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1.
Vet J ; 198(1): 292-4, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23988333

RESUMO

Recent studies in animal models have focused on the role of cholinergic elements, mainly acetylcholinesterase (AChE) and the 'readthrough' acetylcholinesterase isoform (AChE-R), in seizures. A prospective double-masked study was conducted to assess the activity of AChE and AChE-R in cerebrospinal fluid (CSF) of 26 dogs post-seizure, 28 dogs with intervertebral disc disease (IVDD) and 16 healthy dogs. AChE was also measured in the serum in the post-seizure and IVDD groups. The results showed no significant differences in CSF AChE among the three groups. AChE-R was not detected in any dog and AChE in the serum was similar between groups. This preliminary study provides new information on AChE and AChE-R in the CSF and sera of dogs following naturally-occurring seizures.


Assuntos
Acetilcolinesterase/sangue , Acetilcolinesterase/líquido cefalorraquidiano , Doenças do Cão/enzimologia , Convulsões/veterinária , Animais , Colorimetria/veterinária , Ácido Ditionitrobenzoico/metabolismo , Doenças do Cão/sangue , Doenças do Cão/fisiopatologia , Cães , Método Duplo-Cego , Eletroforese/veterinária , Feminino , Israel , Masculino , Estudos Prospectivos , Convulsões/sangue , Convulsões/enzimologia , Convulsões/fisiopatologia , Estatísticas não Paramétricas
2.
Curr Alzheimer Res ; 10(1): 48-56, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23157337

RESUMO

BACKGROUND: Previous studies have demonstrated alterations in the peripheral cholinergic system in Alzheimer's disease (AD), though results have been inconsistent and not linked to in vivo biomarkers of pathology. We examined the relationship between amyloid-beta (Aß) plaques and plasma cholinesterase activity in a heterogeneous dementia population. METHODS: 29 participants with clinical AD and 35 with non-AD diagnoses underwent positron emission tomography (PET) with the amyloid ligand [11C] PIB and plasma measurements of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) activity. Multi-linear regression was used to evaluate the relationship between AChE or BChE activity and PIB binding (adjusted for age, sex, apolipoprotein E4 and vascular risk), applying voxel-wise and region of interest (ROI) approaches. AChE activity was further adjusted for cholinesterase inhibitor (ChE-I) use. Global amyloid load was measured using a PIB Index, representing mean tracer binding in frontal, parietal, lateral temporal and cingulate cortex. RESULTS: AChE activity was correlated with PIB Index (ß=0.39, p < 0.001) and with regional PIB binding in frontal, temporal, parietal and occipital lobes, precuneus and posterior cingulate on both voxel-wise (p < 0.001 uncorrected) and ROI (ß=0.26-0.41, p < 0.005) analysis. Correlations remained significant after covarying clinical diagnosis (ß=0.42, p=0.001), and among participants naive to ChE-I (ß=0.51, p=0.005). No correlation was found between BChE activity and PIB. Among AD participants, disease severity was not correlated with AChE, BChE or PIB Index. CONCLUSION: AChE activity in plasma is correlated with brain Aß load. Activation of the 'anti-inflammatory cholinergic pathway' may provide the link between Aß plaques and peripheral cholinergic measures.


Assuntos
Acetilcolinesterase/sangue , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/metabolismo , Encéfalo/metabolismo , Idoso , Doença de Alzheimer/diagnóstico por imagem , Compostos de Anilina , Benzotiazóis , Encéfalo/diagnóstico por imagem , Mapeamento Encefálico , Butirilcolinesterase/sangue , Radioisótopos de Carbono , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Tomografia por Emissão de Pósitrons/métodos , Escalas de Graduação Psiquiátrica , Compostos Radiofarmacêuticos , Tiazóis
3.
Proc Natl Acad Sci U S A ; 109(52): E3687-95, 2012 Dec 26.
Artigo em Inglês | MEDLINE | ID: mdl-23236169

