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1.
Nat Neurosci ; 22(7): 1066-1074, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31209380

RESUMO

Cannabis is the most frequently used illicit psychoactive substance worldwide; around one in ten users become dependent. The risk for cannabis use disorder (CUD) has a strong genetic component, with twin heritability estimates ranging from 51 to 70%. Here we performed a genome-wide association study of CUD in 2,387 cases and 48,985 controls, followed by replication in 5,501 cases and 301,041 controls. We report a genome-wide significant risk locus for CUD (P = 9.31 × 10-12) that replicates in an independent population (Preplication = 3.27 × 10-3, Pmeta-analysis = 9.09 × 10-12). The index variant (rs56372821) is a strong expression quantitative trait locus for cholinergic receptor nicotinic α2 subunit (CHRNA2); analyses of the genetically regulated gene expression identified a significant association of CHRNA2 expression with CUD in brain tissue. At the polygenic level, analyses revealed a significant decrease in the risk of CUD with increased load of variants associated with cognitive performance. The results provide biological insights and inform on the genetic architecture of CUD.


Assuntos
Abuso de Maconha/genética , Proteínas do Tecido Nervoso/fisiologia , Receptores Nicotínicos/fisiologia , Idade de Início , Alelos , Transtorno do Deficit de Atenção com Hiperatividade/genética , Encéfalo/metabolismo , Estudos de Casos e Controles , Cromossomos Humanos Par 8/genética , Cognição/fisiologia , Estudos de Coortes , Fatores de Confusão (Epidemiologia) , Dinamarca , Escolaridade , Feminino , Perfilação da Expressão Gênica , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Humanos , Islândia , Masculino , Herança Multifatorial , Proteínas do Tecido Nervoso/biossíntese , Proteínas do Tecido Nervoso/genética , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas , Receptores Nicotínicos/biossíntese , Receptores Nicotínicos/genética , Esquizofrenia/genética , Fumar/genética , Transcriptoma
2.
Nat Rev Dis Primers ; 5(1): 30, 2019 05 02.
Artigo em Inglês | MEDLINE | ID: mdl-31048702

RESUMO

Myasthenia gravis (MG) is an autoimmune disease caused by antibodies against the acetylcholine receptor (AChR), muscle-specific kinase (MuSK) or other AChR-related proteins in the postsynaptic muscle membrane. Localized or general muscle weakness is the predominant symptom and is induced by the antibodies. Patients are grouped according to the presence of antibodies, symptoms, age at onset and thymus pathology. Diagnosis is straightforward in most patients with typical symptoms and a positive antibody test, although a detailed clinical and neurophysiological examination is important in antibody-negative patients. MG therapy should be ambitious and aim for clinical remission or only mild symptoms with near-normal function and quality of life. Treatment should be based on MG subgroup and includes symptomatic treatment using acetylcholinesterase inhibitors, thymectomy and immunotherapy. Intravenous immunoglobulin and plasma exchange are fast-acting treatments used for disease exacerbations, and intensive care is necessary during exacerbations with respiratory failure. Comorbidity is frequent, particularly in elderly patients. Active physical training should be encouraged.


Assuntos
Miastenia Gravis/diagnóstico , Miastenia Gravis/terapia , Acetilcolinesterase/genética , Acetilcolinesterase/fisiologia , Corticosteroides/uso terapêutico , Agrina/genética , Agrina/fisiologia , Anti-Inflamatórios não Esteroides/uso terapêutico , Autoanticorpos/análise , Autoanticorpos/sangue , Biomarcadores/análise , Biomarcadores/sangue , Blefaroptose/etiologia , Colágeno/genética , Colágeno/fisiologia , Cortactina/genética , Cortactina/fisiologia , Eletromiografia/métodos , Humanos , Canal de Potássio Kv1.4/genética , Canal de Potássio Kv1.4/fisiologia , Proteínas Relacionadas a Receptor de LDL/genética , Proteínas Relacionadas a Receptor de LDL/fisiologia , Proteínas Musculares/genética , Proteínas Musculares/fisiologia , Miastenia Gravis/fisiopatologia , Receptores Proteína Tirosina Quinases/genética , Receptores Proteína Tirosina Quinases/fisiologia , Receptores Colinérgicos/genética , Receptores Colinérgicos/fisiologia , Receptores Nicotínicos/genética , Fatores de Risco , Canal de Liberação de Cálcio do Receptor de Rianodina/genética , Canal de Liberação de Cálcio do Receptor de Rianodina/fisiologia
3.
J Biomed Sci ; 26(1): 36, 2019 May 11.
Artigo em Inglês | MEDLINE | ID: mdl-31078140

