RESUMO
Dysferlin protein plays a key role in the multimolecular complex responsible for the maintenance of sarcolemma integrity and skeletal muscle cell functioning. We studied the membrane distribution of nicotinic acetylcholine receptors and α2 isoform of Na,K-ATPase in motor endplates of m. soleus in dysferlin-deficient Bla/J mice (a dysferlinopathy model). Endplates of Bla/J mice were characterized by increased area (without changes in fragmentation degree) and reduced density of the membrane distribution of nicotinic acetylcholine receptors in comparison with the corresponding parameters in control С57Bl/6 mice. The density of the membrane distribution of α2 isoform of Na,K-ATPase was also reduced, but the level of the corresponding mRNA remained unchanged. It can be hypothesized that abnormal membrane localization of α2 isoform of Na,K-ATPase results from adaptive skeletal muscle remodeling under conditions of chronic motor dysfunction.
Assuntos
Membrana Celular/metabolismo , Músculo Esquelético/metabolismo , Isoformas de Proteínas/metabolismo , ATPase Trocadora de Sódio-Potássio/metabolismo , Animais , Disferlina/genética , Disferlina/metabolismo , Masculino , Camundongos , Isoformas de Proteínas/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , ATPase Trocadora de Sódio-Potássio/genéticaRESUMO
The third author's name should read: V. V. Matchkov.
RESUMO
BACKGROUND: Abnormalities in circadian rhythms may be causal factors in development of major depressive disorder. The biology underlying a causal relationship between circadian rhythm disturbances and depression is slowly being unraveled. Although there is no direct evidence of dysregulation of clock gene expression in depressive patients, many studies have reported single-nucleotide polymorphisms in clock genes in these patients. METHODS: In the present study we investigated whether a depression-like state in rats is associated with alternations of the diurnal expression of clock genes. The validated chronic mild stress (CMS) animal model of depression was used to investigate rhythmic expression of three clock genes: period genes 1 and 2 (Per1 and Per2) and Bmal1. Brain and liver tissue was collected from 96 animals after 3.5 weeks of CMS (48 control and 48 depression-like rats) at a 4h sampling interval within 24h. We quantified expression of clock genes on brain sections in the prefrontal cortex, nucleus accumbens, pineal gland, suprachiasmatic nucleus, substantia nigra, amygdala, ventral tegmental area, subfields of the hippocampus, and the lateral habenula using in situ hybridization histochemistry. Expression of clock genes in the liver was monitored by real-time quantitative polymerase chain reaction (PCR). RESULTS: We found that the effect of CMS on clock gene expression was selective and region specific. Per1 exhibits a robust diurnal rhythm in most regions of interest, whereas Bmal1 and in particular Per2 were susceptible to CMS. CONCLUSION: The present results suggest that altered expression of investigated clock genes is likely associated with the induction of a depression-like state in the CMS model.
Assuntos
Fatores de Transcrição ARNTL/metabolismo , Encéfalo/metabolismo , Depressão/metabolismo , Fígado/metabolismo , Proteínas Circadianas Period/metabolismo , Fatores de Transcrição ARNTL/genética , Animais , Comportamento Animal , Encéfalo/fisiopatologia , Ritmo Circadiano , Depressão/genética , Depressão/fisiopatologia , Depressão/psicologia , Sacarose Alimentar/administração & dosagem , Modelos Animais de Doenças , Comportamento Alimentar , Regulação da Expressão Gênica , Fígado/fisiopatologia , Masculino , Proteínas Circadianas Period/genética , Ratos Wistar , Fatores de TempoRESUMO
The altered activity of the hypothalamic-pituitary-adrenal (HPA) axis is often observed in stress-related disorders. According to the literature, about 60% of patients with major depressive disorder elicit high levels of cortisol. It is still unclear why high cortisol levels are not observed in all patients. In this study, we used the chronic mild stress (CMS) rat model of depression, which is based on continuous exposure to unpredictable stressors, to track longitudinal changes in HPA function using fecal corticosterone metabolites (FCM) as a read out. The dexamethasone suppression test was used to assess negative feedback inhibition of the HPA axis. Our results show (1) a disturbance in diurnal corticosterone rhythm measured as fluctuations of the diurnal FCM peak, (2) differences in corticosterone levels between stress-susceptible and stress-resilient animals, (3) recovery of diurnal corticosterone rhythm after 8 weeks of CMS, and (4) alterations in sensitivity to dexamethasone in negative feedback regulation of corticosterone secretion during the time course of CMS. Thus, a disruption of HPA axis circadian rhythmicity coincides with the initial state in the development of depression-like behavior. This chronobiological abnormality, as well as the hypersecretion of corticosterone, is state, rather than trait, dependent.