RESUMO

Stress induces long-lasting changes in neuronal gene expression and cholinergic neurotransmission, but the underlying mechanism(s) are incompletely understood. Here, we report that chromatin structure and histone modifications are causally involved in this transcriptional memory. Specifically, the AChE gene encoding the acetylcholine-hydrolyzing enzyme acetylcholinesterase is known to undergo long-lasting transcriptional and alternative splicing changes after stress. In mice subjected to stress, we identified two alternative 5' exons that were down-regulated after stress in the hippocampus, accompanied by reduced acetylation and elevated trimethylation of H3K9 at the corresponding promoter. These effects were reversed completely by daily administration of the histone deacetylase (HDAC) inhibitor sodium butyrate for 1 wk after stress. H3K9 hypoacetylation was associated with a selective, sodium butyrate-reversible promoter accumulation of HDAC4. Hippocampal suppression of HDAC4 in vivo completely abolished the long-lasting AChE-related and behavioral stress effects. Our findings demonstrate long-lasting stress-inducible changes in AChE's promoter choices, identify the chromatin changes that support this long-term transcriptional memory, and reveal HDAC4 as a mediator of these effects in the hippocampus.


Assuntos
Acetilcolinesterase/genética , Epigênese Genética , Histona Desacetilases/metabolismo , Estresse Fisiológico/genética , Transcrição Gênica , Acetilcolinesterase/metabolismo , Animais , Cromatina/metabolismo , Metilação de DNA/genética , Epigênese Genética/efeitos dos fármacos , Éxons/genética , Inativação Gênica/efeitos dos fármacos , Hipocampo/citologia , Inibidores de Histona Desacetilases/farmacologia , Masculino , Camundongos , Neurônios/efeitos dos fármacos , Neurônios/enzimologia , Regiões Promotoras Genéticas , Estresse Fisiológico/efeitos dos fármacos , Natação , Transcrição Gênica/efeitos dos fármacos
4.
Front Mol Neurosci ; 5: 66, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22639569

RESUMO

Recent studies show a key role of brain inflammation in epilepsy. However, the mechanisms controlling brain immune response are only partly understood. In the periphery, acetylcholine (ACh) release by the vagus nerve restrains inflammation by inhibiting the activation of leukocytes. Recent reports suggested a similar anti-inflammatory effect for ACh in the brain. Since brain cholinergic dysfunctions are documented in epileptic animals, we explored changes in brain cholinergic gene expression and associated immune response during pilocarpine-induced epileptogenesis. Levels of acetylcholinesterase (AChE) and inflammatory markers were measured using real-time RT-PCR, in-situ hybridization and immunostaining in wild type (WT) and transgenic mice over-expressing the "synaptic" splice variant AChE-S (TgS). One month following pilocarpine, mice were video-monitored for spontaneous seizures. To test directly the effect of ACh on the brain's innate immune response, cytokines expression levels were measured in acute brain slices treated with cholinergic agents. We report a robust up-regulation of AChE as early as 48 h following pilocarpine-induced status epilepticus (SE). AChE was expressed in hippocampal neurons, microglia, and endothelial cells but rarely in astrocytes. TgS mice overexpressing AChE showed constitutive increased microglial activation, elevated levels of pro-inflammatory cytokines 48 h after SE and accelerated epileptogenesis compared to their WT counterparts. Finally we show a direct, muscarine-receptor dependant, nicotine-receptor independent anti-inflammatory effect of ACh in brain slices maintained ex vivo. Our work demonstrates for the first time, that ACh directly suppresses brain innate immune response and that AChE up-regulation after SE is associated with enhanced immune response, facilitating the epileptogenic process. Our results highlight the cholinergic system as a potential new target for the prevention of seizures and epilepsy.