RESUMO

BACKGROUND: Vascular dementia is the second dementing illness after Alzheimer's disease and caused by reduced blood flow to the brain, and affects cognitive abilities. Our previous study found that auricular electrical stimulation (ES) improved motor and learning impairment, and this phenomenon related with nicotinic acetylcholine receptor (nAChR) expressed cells. However, the underlying mechanism was not clear. In the present study, we investigated the effects of auricular ES on cortical blood flow (CBF) and acetylcholine (ACh) - nAChRs expressed cells. METHODS: Vascular dementia rat animal model was established by permanent occlusions of common carotid arteries with 6-0 nylon suture filament. At 21 day after surgery, motor impairment was confirmed by rotarod test. 15-Hz auricular ES were applied to the ears for 20 min and CBF was recorded at the mean time. The brains were immediately dissected for immunohistochemical stain and western blot analysis. RESULTS: Our results showed that 15-Hz auricular ES rapidly elevated CBF in the middle cerebral artery. The numbers of nAChR α4 immuno-positive cells and western blot levels were significally increased by 15-Hz auricular ES in the hippocampal CA2 output cortex. The numbers of choline acetyltransferase (ChAT) - a key enzyme for biosynthesis of ACh - immuno-positive cells and western blot levels had no significant differences. CONCLUSIONS: The present data suggested that the 15-Hz auricular ES for 20 min rapidly elevated cortical blood flow, promoted the expression of nAChR α4, and would be beneficial for the treatment of Alzheimer type and vascular type dementia.


Assuntos
Córtex Cerebral/irrigação sanguínea , Colina O-Acetiltransferase/genética , Orelha/fisiologia , Habenula/fisiologia , Hipocampo/fisiologia , Receptores Nicotínicos/genética , Animais , Western Blotting , Circulação Cerebrovascular , Colina O-Acetiltransferase/metabolismo , Modelos Animais de Doenças , Estimulação Elétrica , Expressão Gênica , Imuno-Histoquímica , Isquemia/etiologia , Fluxometria por Laser-Doppler , Masculino , Ratos , Ratos Wistar , Receptores Nicotínicos/metabolismo
4.
PLoS Genet ; 15(5): e1008145, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31120900

RESUMO

The interplay of microbiota and the human host is physiologically crucial in health and diseases. The beneficial effects of lactic acid bacteria (LAB), permanently colonizing the human intestine or transiently obtained from food, have been extensively reported. However, the molecular understanding of how LAB modulate human physiology is still limited. G protein-coupled receptors for hydroxycarboxylic acids (HCAR) are regulators of immune functions and energy homeostasis under changing metabolic and dietary conditions. Most mammals have two HCAR (HCA1, HCA2) but humans and other hominids contain a third member (HCA3) in their genomes. A plausible hypothesis why HCA3 function was advantageous in hominid evolution was lacking. Here, we used a combination of evolutionary, analytical and functional methods to unravel the role of HCA3 in vitro and in vivo. The functional studies included different pharmacological assays, analyses of human monocytes and pharmacokinetic measurements in human. We report the discovery of the interaction of D-phenyllactic acid (D-PLA) and the human host through highly potent activation of HCA3. D-PLA is an anti-bacterial metabolite found in high concentrations in LAB-fermented food such as Sauerkraut. We demonstrate that D-PLA from such alimentary sources is well absorbed from the human gut leading to high plasma and urine levels and triggers pertussis toxin-sensitive migration of primary human monocytes in an HCA3-dependent manner. We provide evolutionary, analytical and functional evidence supporting the hypothesis that HCA3 was consolidated in hominids as a new signaling system for LAB-derived metabolites.


Assuntos
Lactobacillales/metabolismo , Receptores Acoplados a Proteínas-G/genética , Receptores Acoplados a Proteínas-G/metabolismo , Receptores Nicotínicos/genética , Receptores Nicotínicos/metabolismo , Dieta , Evolução Molecular , Alimentos Fermentados/microbiologia , Humanos , Lactatos/metabolismo , Filogenia , Receptores Acoplados a Proteínas-G/agonistas , Homologia de Sequência de Aminoácidos , Transdução de Sinais
5.
Medicine (Baltimore) ; 98(20): e15555, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31096457

RESUMO

Acetylcholine receptors (AChRs), including nicotinic acetylcholine receptors (nAChRs) and muscarinic acetylcholine receptors (mAChRs), are highly expressed in bronchial epithelial cells.We used The Cancer Genome Atlas (TCGA) data set to evaluate the expression pattern and prognostic value of the AChR gene family in non-small cell lung cancer (NSCLC). The mined data was validated by quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry (IHC).The survival analysis of TCGA data set showed that only CHRNA7 in the AChR gene family affected prognosis in both lung adenocarcinoma and lung squamous cell carcinoma. Furthermore, qRT-PCR proved that CHRNA7 was significantly upregulated in tumor tissues compared with matched normal tissues at mRNA level (P = .001). The expression level of α7 nAChR (encoded by CHRNA7) in 141 patients was measured by IHC and a high expression of α7 nAChR was associated with unfavorable prognosis (P = .008). Multivariate analysis showed that α7 nAChR was an independent prognostic factor (HR = 2.041; 95% CI 1.188-3.506; P = .007).α7 nAChR was upregulated in NSCLC and was associated with unfavorable prognosis. This gene may be a potential target for lung cancer treatment.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/mortalidade , Neoplasias Pulmonares/mortalidade , Receptores Colinérgicos/genética , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/mortalidade , Idoso , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/mortalidade , Mineração de Dados , Feminino , Humanos , Imuno-Histoquímica , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Prognóstico , RNA Mensageiro/biossíntese , Reação em Cadeia da Polimerase em Tempo Real , Receptores Muscarínicos/genética , Receptores Nicotínicos/genética , Análise de Sobrevida , Regulação para Cima/genética , Receptor Nicotínico de Acetilcolina alfa7
6.
Dev Cell ; 49(1): 100-117.e6, 2019 04 08.
Artigo em Inglês | MEDLINE | ID: mdl-30965033

