Allosteric Control of Gating Mechanisms Revisited: The Large Conductance Ca(2+)-Activated K(+) Channel

Large-conductance Ca(2+)-activated K(+) channels (BK) play a fundamental role in modulating membrane potential in many cell types. The gating of BK channels and its modulation by Ca(2+) and voltage has been the subject of intensive research over almost three decades, yielding several of the most complicated kinetic mechanisms ever proposed. A large number of open and closed states disposed, respectively, in two planes, named tiers, characterize these mechanisms. Transitions between states in the same plane are cooperative and modulated by Ca(2+). Transitions across planes are highly concerted and voltage-dependent. Here we reexamine the validity of the two-tiered hypothesis by restricting attention to the modulation by Ca(2+). Large single channel data sets at five Ca(2+) concentrations were simultaneously analyzed from a Bayesian perspective by using hidden Markov models and Markov-chain Monte Carlo stochastic integration techniques. Our results support a dramatic reduction in model complexity, favoring a simple mechanism derived from the Monod-Wyman-Changeux allosteric model for homotetramers, able to explain the Ca(2+) modulation of the gating process. This model differs from the standard Monod-Wyman-Changeux scheme in that one distinguishes when two Ca(2+) ions are bound to adjacent or diagonal subunits of the tetramer.

Serotonergic mechanisms of the median raphe nucleus-dorsal hippocampus in conditioned fear: Output circuit involves the prefrontal cortex and amygdala

Independent studies have shown that the median raphe nucleus (MRN) and dorsal hippocampus (DH) are involved in the expression of contextual conditioned fear (CFC). However, studies that examine the integrated involvement of serotonergic mechanisms of the MRN-DH are lacking. To address this issue, a CFC paradigm was used to test whether the serotonergic projections from the MRN to DH can influence CFC. Serotoninergic drugs were infused either into the MRN or DH prior to testing sessions in which freezing and startle responses were measured in the same context where 6 h previously rats received footshocks. A reduction of serotonin (5-HT) transmission in the MRN by local infusions of the 5-HT(1A) agonist 8-hydroxy-2-(di-n-propylamino)-tetralin (8-OH-DPAT) decreased freezing in response to the context but did not reduce fear-potentiated startle. This pattern of results is consistent with the hypothesis that MRN serotonergic mechanisms selectively modulate the freezing response to the aversive context. As for the DH, a decrease in postsynaptic 5-HT receptor activity at projection areas has been proposed to be the main consequence of 5-HT(1A) receptor activation in the MIRN. Intra-DH injections of 8-OH-DPAT inhibited both the freezing and fear-potentiated startle response to the context. To reconcile these findings, an inhibitory mechanism may exist between the incoming 5-HT pathway from the MRN to DH and the neurons of the DH output to other structures. The DH-amygdala or medial prefrontal cortex projections could well be this output circuit modulating the expression of CFC as revealed by measurements of Fos immunoreactivity in these areas. (C) 2009 Elsevier B.V. All rights reserved.

Prion protein ablation increases cellular aggregation and embolization contributing to mechanisms of metastasis

Cellular Prion Protein (PrP(C)) is a cell surface protein highly expressed in the nervous system, and to a lesser extent in other tissues. PrP(C) binds to the extracellular matrix laminin and vitronectin, to mediate cell adhesion and differentiation. Herein, we investigate how PrP(C) expression modulates the aggressiveness of transformed cells. Mesenchymal embryonic cells (MEC) from wildtype (Prnp(+/+)) and PrP(C)-null (Prnp(0/0)) mice were immortalized and transformed by co-expression of ras and myc. These cells presented similar growth rates and tumor formation in vivo. When injected in the tail vein, PrnP(0/0)raS/myc cells exhibited increased lung colonization compared with Prnp(+/+)ras/myc cells. Additionally, Prnp(0/0)ras/myc cells form more aggregates with blood components than Prnp(+/+)ras/myc cells, facilitating the arrest of Prnp(0/0)ras/myc cells in the lung vasculature. Integrin alpha(v)beta(3) is more expressed and activated in MEC and in transformed Prnp(0/0) cells than in the respective Prnp(+/+) cells. The blocking of integrin alpha(v)beta(3) by RGD peptide reduces lung colonization in transformed Prnp(0/0) cells to similar levels of those presented by transformed Prnp(+/+) cells. Our data indicate that PrP(C) negatively modulates the expression and activation of integrin alpha(v)beta(3) resulting in a more aggressive phenotype. These results indicate that PrP(C) may have main implications in modulating metastasis formation. (C) 2009 UICC