5.
Brain Behav Immun ; 24(5): 792-9, 2010 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-19729061

RESUMO

Oxygen toxicity contributes to the pathogenesis of adverse neurological outcome in survivors of preterm birth in clinical studies. In infant rodent brains, hyperoxia triggers widespread apoptotic neurodegeneration, induces pro-inflammatory cytokines and inhibits growth factor signaling cascades. Since a tissue-protective effect has been observed for recombinant erythropoietin (rEpo), we hypothesized that rEpo would influence hyperoxia-induced oxidative stress in the developing rat brain. The aim of this study was to investigate the level of glutathione (reduced and oxidized), lipid peroxidation and the expression of heme oxygenase-1 (HO-1) and acetylcholinesterase (AChE) after hyperoxia and rEpo treatment. Six-day-old Wistar rats were exposed to 80% oxygen for 2-48 h and received 20,000 IU/kg rEpo intraperitoneally (i.p.). Sex-matched littermates kept under room air and injected with normal saline or rEpo served as controls. Treatment with rEpo significantly reduced hyperoxia-induced upregulation of oxidized glutathione (GSSG) and malondialdehyde, a product of lipid breakdown, whereas reduced glutathione (GSH) was upregulated by rEpo. In parallel, hyperoxia-treated immature rat brains revealed rEpo-suppressible upregulation of synaptic AChE-S as well as of the stress-inducible AChE-R variant, together predicting rEpo-protected cholinergic signaling and restrained inflammatory reactions. Furthermore, treatment with rEpo induced upregulation of HO-1 on mRNA, protein and activity level in the developing rat brain. Our results suggest that rEpo generates its protective effect against oxygen toxicity by a reduction of diverse oxidative stress parameters and by limiting the stressor-inducible changes in both HO-1 and cholinergic functions.


Assuntos
Encéfalo/patologia , Eritropoetina/farmacologia , Hiperóxia/patologia , Estresse Oxidativo/efeitos dos fármacos , Análise de Variância , Animais , Animais Recém-Nascidos , Western Blotting , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Eritropoetina/uso terapêutico , Feminino , Glutationa/metabolismo , Heme Oxigenase-1/metabolismo , Hiperóxia/tratamento farmacológico , Hiperóxia/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Distribuição Aleatória , Ratos , Ratos Wistar , Proteínas Recombinantes , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Tempo
6.
PLoS One ; 4(5): e5618, 2009 May 19.
Artigo em Inglês | MEDLINE | ID: mdl-19479038

RESUMO

BACKGROUND: Mice, whose ribosomal protein S6 cannot be phosphorylated due to replacement of all five phosphorylatable serine residues by alanines (rpS6(P-/-)), are viable and fertile. However, phenotypic characterization of these mice and embryo fibroblasts derived from them, has established the role of these modifications in the regulation of the size of several cell types, as well as pancreatic beta-cell function and glucose homeostasis. A relatively passive behavior of these mice has raised the possibility that they suffer from muscle weakness, which has, indeed, been confirmed by a variety of physical performance tests. METHODOLOGY/PRINCIPAL FINDINGS: A large variety of experimental methodologies, including morphometric measurements of histological preparations, high throughput proteomic analysis, positron emission tomography (PET) and numerous biochemical assays, were used in an attempt to establish the mechanism underlying the relative weakness of rpS6(P-/-) muscles. Collectively, these experiments have demonstrated that the physical inferiority appears to result from two defects: a) a decrease in total muscle mass that reflects impaired growth, rather than aberrant differentiation of myofibers, as well as a diminished abundance of contractile proteins; and b) a reduced content of ATP and phosphocreatine, two readily available energy sources. The abundance of three mitochondrial proteins has been shown to diminish in the knockin mouse. However, the apparent energy deficiency in this genotype does not result from a lower mitochondrial mass or compromised activity of enzymes of the oxidative phosphorylation, nor does it reflect a decline in insulin-dependent glucose uptake, or diminution in storage of glycogen or triacylglycerol (TG) in the muscle. CONCLUSIONS/SIGNIFICANCE: This study establishes rpS6 phosphorylation as a determinant of muscle strength through its role in regulation of myofiber growth and energy content. Interestingly, a similar role has been assigned for ribosomal protein S6 kinase 1, even though it regulates myoblast growth in an rpS6 phosphorylation-independent fashion.


