed this deficit by determining the distribution and prevalence of T. cruzi infection in urban populations of commensal and wild rodents across New Orleans (Louisiana, USA). We assessed whether T. cruzi prevalence varies according to host species identity and species co-occurrences, and whether T. cruzi prevalence varies across mosaics of abandonment that shape urban rodent demography and assemblage structure in the city. METHODS: Leveraging city-wide population and assemblage surveys, we tested 1428 rodents comprising 5 species (cotton rats, house mice, Norway rats, rice rats and roof rats) captured at 98 trapping sites in 11 study areas across New Orleans including nine residential neighborhoods and a natural area in Orleans Parish and a neighborhood in St. Bernard Parish. We also assayed Norway rats at one site in Baton Rouge (Louisiana, USA). We used chi-square tests to determine whether infection prevalence differed among host species, among study areas, and among trapping sites according to the number of host species present. We used generalized linear mixed models to identify predictors of T. cruzi infection for all rodents and each host species, respectively. RESULTS: We detected T. cruzi in all host species in all study areas in New Orleans, but not in Baton Rouge. Though overall infection prevalence was 11%, it varied by study area and trapping site. There was no difference in prevalence by species, but roof rats exhibited the broadest geographical distribution of infection across the city. Infected rodents were trapped in densely populated neighborhoods like the French Quarter. Infection prevalence seasonally varied with abandonment, increasing with greater abandonment during the summer and declining with greater abandonment during the winter. CONCLUSIONS: Our findings illustrate that T. cruzi can be widespread in urban landscapes, suggesting that transmission and disease risk is greater than is currently recognized. Our findings also suggest that there is disproportionate risk of transmission in historically underserved communities, which could reinforce long-standing socioecological disparities in New Orleans and elsewhere.

4.
Genome ; 2020 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-33086026

RESUMO

Chagas disease is a zoonotic, parasitic, vector-borne neglected tropical disease that affects the lives of over 6 million people throughout the Americas. Trypanosoma cruzi, the causative agent, presents extensive genetic diversity. Here we report the genome sequence of reference strain SC43cl1, a hybrid strain belonging to the TcV discrete typing unit (DTU). The assembled diploid genome was 79 Mbp in size, divided into 1,236 contigs with an average coverage reaching x180. There was extensive synteny of SC43cl1 genome with closely related TcV and TcVI genomes, with limited sequence rearrangements. TcVI genomes included several expansions not present in TcV strains. Comparative analysis of both nuclear and kinetoplast sequences clearly separated TcV from TcVI strains, which strongly supports the current DTU classification.

5.
Vaccine ; 38(29): 4584-4591, 2020 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-32417142

RESUMO

Chagas disease, caused by the protozoan parasite Trypanosoma cruzi is one of the most important neglected parasitic diseases in the Americas. Vaccines represent an attractive complementary strategy for the control of T. cruzi infection and pre-clinical studies in mice demonstrated that trypomastigote surface antigen (TSA-1) and the flagellar calcium-binding (Tc24) parasite antigens are promising candidates for vaccine development. We performed here the first evaluation of the safety and immunogenicity of two recombinant vaccine antigens (named TSA1-C4 and Tc24-C4) in naïve non-human primates. Three rhesus macaques received 3 doses of each recombinant protein, formulated with E6020 (Eisai Co., Ltd.), a novel Toll-like receptor-4 agonist, in a stable emulsion. All parameters from blood chemistry and blood cell counts were stable over the course of the study and unaffected by the vaccine. A specific IgG response against both antigens was detectable after the first vaccine dose, and increased with the second dose. After three vaccine doses, stimulation of PBMCs with a peptide pool derived from TSA1-C4 resulted in the induction of TSA1-C4-specific TNFα-, IL-2- and IFNγ-producing CD4+ in one or two animals while stimulation with a peptide pool derived from Tc24-C4 only activated IFNγ-producing CD4+T cells in one animal. In two animals there was also activation of TSA1-C4-specific IL2-producing CD8+ T cells. This is the first report of the immunogenicity of T. cruzi-derived recombinant antigens formulated as an emulsion with a TLR4 agonist in a non-human primate model. Our results strongly support the need for further evaluation of the preventive efficacy of this type of vaccine in non-human primates and explore the effect of the vaccine in a therapeutic model of naturally-infected Chagasic non-human primates, which would strengthen the rationale for the clinical development as a human vaccine against Chagas disease.