RESUMO

Mechanisms that control aging are important yet poorly defined. To discover longevity control genes, we performed a forward genetic screen for delayed reproductive aging in C. elegans. Here, we show that am117 is a nonsense mutation in the phm-2 gene, which encodes a protein homologous to human scaffold attachment factor B. phm-2(lf) mutant worms have an abnormal pharynx grinder, which allows live bacteria to accumulate in the intestine. This defect shortens lifespan on highly pathogenic bacteria but extends lifespan and health span on the standard E. coli diet by activating innate immunity pathways that lead to bacterial avoidance behavior and dietary restriction. eat-2(lf) mutants displayed a similar phenotype, indicating accumulation of live bacteria also triggers extended longevity in this mutant. The analysis of phm-2 elucidates connections between pathogen response and aging by defining a mechanism of longevity extension in C. elegans-bacterial colonization, innate immune activation, and bacterial avoidance behavior.


Assuntos
Envelhecimento/genética , Proteínas de Caenorhabditis elegans/genética , Longevidade/genética , Receptores Nicotínicos/genética , Envelhecimento/imunologia , Animais , Aprendizagem da Esquiva/fisiologia , Bactérias/imunologia , Bactérias/patogenicidade , Caenorhabditis elegans/genética , Caenorhabditis elegans/imunologia , Caenorhabditis elegans/microbiologia , Dieta , Escherichia coli/química , Regulação da Expressão Gênica/genética , Interações Hospedeiro-Patógeno/genética , Humanos , Imunidade Inata/genética , Intestinos/microbiologia , Longevidade/imunologia
7.
Nat Ecol Evol ; 3(4): 647-656, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30886368

RESUMO

Imidacloprid, the world's most used insecticide, has caused considerable controversy due to harmful effects on non-pest species and increasing evidence showing that insecticides have become the primary selective force in many insect species. The genetic response to insecticides is heterogeneous across populations and environments, leading to more complex patterns of genetic variation than previously thought. This motivated the investigation of imidacloprid resistance at different temperatures in natural populations of Drosophila melanogaster originating from four climate extremes replicated across two continents. Population and quantitative genomic analysis, supported by functional tests, have revealed a mixed genetic architecture to resistance involving major genes (Paramyosin and Nicotinic-Acetylcholine Receptor Alpha 3) and polygenes with a major trade-off with thermotolerance. Reduced genetic differentiation at resistance-associated loci indicated enhanced gene flow at these loci. Resistance alleles showed stronger evidence of positive selection in temperate populations compared to tropical populations in which chromosomal inversions In(2 L)t, In(3 R)Mo and In(3 R)Payne harbour susceptibility alleles. Polygenic architecture and ecological factors should be considered when developing sustainable management strategies for both pest and beneficial insects.


Assuntos
Drosophila melanogaster/fisiologia , Resistência a Inseticidas/fisiologia , Inseticidas , Neonicotinoides , Nitrocompostos , Termotolerância , Animais , Clima , Feminino , Estudo de Associação Genômica Ampla , Receptores Nicotínicos/genética , Tropomiosina/genética
8.
PLoS Genet ; 15(3): e1007857, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30870432

RESUMO

Schwann cells are integral components of vertebrate neuromuscular synapses; in their absence, pre-synaptic nerve terminals withdraw from post-synaptic muscles, leading to muscle denervation and synapse loss at the developing neuromuscular junction (NMJ). Here, we report a rescue of muscle denervation and neuromuscular synapses loss in type III Neuregulin 1 mutant mice (CRD-Nrg1-/-), which lack Schwann cells. We found that muscle denervation and neuromuscular synapse loss were prevented in CRD-Nrg1-/-mice when presynaptic activity was blocked by ablating a specific gene, such as Snap25 (synaptosomal-associated 25 kDa protein) or Chat (choline acetyltransferase). Further, these effects were mediated by a pathway that requires postsynaptic acetylcholine receptors (AChRs), because ablating Chrna1 (acetylcholine receptor α1 subunit), which encodes muscle-specific AChRs in CRD-Nrg1-/-mice also rescued muscle denervation. Moreover, genetically ablating muscle dihydropyridine receptor (DHPR) ß1 subunit (Cacnb1) or ryanodine receptor 1 (Ryr1) also rescued muscle denervation and neuromuscular synapse loss in CRD-Nrg1-/-mice. Thus, these genetic manipulations follow a pathway-from presynaptic to postsynaptic, and, ultimately to muscle activity mediated by DHPRs and Ryr1. Importantly, electrophysiological analyses reveal robust synaptic activity in the rescued, Schwann-cell deficient NMJs in CRD-Nrg1-/-Cacnb1-/-or CRD-Nrg1-/-Ryr1-/-mutant mice. Thus, a blockade of synaptic activity, although sufficient, is not necessary to preserve NMJs that lack Schwann cells. Instead, a blockade of muscle activity mediated by DHRPs and Ryr1 is both necessary and sufficient for preserving NMJs that lack Schwann cells. These findings suggest that muscle activity mediated by DHPRs/Ryr1 may destabilize developing NMJs and that Schwann cells play crucial roles in counteracting such a destabilizing activity to preserve neuromuscular synapses during development.


