RESUMO
Lung inflammation is modulated by cholinergic signaling and exercise training protects mice against pulmonary emphysema development; however, whether exercise training engages cholinergic signaling is unknown. AIMS: As cholinergic signaling is directly linked to the vesicular acetylcholine transporter (VAChT) levels, we evaluated whether the effects of aerobic exercise training depend on the VAChT levels in mice with pulmonary emphysema. MAIN METHODS: Wild-type (WT) and mutant (KDHOM) mice (65-70% of reduction in VAChT levels) were exposed to cigarette smoke (30 min, 2×/day, 5×/week, 12 weeks) and submitted or not to aerobic exercise training on a treadmill (60 min/day, 5×/week, 12 weeks). Lung function and inflammation were evaluated. KEY FINDINGS: Cigarette smoke reduced body mass in mice (p < 0.001) and increased alveolar diameter (p < 0.001), inflammation (p < 0.001) and collagen deposition (p < 0.01) in lung tissue. Both trained groups improved their performance in the final physical test compared to the initial test (p < 0.001). In WT mice, exercise training protected against emphysema development (p < 0.05), reduced mononuclear cells infiltrate (p < 0.001) and increased MAC-2 positive cells in lung parenchyma (p < 0.05); however, these effects were not observed in KDHOM mice. The exercise training reduced iNOS-positive cells (p < 0.001) and collagen fibers deposition (p < 0.05) in lung parenchyma of WT and KDHOM mice, although KDHOM mice showed higher levels of iNOS-positive cells. SIGNIFICANCE: Our data suggest that the protective effects of aerobic exercise training on pulmonary emphysema are, at least in part, dependent on the integrity of the lung cholinergic signaling.
Assuntos
Fumar Cigarros , Enfisema , Enfisema Pulmonar , Animais , Colinérgicos , Inflamação , Pulmão , Camundongos , Camundongos Endogâmicos C57BL , Enfisema Pulmonar/etiologia , Enfisema Pulmonar/prevenção & controle , Proteínas Vesiculares de Transporte de AcetilcolinaRESUMO
INTRODUCTION: Reduced expression of the vesicular acetylcholine transporter (VAChT) leads to changes in the distribution and shape of synaptic vesicles (SVs) at neuromuscular junctions (NMJs), suggesting vesicular acetylcholine (ACh) as a key component of synaptic structure and function. It is poorly understood how long-term changes in cholinergic transmission contribute to age- and disease-related degeneration in the motor system. METHODS: In this study we performed confocal imaging, electrophysiology, electron microscopy, and analyses of respiratory mechanics of the diaphragm NMJ components in 12-month-old wild-type (WT) and VAChTKDHOM mice. RESULTS: Diaphragms of NMJs of the VAChTKDHOM mice were similar to those in WT mice in number, colocalization, and fragmentation of pre-/postsynaptic components. However, they had increased spontaneous SV exocytosis, miniature endplate potential frequency, and diminished MEPP amplitude. No impairment in respiratory mechanics at rest was observed, probably due to the large neurotransmission safety factor of the diaphragm. DISCUSSION: The present findings help us to understand the consequences of reduced ACh release at the NMJs during aging.
Assuntos
Envelhecimento/patologia , Diafragma/ultraestrutura , Síndromes Miastênicas Congênitas/patologia , Junção Neuromuscular/ultraestrutura , Vesículas Sinápticas/ultraestrutura , Acetilcolina/metabolismo , Envelhecimento/metabolismo , Animais , Diafragma/metabolismo , Diafragma/fisiopatologia , Modelos Animais de Doenças , Endocitose , Potenciais Pós-Sinápticos Excitadores/fisiologia , Exocitose , Técnicas de Silenciamento de Genes , Camundongos , Microscopia Confocal , Microscopia Eletrônica de Transmissão , Placa Motora , Síndromes Miastênicas Congênitas/genética , Síndromes Miastênicas Congênitas/metabolismo , Síndromes Miastênicas Congênitas/fisiopatologia , Junção Neuromuscular/metabolismo , Junção Neuromuscular/fisiopatologia , Mecânica Respiratória/fisiologia , Transmissão Sináptica , Vesículas Sinápticas/metabolismo , Proteínas Vesiculares de Transporte de Acetilcolina/genéticaRESUMO
Chronic kidney disease (CKD) is associated with several other long-lasting conditions such as diabetes and cardiovascular diseases and it is a significant contributor to mortality worldwide. Obstructive kidney disease is one of the leading causes of CKD in children and may result from a wide variety of pathologic processes. Recent studies have shown that α7 nicotinic acetylcholine receptor (α7 nAChR) activation in the cholinergic anti-inflammatory pathway reduces production of inflammatory mediators and consequently prevents tissue injury and death. Here, we examined the role of endogenous release of acetylcholine on the development of fibrosis in renal tissue using a model of unilateral ureter obstruction (UUO)-induced CKD, in which obstruction promotes inflammation-mediated kidney damages. To interfere with acetylcholine secretion, we used mice in which the vesicular acetylcholine transporter is genetically reduced (VAChT KD(hom) mice). We observed a higher renal damage in VAChT mutant mice when compared to wild type controls, exemplified by higher proteinuria and increased amount of type 1 collagen in the kidney tissue, indicating accentuated fibrogenesis. These results were accompanied by enhanced localized kidney inflammation, with increased TH1/TH17 profile response. Administration of PNU-282987, a selective agonist of α7 nAChR, significantly attenuated kidney injury after UUO in VAChT KD(hom) mice, indicating that the lack of acetylcholine release decrease the action of the cholinergic anti-inflammatory pathway, promoting an up-regulation of pro-inflammatory and pro-fibrotic pathways. These results suggest that physiological activation of the cholinergic anti-inflammatory pathway regulates inflammatory responses in the kidney suggesting a new therapeutic approach for kidney disease.