6.
Parasit Vectors ; 12(1): 322, 2019 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-31238941

RESUMO

BACKGROUND: Chagas disease is a zoonotic disease caused by the protozoan parasite Trypanosoma cruzi. The role of dogs as sentinels has been proposed in multiple regions, as they are a domestic reservoir for T. cruzi. Our objective was to determine the prevalence of T. cruzi infection in shelter dogs from southern Louisiana, and assess its magnitude and distribution. RESULTS: A total of 540 dogs were enrolled, from 20 animal shelters, and tested for T. cruzi infection by serological tests (rapid test, ELISA and western blot) and PCR. We documented a high prevalence of T. cruzi infection with at least 6.9% (95% CI: 5.0-9.3%) seropositive and 15.7% (95% CI: 12.9-19.1%) PCR-positive dogs. Serological tests showed limited agreement, and concordance between serology and PCR was higher when considering reactivity to single serological tests. Trypanosoma cruzi infection was distributed evenly among shelters. Infection was significantly correlated with age (R2 = 0.99), indicating an incidence of new cases of 2.27 ± 0.25% per year. CONCLUSION: Trypanosoma cruzi infection is a significant and widespread veterinary problem in shelter dogs in the region, although it is mostly unnoticed by health professionals. This highlights the need for greater awareness of T. cruzi infection among the veterinary community and dog owners.


Assuntos
Doença de Chagas/veterinária , Doenças do Cão/epidemiologia , Cães/parasitologia , Animais , Anticorpos Antiprotozoários/sangue , Doença de Chagas/epidemiologia , Doenças do Cão/parasitologia , Ensaio de Imunoadsorção Enzimática , Feminino , Louisiana/epidemiologia , Masculino , Prevalência , Testes Sorológicos , Trypanosoma cruzi/genética , Trypanosoma cruzi/isolamento & purificação
7.
Mol Brain ; 7: 71, 2014 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-25269856

RESUMO

BACKGROUND: Mutations in the human FOXP2 gene cause speech and language impairments. The FOXP2 protein is a transcription factor that regulates the expression of many downstream genes, which may have important roles in nervous system development and function. An adequate amount of functional FOXP2 protein is thought to be critical for the proper development of the neural circuitry underlying speech and language. However, how FOXP2 gene expression is regulated is not clearly understood. The FOXP2 mRNA has an approximately 4-kb-long 3' untranslated region (3' UTR), twice as long as its protein coding region, indicating that FOXP2 can be regulated by microRNAs (miRNAs). FINDINGS: We identified multiple miRNAs that regulate the expression of the human FOXP2 gene using sequence analysis and in vitro cell systems. Focusing on let-7a, miR-9, and miR-129-5p, three brain-enriched miRNAs, we show that these miRNAs regulate human FOXP2 expression in a dosage-dependent manner and target specific sequences in the FOXP2 3' UTR. We further show that these three miRNAs are expressed in the cerebellum of the human fetal brain, where FOXP2 is known to be expressed. CONCLUSIONS: Our results reveal novel regulatory functions of the human FOXP2 3' UTR sequence and regulatory interactions between multiple miRNAs and the human FOXP2 gene. The expression of let-7a, miR-9, and miR-129-5p in the human fetal cerebellum is consistent with their roles in regulating FOXP2 expression during early cerebellum development. These results suggest that various genetic and environmental factors may contribute to speech and language development and related neural developmental disorders via the miRNA-FOXP2 regulatory network.