Assuntos
Canais de Cálcio Tipo L/genética , Neuregulina-1/genética , Canal de Liberação de Cálcio do Receptor de Rianodina/genética , Sinapses/genética , Animais , Axônios/metabolismo , Eletrofisiologia , Humanos , Camundongos , Neurônios Motores/metabolismo , Denervação Muscular , Músculo Esquelético/metabolismo , Músculo Esquelético/fisiologia , Regeneração Nervosa/genética , Junção Neuromuscular/genética , Terminações Pré-Sinápticas/metabolismo , Receptores Nicotínicos/genética , Células de Schwann/metabolismo , Sinapses/fisiologia , Proteína 25 Associada a Sinaptossoma/genética
9.
BMC Evol Biol ; 19(1): 38, 2019 01 30.
Artigo em Inglês | MEDLINE | ID: mdl-30700248

RESUMO

BACKGROUND: Many physiological processes are influenced by nicotinic acetylcholine receptors (nAChR), ranging from neuromuscular and parasympathetic signaling to modulation of the reward system and long-term memory. Due to the complexity of the nAChR family and variable evolutionary rates among its members, their evolution in vertebrates has been difficult to resolve. In order to understand how and when the nAChR genes arose, we have used a broad approach of analyses combining sequence-based phylogeny, chromosomal synteny and intron positions. RESULTS: Our analyses suggest that there were ten subunit genes present in the vertebrate predecessor. The two basal vertebrate tetraploidizations (1R and 2R) then expanded this set to 19 genes. Three of these have been lost in mammals, resulting in 16 members today. None of the ten ancestral genes have kept all four copies after 2R. Following 2R, two of the ancestral genes became triplicates, five of them became pairs, and three seem to have remained single genes. One triplet consists of CHRNA7, CHRNA8 and the previously undescribed CHRNA11, of which the two latter have been lost in mammals but are still present in lizards and ray-finned fishes. The other triplet consists of CHRNB2, CHRNB4 and CHRNB5, the latter of which has also been lost in mammals. In ray-finned fish the neuromuscular subunit gene CHRNB1 underwent a local gene duplication generating CHRNB1.2. The third tetraploidization in the predecessor of teleosts (3R) expanded the repertoire to a total of 31 genes, of which 27 remain in zebrafish. These evolutionary relationships are supported by the exon-intron organization of the genes. CONCLUSION: The tetraploidizations explain all gene duplication events in vertebrates except two. This indicates that the genome doublings have had a substantial impact on the complexity of this gene family leading to a very large number of members that have existed for hundreds of millions of years.


Assuntos
Evolução Molecular , Receptores Nicotínicos/genética , Vertebrados/genética , Animais , Sequência de Bases , Cromossomos/genética , Éxons/genética , Duplicação Gênica , Humanos , Íntrons/genética , Funções Verossimilhança , Filogenia , Poliploidia , Subunidades Proteicas/genética , Sintenia/genética , Fatores de Tempo
10.
Cell Mol Life Sci ; 76(6): 1151-1167, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30600358

RESUMO

Neuronal nicotinic receptors containing α4 and ß2 subunits assemble in two pentameric stoichiometries, (α4)3(ß2)2 and (α4)2(ß2)3, each with distinct pharmacological signatures; (α4)3(ß2)2 receptors are strongly potentiated by the drug NS9283, whereas (α4)2(ß2)3 receptors are unaffected. Despite this stoichiometry-selective pharmacology, the molecular identity of the target for NS9283 remains elusive. Here, studying (α4)3(ß2)2 receptors, we show that mutations at either the principal face of the ß2 subunit or the complementary face of the α4 subunit prevent NS9283 potentiation of ACh-elicited single-channel currents, suggesting the drug targets the ß2-α4 pseudo-agonist sites, the α4-α4 agonist site, or both sites. To distinguish among these possibilities, we generated concatemeric receptors with mutations at specified subunit interfaces, and monitored the ability of NS9283 to potentiate ACh-elicited single-channel currents. We find that a mutation at the principal face of the ß2 subunit at either ß2-α4 pseudo-agonist site suppresses potentiation, whereas mutation at the complementary face of the α4 subunit at the α4-α4 agonist site allows a significant potentiation. Thus, monitoring potentiation of single concatemeric receptor channels reveals that the ß2-α4 pseudo-agonist sites are required for stoichiometry-selective drug action. Together with the recently determined structure of the (α4)3(ß2)2 receptor, the findings have implications for structure-guided drug design.