Mechanisms mediating the diuretic and natriuretic actions of the incretin hormone glucagon-like peptide-1

Crajoinas RO, Oricchio FT, Pessoa TD, Pacheco BP, Lessa LM, Malnic G, Girardi AC. Mechanisms mediating the diuretic and natriuretic actions of the incretin hormone glucagon-like peptide-1. Am J Physiol Renal Physiol 301: F355-F363, 2011. First published May 18, 2011; doi: 10.1152/ajprenal.00729.2010.-Glucagon-like peptide-1 (GLP-1) is a gut incretin hormone considered a promising therapeutic agent for type 2 diabetes because it stimulates beta cell proliferation and insulin secretion in a glucose-dependent manner. Cumulative evidence supports a role for GLP-1 in modulating renal function; however, the mechanisms by which GLP-1 induces diuresis and natriuresis have not been completely established. This study aimed to define the cellular and molecular mechanisms mediating the renal effects of GLP-1. GLP-1 (1 mu g.kg(-1).min(-1)) was intravenously administered in rats for the period of 60 min. GLP-1-infused rats displayed increased urine flow, fractional excretion of sodium, potassium, and bicarbonate compared with those rats that received vehicle (1% BSA/saline). GLP-1-induced diuresis and natriuresis were also accompanied by increases in renal plasma flow and glomerular filtration rate. Real-time RT-PCR in microdissected rat nephron segments revealed that GLP-1 receptor-mRNA expression was restricted to glomerulus and proximal convoluted tubule. In rat renal proximal tubule, GLP-1 significantly reduced Na(+)/H(+) exchanger isoform 3 (NHE3)-mediated bicarbonate reabsorption via a protein kinase A (PKA)-dependent mechanism. Reduced proximal tubular bicarbonate flux rate was associated with a significant increase of NHE3 phosphorylation at the PKA consensus sites in microvillus membrane vesicles. Taken together, these data suggest that GLP-1 has diuretic and natriuretic effects that are mediated by changes in renal hemodynamics and by downregulation of NHE3 activity in the renal proximal tubule. Moreover, our findings support the view that GLP-1-based agents may have a potential therapeutic use not only as antidiabetic drugs but also in hypertension and other disorders of sodium retention.

Myofascial trigger point: A possible way of modulating tinnitus

In order to investigate whether myofascial trigger points can modulate tinnitus, as well as the association between tinnitus and myofascial trigger points, 94 individuals with and 94 without tinnitus, matched by age and gender, were analyzed by means of bilateral digital pressure of 9 muscles. Temporary modulation of tinnitus was frequently observed (55.9%) during digital pressure, mainly in the masseter. The rate of tinnitus modulation was significantly higher on the same side of the myofascial trigger point subject to examination in 6 out of 9 muscles. An association between tinnitus and the presence of myofascial trigger points was observed (p < 0.001), as well as a laterality association between the ear with the worst tinnitus and the side of the body with more myofascial trigger points (p < 0.001). Thus, this relationship could be explained not only by somatosensory-auditory system interactions but also by the influence of the sympathetic system. Copyright (c) 2007 S. Karger AG, Basel.