Assuntos
Regiões 3' não Traduzidas/genética , Fatores de Transcrição Forkhead/genética , Regulação da Expressão Gênica , MicroRNAs/metabolismo , Sequência de Bases , Sítios de Ligação/genética , Cerebelo/embriologia , Cerebelo/metabolismo , Regulação para Baixo/genética , Feto/metabolismo , Fatores de Transcrição Forkhead/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Humanos , MicroRNAs/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
8.
Neuron ; 73(4): 774-88, 2012 Feb 23.
Artigo em Inglês | MEDLINE | ID: mdl-22365550

RESUMO

EPAC proteins are the guanine nucleotide exchange factors that act as the intracellular receptors for cyclic AMP. Two variants of EPAC genes including EPAC1 and EPAC2 are cloned and are widely expressed throughout the brain. But, their functions in the brain remain unknown. Here, we genetically delete EPAC1 (EPAC1(-/-)), EPAC2 (EPAC2(-/-)), or both EPAC1 and EPAC2 genes (EPAC(-/-)) in the forebrain of mice. We show that EPAC null mutation impairs long-term potentiation (LTP) and that this impairment is paralleled with the severe deficits in spatial learning and social interactions and is mediated in a direct manner by miR-124 transcription and Zif268 translation. Knockdown of miR-124 restores Zif268 and hence reverses all aspects of the EPAC(-/-) phenotypes, whereas expression of miR-124 or knockdown of Zif268 reproduces the effects of EPAC null mutation. Thus, EPAC proteins control miR-124 transcription in the brain for processing spatial learning and social interactions.


Assuntos
Proteína 1 de Resposta de Crescimento Precoce/metabolismo , Fatores de Troca do Nucleotídeo Guanina/deficiência , Relações Interpessoais , Deficiências da Aprendizagem/genética , Deficiências da Aprendizagem/psicologia , MicroRNAs/metabolismo , Análise de Variância , Animais , Biofísica , Imunoprecipitação da Cromatina , Corpo Estriado/metabolismo , Corpo Estriado/patologia , Espinhas Dendríticas/patologia , Espinhas Dendríticas/ultraestrutura , Modelos Animais de Doenças , Proteína 1 de Resposta de Crescimento Precoce/genética , Estimulação Elétrica , Comportamento Exploratório/fisiologia , Perfilação da Expressão Gênica , Hipocampo/patologia , Hipocampo/ultraestrutura , Técnicas In Vitro , Deficiências da Aprendizagem/patologia , Potenciação de Longa Duração/efeitos dos fármacos , Potenciação de Longa Duração/genética , Potenciação de Longa Duração/fisiologia , Masculino , Aprendizagem em Labirinto/fisiologia , Camundongos , Camundongos Transgênicos , MicroRNAs/genética , Microscopia Eletrônica , Neurônios/fisiologia , Análise de Sequência com Séries de Oligonucleotídeos , Técnicas de Patch-Clamp , Biossíntese de Proteínas/genética , Tempo de Reação/genética , Transdução de Sinais/fisiologia , Coloração pela Prata , Transfecção , Proteínas rap1 de Ligação ao GTP/metabolismo
9.
Stem Cells Dev ; 21(3): 411-22, 2012 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-21740234

RESUMO

Chronic intake of nicotine can impair hippocampal plasticity, but the underlying mechanism is poorly understood. Here, we demonstrate that chronic nicotine administration in adult rats inactivates the cyclic AMP-response element binding protein (CREB), a transcription factor that regulates neurogenesis and other plasticity-related processes necessary for learning and memory. Consequently, we showed that impaired CREB signaling is associated with a significant decline in the production of new neurons in the dentate gyrus. Combining retrovirus labeling with gene expression approaches, we found that chronic nicotine administration reduces the number of adult-generated granule neurons by decreasing the survival of newborn cells but not the proliferation of progenitor cells. Additionally, we found that retroviral-mediated expression of a constitutively active CREB in the dentate gyrus rescues survival of newborn cells and reverses the nicotine-induced decline in the number of mature granule neurons. Prolonged nicotine exposure also compromises CREB activation and reduces the viability of progenitor cells in vitro, thereby suggesting that nicotine may exert its adverse effects directly on immature cells in vivo. Taken together, these data demonstrate that inhibition of CREB activation is responsible for the nicotine-induced impairment of hippocampal plasticity.