Assuntos
Neurônios/fisiologia , Agonistas Nicotínicos/metabolismo , Receptores Nicotínicos/metabolismo , Acetilcolina/metabolismo , Acetilcolina/farmacologia , Potenciais de Ação/efeitos dos fármacos , Sítios de Ligação/genética , Sinergismo Farmacológico , Células HEK293 , Humanos , Modelos Moleculares , Mutação , Neurônios/metabolismo , Agonistas Nicotínicos/farmacologia , Oxidiazóis/metabolismo , Oxidiazóis/farmacologia , Conformação Proteica , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Subunidades Proteicas/química , Subunidades Proteicas/genética , Subunidades Proteicas/metabolismo , Piridinas/metabolismo , Piridinas/farmacologia , Receptores Nicotínicos/química , Receptores Nicotínicos/genética
11.
Artigo em Inglês | MEDLINE | ID: mdl-30682641

RESUMO

The ongoing and widespread emergence of resistance to the existing anti-nematodal pharmacopeia has made it imperative to develop new anthelminthic agents. Historically, plants have been important sources of therapeutic compounds and offer an alternative to synthetic drugs. Monoterpenoids are phytochemicals that have been shown to produce acute toxic effects in insects and nematodes. Previous studies have shown nicotinic acetylcholine receptors (nAChRs) to be possible targets for naturally occurring plant metabolites such as carvacrol and carveol. In this study we examined the effects of monoterpenoid compounds on a levamisole sensitive nAChR from Oesophagostomum dentatum and a nicotine sensitive nAChR from Ascaris suum. We expressed the receptors in Xenopus laevis oocytes and used two-electrode voltage-clamp to characterize the effect of various compounds on these cys-loop receptors. At 100 µM the majority of these compounds acted as antagonists. Interestingly, further experiments revealed that both 0.1 µM and 10 µM menthol potentiated acetylcholine and levamisole responses in the levamisole sensitive receptor but not the nicotine sensitive receptor. We also investigated the effects of 0.1 µM menthol on the contractility of A. suum somatic muscle strips. Menthol produced significant potentiation of peak contractions at each concentration of acetylcholine. The positive allosteric modulatory effects of menthol in both in vivo and in vitro experiments suggests menthol as a promising candidate for combination therapy with cholinergic anthelmintics.


Assuntos
Anti-Helmínticos/farmacologia , Levamisol/farmacologia , Mentol/farmacologia , Receptores Nicotínicos/efeitos dos fármacos , Regulação Alostérica , Animais , Ascaris suum/efeitos dos fármacos , Ascaris suum/genética , Colinérgicos/farmacologia , Feminino , Monoterpenos/farmacologia , Oesophagostomum/efeitos dos fármacos , Oesophagostomum/genética , Receptores Nicotínicos/genética , Xenopus laevis
12.
Behav Brain Res ; 360: 262-269, 2019 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-30552947

RESUMO

The objective of the present study is to investigate the role of α4, α5, α6 or ß2 nAChR subunits in the antidepressant-like effect of bupropion. Adult male mice were treated with subcutaneous acute doses of bupropion (3 and 10 mg/kg) 30 min before the forced swim test (FST) in α4, α5, α6, or ß2 nAChR subunit knockout (KO) and wild-type (WT) mice. In addition, the effects of ß2* antagonist dihydro-ß-erythroidine (DHßE, 3 mg/kg) on antidepressant-like effects of bupropion in C57BL/6 J mice were assessed. Our results showed that baseline immobility and climbing time did not differ between KO and corresponding WT mice except for ß2 KO. Bupropion significantly decreased immobility time and increased climbing time in the α4, α6 and ß2 nAChR KO mice in comparison to WT littermates, indicating that lack of these nAChR subunits enhanced antidepressant effects of bupropion. On the contrary, the α5 nAChR subunit deletion did not alter the FST behavior in the bupropion-treated mice. Not only in the transgenic mice, bupropion also showed antidepressant-like effects in the WT mice. In addition, DHßE pretreatment before bupropion administration resulted in decreased immobility time and increased climbing time. Taken together, the present study provides evidence on the involvement of α4*, α6*, and ß2* (* indicates possible presence of other subunits) nAChRs in the antidepressant-like effects of bupropion in the FST.