Mechanisms of pigmentation loss in subterranean fishes

Troglobitic (exclusively subterranean) organisms usually present, among their apomorphies related to the subterranean life (troglomorphisms), the regression of eyes and melanic pigmentation. The degree of regression varies among species, from a slight reduction to the complete loss of eyes and dark pigmentation, without a taxonomic correlation. While mechanisms of eye reduction have been intensively investigated in some troglobites such as the Mexican blind tetra characins, genus Astyanax, and the European salamander, Proteus anguinus, few studies have focused on pigmentation. The Brazilian subterranean ichthyofauna distinguishes not only by the species richness (23 troglobitic fishes so far known) but also by the variation in the degree of reduction of eyes and pigmentation. This study focused on Brazilian fishes completely devoid of melanic pigmentation: the characiform Stygichthys typhlops (Characidae) and the siluriforms Ancistrus formoso (Loricariidae), Rhamdiopsis sp.1 (Heptapteridae; from caves in the Chapada Diamantina, Bahia) and Rhamdiopsis sp. 2 (cave in Campo Formoso, Bahia). In order to investigate if such depigmentation is the result of blockage in some step in the melanogenesis, in vitro tests of administration of L-DOPA were done, using caudal-fin fragments extracted from living fish. Except for Rhamdiopsis sp. 2, all the studied species were DOPA(+), i.e., melanin was synthesized after L-DOPA administration. This indicates these fish do have melanophores but they are unable to convert L-tyrosine to L-DOPA. On the other hand, Rhamdiopsis sp. 2, like the albino specimens of Trichomycterus itacarambiensis previously studied (which correspond to one third of the population), are DOPA(-), either because the block of melanin synthesis occurs downstream in melanogenesis, which is probably the case with T. itacarambiensis (monogenic system in view of the phenotypic discontinuity), or because the so-called albinos do no possess melanophores. The physiological loss in the ability to synthesize melanin, apparently caused by different genetic processes in DOPA(+) and in DOPA(-) fishes, may co-exist in subterranean populations with a decrease in the density of melanophores, as observed in the pigmented two thirds of T. itacarambiensis population, a morphological reduction apparently controlled by polygenic systems producing a continuous phenotypic variation.

Central chemoreceptors and neural mechanisms of cardiorespiratory control

The arterial partial pressure (P CO2) of carbon dioxide is virtually constant because of the close match between the metabolic production of this gas and its excretion via breathing. Blood gas homeostasis does not rely solely on changes in lung ventilation, but also to a considerable extent on circulatory adjustments that regulate the transport of CO2 from its sites of production to the lungs. The neural mechanisms that coordinate circulatory and ventilatory changes to achieve blood gas homeostasis are the subject of this review. Emphasis will be placed on the control of sympathetic outflow by central chemoreceptors. High levels of CO2 exert an excitatory effect on sympathetic outflow that is mediated by specialized chemoreceptors such as the neurons located in the retrotrapezoid region. In addition, high CO2 causes an aversive awareness in conscious animals, activating wake-promoting pathways such as the noradrenergic neurons. These neuronal groups, which may also be directly activated by brain acidification, have projections that contribute to the CO2-induced rise in breathing and sympathetic outflow. However, since the level of activity of the retrotrapezoid nucleus is regulated by converging inputs from wake-promoting systems, behavior-specific inputs from higher centers and by chemical drive, the main focus of the present manuscript is to review the contribution of central chemoreceptors to the control of autonomic and respiratory mechanisms.

Stress-induced neuroinflammation: mechanisms and new pharmacological targets

Stress is triggered by numerous unexpected environmental, social or pathological stimuli occurring during the life of animals, including humans, which determine changes in all of their systems. Although acute stress is essential for survival, chronic, long-lasting stress can be detrimental. In this review, we present data supporting the hypothesis that stress-related events are characterized by modifications of oxidative/nitrosative pathways in the brain in response to the activation of inflammatory mediators. Recent findings indicate a key role for nitric oxide (NO) and an excess of pro-oxidants in various brain areas as responsible for both neuronal functional impairment and structural damage. Similarly, cyclooxygenase-2 (COX-2), another known source of oxidants, may account for stress-induced brain damage. Interestingly, some of the COX-2-derived mediators, such as the prostaglandin 15d-PGJ2 and its peroxisome proliferator-activated nuclear receptor PPARγ, are activated in the brain in response to stress, constituting a possible endogenous anti-inflammatory mechanism of defense against excessive inflammation. The stress-induced activation of both biochemical pathways depends on the activation of the N-methyl-D-aspartate (NMDA) glutamate receptor and on the activation of the transcription factor nuclear factor kappa B (NFκB). In the case of inducible NO synthase (iNOS), release of the cytokine TNF-α also accounts for its expression. Different pharmacological strategies directed towards different sites in iNOS or COX-2 pathways have been shown to be neuroprotective in stress-induced brain damage: NMDA receptor blockers, inhibitors of TNF-α activation and release, inhibitors of NFκB, specific inhibitors of iNOS and COX-2 activities and PPARγ agonists. This article reviews recent contributions to this area addressing possible new pharmacological targets for the treatment of stress-induced neuropsychiatric disorders.