Assuntos
Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Giro Denteado/citologia , Giro Denteado/efeitos dos fármacos , Nicotina/administração & dosagem , Animais , Bromodesoxiuridina/administração & dosagem , Contagem de Células , Morte Celular , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Meios de Cultura/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Giro Denteado/metabolismo , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/citologia , Neurônios/efeitos dos fármacos , Nicotina/efeitos adversos , Ratos , Retroviridae/genética , Retroviridae/metabolismo , Células-Tronco/citologia , Células-Tronco/efeitos dos fármacos , Células-Tronco/metabolismo , Ativação Transcricional , Transfecção
10.
Cell ; 140(2): 222-34, 2010 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-20141836

RESUMO

N-methyl-D-aspartate (NMDA) receptors constitute a major subtype of glutamate receptors at extrasynaptic sites that link multiple intracellular catabolic processes responsible for irreversible neuronal death. Here, we report that cerebral ischemia recruits death-associated protein kinase 1 (DAPK1) into the NMDA receptor NR2B protein complex in the cortex of adult mice. DAPK1 directly binds with the NMDA receptor NR2B C-terminal tail consisting of amino acid 1292-1304 (NR2B(CT)). A constitutively active DAPK1 phosphorylates NR2B subunit at Ser-1303 and in turn enhances the NR1/NR2B receptor channel conductance. Genetic deletion of DAPK1 or administration of NR2B(CT) that uncouples an activated DAPK1 from an NMDA receptor NR2B subunit in vivo in mice blocks injurious Ca(2+) influx through NMDA receptor channels at extrasynaptic sites and protects neurons against cerebral ischemic insults. Thus, DAPK1 physically and functionally interacts with the NMDA receptor NR2B subunit at extrasynaptic sites and this interaction acts as a central mediator for stroke damage.


Assuntos
Proteínas Reguladoras de Apoptose/metabolismo , Isquemia Encefálica/metabolismo , Proteínas Quinases Dependentes de Cálcio-Calmodulina/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Acidente Vascular Cerebral/metabolismo , Animais , Proteínas Reguladoras de Apoptose/antagonistas & inibidores , Proteínas Reguladoras de Apoptose/genética , Encéfalo/metabolismo , Encéfalo/patologia , Isquemia Encefálica/tratamento farmacológico , Proteínas Quinases Dependentes de Cálcio-Calmodulina/antagonistas & inibidores , Proteínas Quinases Dependentes de Cálcio-Calmodulina/genética , Morte Celular , Proteínas Quinases Associadas com Morte Celular , Camundongos , Neurônios/citologia , Neurônios/metabolismo , Peptídeos/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Acidente Vascular Cerebral/tratamento farmacológico , Acidente Vascular Cerebral/patologia , Produtos do Gene tat do Vírus da Imunodeficiência Humana/genética , Produtos do Gene tat do Vírus da Imunodeficiência Humana/metabolismo
11.
Ageing Res Rev ; 9(1): 20-40, 2010 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19800420

RESUMO

Mitochondrial dysfunction and reactive oxygen species (ROS) production are at the heart of the aging process and are thought to underpin age-related diseases. Mitochondria are not only the primary energy-generating system but also the dominant cellular source of metabolically derived ROS. Recent studies unravel the existence of mechanisms that serve to modulate the balance between energy metabolism and ROS production. Among these is the regulation of proton conductance across the inner mitochondrial membrane that affects the efficiency of respiration and heat production. The field of mitochondrial respiration research has provided important insight into the role of altered energy balance in obesity and diabetes. The notion that respiration and oxidative capacity are mechanistically linked is making significant headway into the field of aging and age-related diseases. Here we review the regulation of cellular energy and ROS balance in biological systems and survey some of the recent relevant studies that suggest that respiratory adaptation and thermodynamics are important in aging and age-related diseases.