Assuntos
Metabolismo Energético , Debilidade Muscular/metabolismo , Músculo Esquelético/crescimento & desenvolvimento , Músculo Esquelético/metabolismo , Proteína S6 Ribossômica/deficiência , Proteína S6 Ribossômica/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Metabolismo Energético/efeitos dos fármacos , Glucose/metabolismo , Glicogênio/metabolismo , Insulina/farmacologia , Camundongos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Contração Muscular/efeitos dos fármacos , Fibras Musculares Esqueléticas/efeitos dos fármacos , Fibras Musculares Esqueléticas/metabolismo , Músculo Esquelético/efeitos dos fármacos , Tamanho do Órgão/efeitos dos fármacos , Fosforilação Oxidativa/efeitos dos fármacos , Fosfocreatina/metabolismo , Transdução de Sinais/efeitos dos fármacos , Triglicerídeos/metabolismo
7.
Eur J Neurosci ; 27(4): 965-75, 2008 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-18333967

RESUMO

The entorhinal cortex (EC) plays an important role in temporal lobe epilepsy. Under normal conditions, the enriched cholinergic innervation of the EC modulates local synchronized oscillatory activity; however, its role in epilepsy is unknown. Enhanced neuronal activation has been shown to induce transcriptional changes of key cholinergic genes and thus alter cholinergic responses. To examine cholinergic modulations in epileptic tissue we studied molecular and electrophysiological cholinergic responses in the EC of chronically epileptic rats following exposure to pilocarpine or kainic acid. We confirmed that while the total activity of the acetylcholine (ACh)-hydrolysing enzyme, acetylcholinesterase (AChE) was not altered, epileptic rats showed alternative splicing of AChE pre-mRNA transcripts, accompanied by a shift from membrane-bound AChE tetramers to soluble monomers. This was associated with increased sensitivity to ACh application: thus, in control rats, ACh (10-100 microm) induced slow (< 1Hz), periodic events confined to the EC; however, in epileptic rats, ACh evoked seconds-long seizure-like events with initial appearance in the EC, and frequent propagation to neighbouring cortical regions. ACh-induced seizure-like events could be completely blocked by the non-specific muscarinic antagonist, atropine, and were partially blocked by the muscarinic-1 receptor antagonist, pirenzepine; but were not affected by the non-specific nicotinic antagonist, mecamylamine. Epileptic rats presented reduced transcript levels of muscarinic receptors with no evidence of mRNA editing or altered mRNA levels for nicotinic ACh receptors. Our findings suggest that altered cholinergic modulation may initiate seizure events in the epileptic temporal cortex.


Assuntos
Acetilcolina/metabolismo , Acetilcolinesterase/metabolismo , Córtex Entorrinal/fisiopatologia , Epilepsia/fisiopatologia , Convulsões/fisiopatologia , Acetilcolinesterase/química , Acetilcolinesterase/genética , Processamento Alternativo , Animais , Doença Crônica , Convulsivantes/toxicidade , Eletrofisiologia , Epilepsia/induzido quimicamente , Epilepsia/complicações , Expressão Gênica , Ácido Caínico/toxicidade , Microeletrodos , Antagonistas Muscarínicos/farmacologia , Antagonistas Nicotínicos/farmacologia , Técnicas de Cultura de Órgãos , Pilocarpina/toxicidade , RNA Mensageiro/análise , Ratos , Ratos Sprague-Dawley , Receptores Muscarínicos/efeitos dos fármacos , Receptores Muscarínicos/genética , Receptores Muscarínicos/metabolismo , Receptores Nicotínicos/efeitos dos fármacos , Receptores Nicotínicos/genética , Receptores Nicotínicos/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Convulsões/induzido quimicamente , Convulsões/etiologia
8.
Ann N Y Acad Sci ; 1148: 269-81, 2008 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19120119

RESUMO

Mammalian stress responses present a case study for investigating alternative splicing reactions in general, and changes in acetylcholinesterase (AChE) gene expression in particular, under endangered homeostasis. Acetylcholine (ACh) is a major regulator of stress responses, which was recently found to function as an essential route by which neurons can "talk" to immune cells. Therefore, chemical, physical, or psychological insults to the brain might all be traced in peripheral immune cells, which serve as key determinants in the physiological reactions to stress. Stress-induced changes in the alternative splicing patterns of AChE pre-mRNA give this gene and its different protein products diverse stress responsive functions that are associated with both the enzymatic and noncatalytic properties of AChE variants. Transgenic manipulations of AChE gene expression uncovered previously nonperceived aspects of stress responses, including brain-to-blood as well as immune-to-neuronal communication. Herein we discuss the newly gained understanding achieved by using genomic manipulations of AChE gene expression as tools for approaching the alternative splicing features of mammalian stress responses.