RESUMO
The aim of the present study was to evaluate the effects of changes to the autonomic nervous system in mice during the acute phase of Chagas disease, which is an infection caused by the parasite Trypanosoma cruzi. The following types of mice were inoculated with T. cruzi (CHG): wild-type (WT) and vesicular acetylcholine transporter knockdown (KDVAChT) C57BL/6j mice; wild-type non-treated (NT) FVB mice; FVB mice treated with pyridostigmine bromide (PYR) or salbutamol (SALB); and ß(2)-adrenergic receptor knockout (KOß2) FVB mice. During infection and at 18-21 days after infection (acute phase), the survival curves, parasitaemia, electrocardiograms, heart rate variability, autonomic tonus and histopathology of the animals were evaluated. Negative control groups were matched for age, genetic background and treatment. The KDVAChT-CHG mice exhibited a significant shift in the electrocardiographic, autonomic and histopathological profiles towards a greater inflammatory immune response that was associated with a reduction in blood and tissue parasitism. In contrast, the CHG-PYR mice manifested reduced myocardial inflammation and lower blood and tissue parasitism. Similar results were observed in CHG-SALB animals. Unexpectedly, the KOß2-CHG mice exhibited less myocardial inflammation and higher blood and tissue parasitism, which were associated with reduced mortality. These findings could have been due to the increase in vagal tone observed in the KOß2 mice, which rendered them more similar to the CHG-PYR animals. In conclusion, our results indicate a marked immunomodulatory role for the parasympathetic and sympathetic autonomic nervous systems, which inhibit both the inflammatory immune response and parasite clearance during the acute phase of experimental Chagas heart disease in mice.
Assuntos
Doença de Chagas/imunologia , Doença de Chagas/fisiopatologia , Inflamação/imunologia , Inflamação/fisiopatologia , Sistema Nervoso Parassimpático/fisiopatologia , Sistema Nervoso Simpático/fisiopatologia , Doença Aguda , Animais , Atenolol/farmacologia , Doença de Chagas/metabolismo , Doença de Chagas/parasitologia , Eletrocardiografia/métodos , Frequência Cardíaca/efeitos dos fármacos , Frequência Cardíaca/fisiologia , Inflamação/metabolismo , Inflamação/parasitologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Miocárdio/metabolismo , Miocárdio/patologia , Parasitemia/imunologia , Parasitemia/metabolismo , Parasitemia/parasitologia , Parasitemia/fisiopatologia , Sistema Nervoso Parassimpático/efeitos dos fármacos , Sistema Nervoso Parassimpático/imunologia , Sistema Nervoso Parassimpático/metabolismo , Propranolol/farmacologia , Brometo de Piridostigmina/farmacologia , Receptores Adrenérgicos beta 2/metabolismo , Sistema Nervoso Simpático/efeitos dos fármacos , Sistema Nervoso Simpático/imunologia , Sistema Nervoso Simpático/metabolismoRESUMO
We isolated from a brain library a cDNA encoding an isoform of rat CED-6 that has not been previously described. This transcript results from alternative splicing of the ced-6 gene present on chromosome 9. We expressed this isoform as his-tagged protein in E. coli and used the purified protein to raise antibodies to investigate the expression of CED-6 in rat brain. Immunoblot analysis showed the presence of CED-6 as a doublet of approximately 34 and 33 kDa in cortex, hippocampus and cerebellum, indicating that the protein was present in different regions of the brain. Subcellular fractionation experiments showed that CED-6 immunoreactivity did not concentrate in GFAP-containing glial vesicles, whereas it showed a distribution similar to the synaptotagmin in synaptosomes-enriched fractions, suggesting that CED-6 is present in neurons. CED-6 immunoreactivity was also investigated using immunohistochemistry analysis and it was found in several brain regions, being particularly strong in the cell body of some groups of neurons such as Purkinje cell layer of cerebellum, and pyramidal cells of the hippocampal formation and also in epithelial cells from the choroid plexus. Importantly, CED-6 immunoreactivity colocalized with a neuronal marker but not with a glial marker. Considering that several PTB-containing proteins bind clathrin, we investigated whether rat CED-6 would also have this property. Yeast two-hybrid and GST pull-down analysis indicated that ratCED-6 interacts with clathrin and in cultured cells we detected colocalization between CED-6 and clathrin-coated vesicles. The present findings suggest that CED-6 may have a role in endocytic trafficking or signaling in neurons.