Assuntos
Adaptação Fisiológica , Envelhecimento/metabolismo , Regulação da Temperatura Corporal/fisiologia , Mitocôndrias/metabolismo , Doenças Mitocondriais/metabolismo , Animais , Restrição Calórica , Respiração Celular , Diabetes Mellitus Tipo 2/metabolismo , Metabolismo Energético , Humanos , Canais Iônicos/metabolismo , Camundongos , Proteínas Mitocondriais/metabolismo , Doenças Neurodegenerativas/metabolismo , Obesidade/metabolismo , Prótons , Ratos , Espécies Reativas de Oxigênio/metabolismo , Proteína Desacopladora 1
12.
Neuron ; 60(5): 803-17, 2008 Dec 10.
Artigo em Inglês | MEDLINE | ID: mdl-19081376

RESUMO

Aberrant cell-cycle activity and DNA damage are emerging as important pathological components in various neurodegenerative conditions. However, their underlying mechanisms are poorly understood. Here, we show that deregulation of histone deacetylase 1 (HDAC1) activity by p25/Cdk5 induces aberrant cell-cycle activity and double-strand DNA breaks leading to neurotoxicity. In a transgenic model for neurodegeneration, p25/Cdk5 activity elicited cell-cycle activity and double-strand DNA breaks that preceded neuronal death. Inhibition of HDAC1 activity by p25/Cdk5 was identified as an underlying mechanism for these events, and HDAC1 gain of function provided potent protection against DNA damage and neurotoxicity in cultured neurons and an in vivo model for ischemia. Our findings outline a pathological signaling pathway illustrating the importance of maintaining HDAC1 activity in the adult neuron. This pathway constitutes a molecular link between aberrant cell-cycle activity and DNA damage and is a potential target for therapeutics against diseases and conditions involving neuronal death.


Assuntos
Quinase 5 Dependente de Ciclina/fisiologia , Histona Desacetilases/metabolismo , Degeneração Neural/enzimologia , Animais , Animais Recém-Nascidos , Ciclo Celular/fisiologia , Células Cultivadas , Córtex Cerebral/citologia , Imunoprecipitação da Cromatina/métodos , Ensaio Cometa , Condicionamento Psicológico/fisiologia , Quinase 5 Dependente de Ciclina/genética , Quebras de DNA de Cadeia Dupla , Dano ao DNA/genética , Medo/fisiologia , Expressão Gênica/genética , Perfilação da Expressão Gênica/métodos , Proteínas de Fluorescência Verde/biossíntese , Proteínas de Fluorescência Verde/genética , Histona Desacetilase 1 , Humanos , Isquemia/patologia , Antígeno Ki-67/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Degeneração Neural/genética , Tecido Nervoso/metabolismo , Neurônios/fisiologia , Antígeno Nuclear de Célula em Proliferação/metabolismo , Prosencéfalo/metabolismo , Ratos , Transfecção
13.
J Neurochem ; 103(5): 1982-8, 2007 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-17883401

RESUMO

The Kir6.1 channels are a subtype of ATP-sensitive inwardly rectifying potassium (K(ATP)) channels that play an essential role in coupling the cell's metabolic events to electrical activity. In this study, we show that functional Kir6.1 channels are located at excitatory pre-synaptic terminals as a complex with type-1 Sulfonylurea receptors (SUR1) in the hippocampus. The mutant mice with deficiencies in expressing the Kir6.1 or the SUR1 gene are more vulnerable to generation of epileptic form of seizures, compared to wild-type controls. Whole-cell patch clamp recordings demonstrate that genetic deletion of the Kir6.1/SUR1 channels enhances glutamate release at CA3 synapses. Hence, expression of functional Kir6.1/SUR1 channels inhibits seizure responses and possibly acts via limiting excitatory glutamate release.


Assuntos
Regulação da Expressão Gênica/fisiologia , Ácido Glutâmico/metabolismo , Hipocampo/citologia , Canais de Potássio Corretores do Fluxo de Internalização/fisiologia , Convulsões/etiologia , Sinapses/metabolismo , Animais , Comportamento Animal , Agonistas de Aminoácidos Excitatórios/farmacologia , Hipocampo/metabolismo , Humanos , Imunoprecipitação/métodos , Técnicas In Vitro , Canais KATP , Ácido Caínico/farmacologia , Masculino , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/efeitos da radiação , Camundongos , Camundongos Knockout , Técnicas de Patch-Clamp/métodos , Canais de Potássio Corretores do Fluxo de Internalização/deficiência , Ratos , Ratos Sprague-Dawley , Convulsões/genética , Convulsões/fisiopatologia , Sinapses/efeitos dos fármacos
14.
Neuron ; 49(5): 719-33, 2006 Mar 02.
Artigo em Inglês | MEDLINE | ID: mdl-16504947