Assuntos
Ritmo Circadiano/fisiologia , Corticosterona/metabolismo , Depressão/fisiopatologia , Sistema Hipotálamo-Hipofisário/fisiologia , Sistema Hipófise-Suprarrenal/fisiologia , Animais , Dexametasona , Fezes/química , Masculino , RatosRESUMO
Major depressive disorder (MDD) is a growing problem worldwide. Though, the etiology remains unresolved, circadian rhythm disturbances are frequently observed in MDD and thus is speculated to play a key role herein. The present study focuses on circadian rhythm disturbances in the chronic mild stress (CMS) animal model of depression and examined whether the atypical antidepressant, agomelatine, which is mediating its action via melatonergic and serotonergic receptors, is capable of resynchronizing the perturbed rhythm. Melatonin is often used as a marker of the circadian phase, but the functional and behavioral output is dictated on a cellular level by the molecular clock, driven by the clock genes. We applied in situ hybridization histochemistry to measure the expression levels of the core clock genes, period (Per) 1 and 2 and bone and muscle ARNT-like protein 1 (Bmal1), in multiple brain regions believed to be implicated in depression. Agomelatine showed an antidepressant-like effect in the sucrose consumption test and an anxiolytic-like profile in the elevated zero maze. We found that CMS increased nighttime melatonin release in rats and that agomelatine attenuated this effect. Stress was shown to have a time and region-specific effect on clock gene expression in the brain. Treatment with agomelatine failed to normalize clock gene expression, and the observed modifying effect on gene expression did not associate with the antidepressant-like effect. This suggests that the antidepressant actions of agomelatine are mainly independent of circadian rhythm synchronization and, in this regard, not superior to traditional antidepressants tested in our model.
Assuntos
Acetamidas/uso terapêutico , Antidepressivos/uso terapêutico , Proteínas CLOCK/biossíntese , Ritmo Circadiano/efeitos dos fármacos , Depressão/tratamento farmacológico , Modelos Animais de Doenças , Acetamidas/farmacologia , Animais , Antidepressivos/farmacologia , Proteínas CLOCK/genética , Ritmo Circadiano/fisiologia , Depressão/genética , Depressão/metabolismo , Masculino , Melatonina/farmacologia , Proteínas Circadianas Period/biossíntese , Ratos , Ratos Wistar , Resultado do TratamentoRESUMO
AIMS: The Na,K-ATPase is involved in a large number of regulatory activities including cSrc-dependent signalling. Upon inhibition of the Na,K-ATPase with ouabain, cSrc activation is shown to occur in many cell types. This study tests the hypothesis that acute potentiation of agonist-induced contraction by ouabain is mediated through Na,K-ATPase-cSrc signalling-dependent sensitization of vascular smooth muscle cells to Ca2+ . METHODS: Agonist-induced rat mesenteric small artery contraction was examined in vitro under isometric conditions and in vivo in anaesthetized rats. Arterial wall tension and [Ca2+ ]i in vascular smooth muscle cells were measured simultaneously. Changes in cSrc and myosin phosphatase targeting protein 1 (MYPT1) phosphorylation were analysed by Western blot. Protein expression was examined with immunohistochemistry. The α1 and α2 isoforms of the Na,K-ATPase were transiently downregulated by siRNA transfection in vivo. RESULTS: Ten micromolar ouabain, but not digoxin, potentiated contraction to noradrenaline. This effect was not endothelium-dependent. Ouabain sensitized smooth muscle cells to Ca2+ , and this was associated with increased phosphorylation of cSrc and MYPT1. Inhibition of tyrosine kinase by genistein, PP2 or pNaKtide abolished the potentiating effect of ouabain on arterial contraction and Ca2+ sensitization. Downregulation of the Na,K-ATPase α2 isoform made arterial contraction insensitive to ouabain and tyrosine kinase inhibition. CONCLUSION: Data suggest that micromolar ouabain potentiates agonist-induced contraction of rat mesenteric small artery via Na,K-ATPase-dependent cSrc activation, which increases Ca2+ sensitization of vascular smooth muscle cells by MYPT1 phosphorylation. This mechanism may be critical for acute control of vascular tone.
Assuntos
Sinalização do Cálcio , Artérias Mesentéricas/enzimologia , ATPase Trocadora de Sódio-Potássio/metabolismo , Vasoconstrição , Quinases da Família src/metabolismo , Animais , Sinalização do Cálcio/efeitos dos fármacos , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/enzimologia , Inibidores Enzimáticos/farmacologia , Masculino , Artérias Mesentéricas/efeitos dos fármacos , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/enzimologia , Fosforilação , Proteína Fosfatase 1/metabolismo , Interferência de RNA , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Ratos Wistar , ATPase Trocadora de Sódio-Potássio/antagonistas & inibidores , ATPase Trocadora de Sódio-Potássio/genética , Vasoconstrição/efeitos dos fármacos , Vasoconstritores/farmacologia , Quinases da Família src/antagonistas & inibidoresRESUMO
Disturbances of circadian rhythms have been suggested to be a causal factor in the development of major depressive disorder. However, the mechanisms underlying the association between circadian rhythm abnormalities and mood disorders are still unknown. In the current study the association between diurnal pattern of key phase markers (melatonin, corticosterone, and core body temperature) and anhedonic-like behavior was investigated using the highly validated rat chronic mild stress (CMS) model of depression. Phase marker measurements were done after 3.5 weeks of CMS in 48 control rats and 48 anhedonic-like rats at 6 time points within 24h. The results showed that anhedonic-like behavior associates with changes in all three phase markers: an increased dark phase melatonin secretion, an additional peak in corticosterone level in the beginning of the light phase, and hypothermia in the dark phase. The result adds to the validity of the CMS model in general and in particular to be adequate as a model for studying the chronobiology of depressive disorder.