Assuntos
Antidepressivos de Segunda Geração/farmacologia , Bupropiona/farmacologia , Receptores Nicotínicos/metabolismo , Natação/psicologia , Análise de Variância , Animais , Di-Hidro-beta-Eritroidina/farmacologia , Relação Dose-Resposta a Droga , Resposta de Imobilidade Tônica/fisiologia , Locomoção/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mutação/genética , Subunidades Proteicas/genética , Subunidades Proteicas/metabolismo , Desempenho Psicomotor/efeitos dos fármacos , Receptores Nicotínicos/genética
13.
Neurol Sci ; 40(3): 503-507, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30542963

RESUMO

INTRODUCTION: The slow-channel congenital myasthenic syndrome (SCCMS) is a postsynaptic form of congenital myasthenic syndromes (CMSs), a clinically heterogeneous group of disorders caused by genetic defects leading to an abnormal signal transmission at the endplate. METHODS: We report clinical and molecular data of a multigenerational family in which the presentation of a progressive proximal-distal weakness with ocular involvement led to a number of different clinical diagnoses. RESULTS: A comprehensive genetic study which included whole-genome linkage analysis and whole-exome sequencing identified a heterozygous missense substitution (c.721C>T, p.L241F) in the ε subunit of the acetylcholine receptor (CHRNE) that was consistent with clinical weakness in all patients. DISCUSSION: SCCMS is characterized by a broad and heterogeneous clinical phenotype in which disease onset, symptoms, severity, and progression can be highly variable even between family members. The identification of a CHRNE mutation allowed to make the definitive diagnosis of CMS in this family and contributed to define the clinical spectrum of this disease.


Assuntos
Saúde da Família , Mutação de Sentido Incorreto/genética , Síndromes Miastênicas Congênitas/genética , Receptores Nicotínicos/genética , Adulto , Idoso , Eletromiografia , Feminino , Ligação Genética , Humanos , Itália , Masculino , Pessoa de Meia-Idade , Músculo Esquelético/fisiopatologia , Síndromes Miastênicas Congênitas/patologia , Síndromes Miastênicas Congênitas/fisiopatologia , Condução Nervosa/genética , Sequenciamento Completo do Exoma
14.
PLoS Negl Trop Dis ; 12(12): e0007021, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30557347

RESUMO

Nematode parasites cause substantial morbidity to billions of people and considerable losses in livestock and food crops. The repertoire of effective anthelmintic compounds for treating these parasitoses is very limited, as drug development has been delayed for decades. Moreover, resistance has become a global concern in livestock parasites and is an emerging issue for human helminthiasis. Therefore, anthelmintics with novel mechanisms of action are urgently needed. Taking advantage of Caenorhabditis elegans as an established model system, we here screened the nematicidal potential of novel imidazolium and imidazole derivatives. One of these derivatives, diisopropylphenyl-imidazole (DII), is lethal to C. elegans at both mature and immature stages. This lethal effect appears to be specific because DII concentrations which prove to be toxic to C. elegans do not induce significant lethality on bacteria, Drosophila melanogaster, and HEK-293 cells. Our analysis of DII action on C. elegans mutant strains determined that, in the adult stage, null mutants of unc-29 are resistant to the drug. Muscle expression of this gene completely restores DII sensitivity. UNC-29 has been largely reported as an essential constituent of the levamisole-sensitive muscle nicotinic receptor (L-AChR). Nevertheless, null mutants in unc-63 and lev-8 (essential and non-essential subunits of L-AChRs, respectively) are as sensitive to DII as the wild-type strain. Therefore, our results suggest that DII effects on adult nematodes rely on a previously unidentified UNC-29-containing muscle AChR, different from the classical L-AChR. Interestingly, DII targets appear to be different between larvae and adults, as unc-29 null mutant larvae are sensitive to the drug. The existence of more than one target could delay resistance development. Its lethality on C. elegans, its harmlessness in non-nematode species and its novel and dual mechanism of action make DII a promising candidate compound for anthelmintic therapy.


Assuntos
Anti-Helmínticos/farmacologia , Caenorhabditis elegans/efeitos dos fármacos , Imidazóis/farmacologia , Animais , Anti-Helmínticos/síntese química , Anti-Helmínticos/química , Caenorhabditis elegans/genética , Caenorhabditis elegans/crescimento & desenvolvimento , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Drosophila melanogaster/efeitos dos fármacos , Drosophila melanogaster/genética , Feminino , Células HEK293 , Humanos , Imidazóis/síntese química , Imidazóis/química , Masculino , Estrutura Molecular , Receptores Nicotínicos/genética , Receptores Nicotínicos/metabolismo
15.
PLoS One ; 13(12): e0208982, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30543688