RESUMO

ADAR2 is a nuclear enzyme essential for GluR2 pre-mRNA editing at Q/R site-607, which gates Ca2+ entry through AMPA receptor channels. Here, we show that forebrain ischemia in adult rats selectively reduces expression of ADAR2 enzyme and, hence, disrupts RNA Q/R site editing of GluR2 subunit in vulnerable neurons. Recovery of GluR2 Q/R site editing by expression of exogenous ADAR2b gene or a constitutively active CREB, VP16-CREB, which induces expression of endogenous ADAR2, protects vulnerable neurons in the rat hippocampus from forebrain ischemic insult. Generation of a stable ADAR2 gene silencing by delivering small interfering RNA (siRNA) inhibits GluR2 Q/R site editing, leading to degeneration of ischemia-insensitive neurons. Direct introduction of the Q/R site edited GluR2 gene, GluR2(R607), rescues ADAR2 degeneration. Thus, ADAR2-dependent GluR2 Q/R site editing determines vulnerability of neurons in the rat hippocampus to forebrain ischemia.


Assuntos
Adenosina Desaminase/metabolismo , Ataque Isquêmico Transitório/patologia , Neurônios/citologia , Prosencéfalo/citologia , Edição de RNA/fisiologia , Receptores de AMPA/metabolismo , Animais , Animais Recém-Nascidos , Northern Blotting/métodos , Western Blotting/métodos , Proteína de Ligação a CREB/metabolismo , Cálcio/metabolismo , Contagem de Células/métodos , Sobrevivência Celular/fisiologia , Células Cultivadas , Modelos Animais de Doenças , Estimulação Elétrica/métodos , Expressão Gênica/efeitos dos fármacos , Expressão Gênica/fisiologia , Proteínas de Fluorescência Verde/metabolismo , Hipocampo/citologia , Imuno-Histoquímica/métodos , Técnicas In Vitro , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Potenciais da Membrana/efeitos da radiação , Neurônios/metabolismo , Fosfopiruvato Hidratase/metabolismo , RNA Mensageiro/biossíntese , RNA Interferente Pequeno/farmacologia , Proteínas de Ligação a RNA , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos
15.
J Neurosci ; 23(3): 826-36, 2003 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-12574411

RESUMO

Long-term depression (LTD) is an activity-dependent weakening of synaptic efficacy at individual inhibitory synapses, a possible cellular model of learning and memory. Here, we show that the induction of LTD of inhibitory transmission recruits activated calcineurin (CaN) to dephosphorylate type-A GABA receptor (GABA(A)Rs) via the direct binding of CaN catalytic domain to the second intracellular domain of the GABA(A)R-gamma(2) subunits. Prevention of the CaN-GABA(A) receptor complex formation by expression of an autoinhibitory domain of CaN in the hippocampus of transgenic mice blocks the induction of LTD. Conversely, genetic expression of the CaN catalytic domain in the hippocampus depresses inhibitory synaptic responses, occluding LTD. Thus, an activity-dependent physical and functional interaction between CaN and GABA(A) receptors is both necessary and sufficient for inducing LTD at CA1 individual inhibitory synapses.


Assuntos
Calcineurina/metabolismo , Hipocampo/metabolismo , Depressão Sináptica de Longo Prazo/fisiologia , Inibição Neural/fisiologia , Receptores de GABA-A/metabolismo , Sinapses/metabolismo , Animais , Estimulação Elétrica , Hipocampo/citologia , Técnicas In Vitro , Substâncias Macromoleculares , Camundongos , Camundongos Mutantes , Camundongos Transgênicos , Técnicas de Patch-Clamp , Fosforilação , Ligação Proteica/fisiologia , Subunidades Proteicas/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo
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