Assuntos
Acetilcolinesterase/genética , Acetilcolinesterase/metabolismo , Processamento Alternativo , Encéfalo/fisiologia , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Estresse Psicológico , Acetilcolinesterase/química , Animais , Comportamento Animal/fisiologia , Cognição/fisiologia , Regulação Enzimológica da Expressão Gênica , Homeostase , Humanos , Modelos Moleculares , Conformação Proteica , Isoformas de Proteínas/química
9.
J Mol Neurosci ; 30(1-2): 197-200, 2006.
Artigo em Inglês | MEDLINE | ID: mdl-17192675

RESUMO

Stress insults induce hyperexcitation of cholinergic circuits, both peripherally in the sympathetic pathway (Tracey, 2002) and at the central nervous system (CNS) (Sapolsky, 1996). This reaction can serve to ensure survival but might also entail a risk to the hyperactivated neurons. Consequent changes in the expression of a series of proteins related to acetylcholine (ACh) metabolism might protect the organism from the potentially detrimental effects of this increase in ACh. Of particular interest among these effects is the induction by alternative splicing of the alternative, usually rare, readthrough variant of acetylcholinesterase (AChE), AChE-R. AChE-R is one of the first proteins conveying the signal that the organism has entered a state of alert. Viewing AChER as a stress signal can therefore serve to answer the question "How do we expect a stress signal to operate"? This facilitates the generation of hypotheses regarding the triggering of such signals and the effects it exerts at the molecular, cellular, and physiological levels.


Assuntos
Acetilcolinesterase/genética , Acetilcolinesterase/metabolismo , Transtornos de Estresse Pós-Traumáticos/enzimologia , Estresse Fisiológico/enzimologia , Variação Genética , Humanos , Transdução de Sinais
10.
Cell Tissue Res ; 326(2): 655-69, 2006 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-16802134

RESUMO

Termination of synaptic transmission by neurotransmitter hydrolysis is a substantial characteristic of cholinergic synapses. This unique termination mechanism makes acetylcholinesterase (AChE), the enzyme in charge of executing acetylcholine breakdown, a key component of cholinergic signaling. AChE is now known to exist not as a single entity, but rather as a combinatorial complex of protein products. The diverse AChE molecular forms are generated by a single gene that produces over ten different transcripts by alternative splicing and alternative promoter choices. These transcripts are translated into six different protein subunits. Mature AChE proteins are found as soluble monomers, amphipatic dimers, or tetramers of these subunits and become associated to the cellular membrane by specialized anchoring molecules or members of other heteromeric structural components. A substantial increasing body of research indicates that AChE functions in the central nervous system go far beyond the termination of synaptic transmission. The non-enzymatic neuromodulatory functions of AChE affect neurite outgrowth and synaptogenesis and play a major role in memory formation and stress responses. The structural homology between AChE and cell adhesion proteins, together with the recently discovered protein partners of AChE, predict the future unraveling of the molecular pathways underlying these multileveled functions.


Assuntos
Acetilcolina/metabolismo , Acetilcolinesterase/biossíntese , Neuritos/enzimologia , Neurotransmissores/metabolismo , Sinapses/enzimologia , Transmissão Sináptica/fisiologia , Acetilcolina/genética , Acetilcolinesterase/genética , Animais , Humanos , Memória/fisiologia , Neurotransmissores/genética , Estrutura Quaternária de Proteína/genética , Subunidades Proteicas/biossíntese , Subunidades Proteicas/genética , Homologia Estrutural de Proteína , Sinapses/genética
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