RESUMO

Cholinergic Receptor Nicotinic Alpha 5 (CHRNA5) is an important susceptibility locus for nicotine addiction and lung cancer. Depletion of CHRNA5 has been associated with reduced cell viability, increased apoptosis and alterations in cellular motility in different cancers yet not in breast cancer. Herein we first showed the expression of CHRNA5 was variable and positively correlated with the fraction of total genomic alterations in breast cancer cell lines and tumors indicating its potential role in DNA damage response (DDR). Next, we demonstrated that silencing of CHRNA5 expression in MCF7 breast cancer cell line by RNAi affected expression of genes involved in cytoskeleton, TP53 signaling, DNA synthesis and repair, cell cycle, and apoptosis. The transcription profile of CHRNA5 depleted MCF7 cells showed a significant positive correlation with that of A549 lung cancer cell line while exhibiting a negative association with the CHRNA5 co-expression profile obtained from Cancer Cell Line Encylopedia (CCLE). Moreover, it exhibited high similarities with published MCF7 expression profiles obtained from exposure to TP53 inducer nutlin-3a and topoisomerase inhibitors. We then demonstrated that CHRNA5 siRNA treatment reduced cell viability and DNA synthesis indicating G1 arrest while it significantly increased apoptotic sub-G1 cell population. Accordingly, we observed lower levels of phosphorylated RB (Ser807/811) and an increased BAX/BCL2 ratio in RNAi treated MCF7 cells. We also showed that CHRNA5 RNAi transcriptome correlated negatively with DDR relevant gene expression profile in breast cancer gene expression datasets while the coexposure to topoisomerase inhibitors in the presence of CHRNA5 RNAi enhanced chemosensitivity potentially due to reduced DDR. CHRNA5 RNAi consistently lowered total CHEK1 mRNA and protein levels as well as phosphorylated CHEK1 (Ser345) in MCF7 cells. We also detected a significant positive correlation between the expression levels of CHRNA5 and CHEK1 in CCLE, TCGA and METABRIC breast cancer datasets. Our study suggests CHRNA5 RNAi is associated with cell cycle inhibition, apoptosis as well as reduced DDR and increased drug sensitivity in breast cancer yet future studies are warranted since dose- and cell line-specific differences exist in response to CHRNA5 depletion. Gene expression microarray data can be accessed from GEO database under the accession number GSE89333.


Assuntos
Apoptose/genética , Neoplasias da Mama/patologia , Ciclo Celular/genética , Dano ao DNA/genética , Proteínas do Tecido Nervoso/deficiência , Proteínas do Tecido Nervoso/genética , Interferência de RNA , Receptores Nicotínicos/deficiência , Receptores Nicotínicos/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Sobrevivência Celular/genética , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Transdução de Sinais/genética
16.
Zool Res ; 39(6): 431-436, 2018 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-30084433

RESUMO

Three-finger toxins (TFTs) are well-recognized non-enzymatic venom proteins found in snakes. However, although TFTs exhibit accelerated evolution, the drivers of this evolution remain poorly understood. The structural complexes between long-chain α-neurotoxins, a subfamily of TFTs, and their nicotinic acetylcholine receptor targets have been determined in previous research, providing an opportunity to address such questions. In the current study, we observed several previously identified positively selected sites (PSSs) and the highly variable C-terminal loop of these toxins at the toxin/receptor interface. Of interest, analysis of the molecular adaptation of the toxin-recognition regions in the corresponding receptors provided no statistical evidence for positive selection. However, these regions accumulated abundant amino acid variations in the receptors from the prey of snakes, suggesting that accelerated substitution of TFTs could be a consequence of adaptation to these variations. To the best of our knowledge, this atypical evolution, initially discovered in scorpions, is reported in snake toxins for the first time and may be applicable for the evolution of toxins from other venomous animals.


Assuntos
Venenos de Serpentes/genética , Animais , Evolução Molecular , Variação Genética/genética , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Receptores Nicotínicos/genética , Receptores Nicotínicos/metabolismo , Venenos de Serpentes/metabolismo , Serpentes/genética
17.
J Clin Neuromuscul Dis ; 20(1): 14-27, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30124556

RESUMO

OBJECTIVES: To investigate the mutational spectrum and genotype-phenotype correlation in Indian patients with congenital myasthenic syndrome (CMS), using next-generation sequencing of 5 genes. METHODS: CHRNE, COLQ, DOK7, RAPSN, and GFPT1 were sequenced in 25 affected patients. RESULTS: We found clinically significant variants in 18 patients, of which variants in CHRNE were the most common, and 9 were novel. A common pathogenic COLQ variant was also detected in 4 patients with isolated limb-girdle congenital myasthenia. CONCLUSIONS: Targeted screening of 5 genes is an effective alternate test for CMS, and an affordable one even in a developing country such as India. In addition, we recommend that patients with isolated limb-girdle congenital myasthenia be screened initially for the common COLQ pathogenic variant. This study throws the first light on the genetic landscape of CMSs in India.


Assuntos
Proteínas Musculares/genética , Mutação/genética , Síndromes Miastênicas Congênitas/epidemiologia , Síndromes Miastênicas Congênitas/genética , Acetilcolinesterase/genética , Adolescente , Adulto , Criança , Pré-Escolar , Estudos de Coortes , Colágeno/genética , Feminino , Estudos de Associação Genética , Glutamina-Frutose-6-Fosfato Transaminase (Isomerizante)/metabolismo , Humanos , Índia/epidemiologia , Masculino , Pessoa de Meia-Idade , Receptores Nicotínicos/genética , Índice de Gravidade de Doença , Adulto Jovem
18.
Int J Environ Health Res ; 28(6): 683-696, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30103633

RESUMO

Neonicotinoids, such as acetamiprid (ACE), a pesticide used worldwide, are believed to be safe for human use. These molecules are structurally similar to nicotine, act as nicotinic acetylcholine receptor (nAChR) agonists, and were shown to be associated with neuromuscular and reproductive disorders, but these experiments were primarily performed in mature animals. In this study, the effects of ACE on the testes of immature mice were examined. The exposure of 3-week-old mice to ACE-containing water for 180 days led to a decrease in body weight and mildly affected spermatogenesis. Additionally, the expression of testosterone-metabolism genes, nAChR subunit genes, and proliferation-associated genes decreased in the testes of ACE-treated mice. Our results show that immature rodents may be less sensitive to ACE than mature ones, that mice may be more likely to accumulate ACE than rats, and that the development of disorders may be affected by the accumulation of ACE in the testes.


Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Inseticidas/toxicidade , Neonicotinoides/toxicidade , Espermatogênese/efeitos dos fármacos , Testículo/efeitos dos fármacos , Animais , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Inseticidas/metabolismo , Masculino , Camundongos , Neonicotinoides/metabolismo , Receptores Nicotínicos/genética , Testosterona/metabolismo , Perda de Peso/efeitos dos fármacos
19.
Int J Mol Sci ; 19(8)2018 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-30115897

RESUMO

To further investigate the importance of Schistosoma japonicum acetylcholinesterase (SjAChE) in cholinergic signaling for parasite growth and development, we used RNA interference (RNAi) to knock-down its expression in adults and eggs in vitro. This resulted in its reduced transcription but also expression of other important genes involved both in cholinergic signaling and glucose uptake were impacted substantially. Significant decreases in AChE protein expression, AChE enzymatic activity, and glucose uptake were observed in the SjAChE-knockdown parasites compared with luciferase controls. In vaccine/challenge experiments, we found that immunization of mice with recombinant SjAChE (rSjAChE) expressed in Escherichia coli elicited reductions in male worm numbers (33%), liver granuloma density (41%), and reduced numbers of mature intestinal eggs (73%) in the vaccinated group compared with the control group. These results indicate AChE plays an important role in the metabolism of male worms, and impacts indirectly on female fecundity leading to increased numbers of immature eggs being released and reduced sizes of liver granulomas. Furthermore, cytokine analysis showed that immunization of mice with rSjAChE elicited a predominantly Th1-type immune response characterized by increased production of IFNγ in splenic CD4⁺ T cells of vaccinated mice. The study confirms the potential of SjAChE as a vaccine/drug candidate against zoonotic schistosomiasis japonica.


Assuntos
Acetilcolinesterase/metabolismo , Parasitos/enzimologia , Parasitos/crescimento & desenvolvimento , Schistosoma japonicum/enzimologia , Schistosoma japonicum/crescimento & desenvolvimento , Animais , Formação de Anticorpos , Citocinas/metabolismo , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Glucose/metabolismo , Fígado/parasitologia , Fígado/patologia , Camundongos Endogâmicos CBA , Óvulo/metabolismo , Parasitos/genética , Interferência de RNA , RNA de Cadeia Dupla/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Nicotínicos/genética , Receptores Nicotínicos/metabolismo , Schistosoma japonicum/genética , Esquistossomose Japônica/imunologia , Esquistossomose Japônica/parasitologia , Baço/metabolismo , Transcrição Genética , Resultado do Tratamento , Vacinação , Vacinas/imunologia
20.
Cell Mol Life Sci ; 75(23): 4465-4478, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30069700

RESUMO

Snake venom α-neurotoxins potently inhibit rodent nicotinic acetylcholine receptors (nAChRs), but their activity on human receptors and their role in human paralysis from snakebite remain unclear. We demonstrate that two short-chain α-neurotoxins (SαNTx) functionally inhibit human muscle-type nAChR, but are markedly more reversible than against rat receptors. In contrast, two long-chain α-neurotoxins (LαNTx) show no species differences in potency or reversibility. Mutant studies identified two key residues accounting for this. Proteomic and clinical data suggest that paralysis in human snakebites is not associated with SαNTx, but with LαNTx, such as in cobras. Neuromuscular blockade produced by both subclasses of α-neurotoxins was reversed by antivenom in rat nerve-muscle preparations, supporting its effectiveness in human post-synaptic paralysis.


Assuntos
Neurotoxinas/envenenamento , Paralisia/fisiopatologia , Mordeduras de Serpentes/fisiopatologia , Venenos de Serpentes/envenenamento , Transmissão Sináptica/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Antivenenos/farmacologia , Humanos , Junção Neuromuscular/efeitos dos fármacos , Junção Neuromuscular/metabolismo , Neurotoxinas/genética , Paralisia/induzido quimicamente , Proteômica/métodos , Ratos , Receptores Nicotínicos/genética , Receptores Nicotínicos/metabolismo , Homologia de Sequência de Aminoácidos , Venenos de Serpentes/genética , Especificidade da Espécie
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