Protector mechanisms of the association between gastroesophageal reflux disease and asthma: experimental study in rats

BACKGROUND: It is well known the association between gastroesophageal reflux disease and asthma. The hyperreactivity of the airways is a characteristic of an asthmatic. Many studies associate the increase of the airways reactivity with gastroesophageal reflux disease. AIM: In this study we have evaluated the effect of the intraluminal exposition to gastric juice of trachea on the reactivity to methacholine from rats submitted to a pulmonary allergic inflammation. METHODS: Group of rats were sensitized and challenged with ovalbumin. After 24 hours the animals were sacrificed, and their tracheae were removed to be cultured with gastric juice. The gastric juice was obtained from a donor rat. Subsequently the segments were placed into plastic plates with RPMI-1640 for incubation, under suitable atmosphere and time. After the period of incubation the segments were put into chambers for the analysis of the contractile response to methacholine. RESULTS: We observed reduction in the contractile response of trachea cultured with gastric juice from allergic rats. This result was confirmed by the pharmacological treatments with compound 48/80 and dissodium cromoglicate (mast cells blockade), L-NAME (nitric oxide inhibitor, NO), capsaicin (neuropeptides depletion) and indomethacin (ciclooxigenase inhibitor). CONCLUSIONS: Our results highlight to the existence of a complex interaction between pulmonary allergy and gastric juice in the airways. The involvement of the non-adrenergic non-cholinergic system, NO, prostanoids and mast cells are directly related to this interaction. We suggest that the reduced contractile response observed in vitro may represent a protector mechanism of the airways. Despite its presence in the human body it can not be observed due to the predominant effects of excitatory the non-adrenergic non-cholinergic system.

Pode a atividade física materna modular a programação fetal induzida pela nutrição?

Existe considerável evidência para a indução de diferentes fenótipos em reposta às variações no ambiente fetal e neonatal. O aporte inadequado de nutrientes no período crítico do desenvolvimento está associado ao risco alto de doenças metabólicas na vida adulta, este fenômeno biológico é chamado de programação. A atividade física durante a gestação resulta em adaptações fisiológicas da mãe e no aumento da disponibilidade de nutrientes e oxigênio no espaço feto-placentário. Este trabalho tem como objetivo discutir os mecanismos da indução de programação fetal pela nutrição e o provável efeito modulador da atividade física durante a gestação. Foram utilizadas as bases de dados do Medline Pubmed, Lilacs e Bireme, com publicações entre 1990 até 2008. Os termos de indexação utilizados foram: nutrition, fetal programming, gestation, physical activity, physical exercise, metabolism. Em conclusão, o aporte inadequado de nutrientes programa o aparecimento de doenças metabólicas na vida adulta, enquanto que a atividade física durante a gestação aumenta a disponibilidade de nutrientes e oxigênio, repercutindo positivamente no crescimento fetal e no peso ao nascer.

Cyclic nitroxides inhibit the toxicity of nitric oxide-derived oxidants: mechanisms and implications

The substantial therapeutic potential of tempol (4-hydroxy-2,2,6,6-tetramethyl-1-piperidinyloxy) and related cyclic nitroxides as antioxidants has stimulated innumerous studies of their reactions with reactive oxygen species. In comparison, reactions of nitroxides with nitric oxide-derived oxidants have been less frequently investigated. Nevertheless, this is relevant because tempol has also been shown to protect animals from injuries associated with inflammatory conditions, which are characterized by the increased production of nitric oxide and its derived oxidants. Here, we review recent studies addressing the mechanisms by which cyclic nitroxides attenuate the toxicity of nitric oxidederived oxidants. As an example, we present data showing that tempol protects mice from acetaminophen-induced hepatotoxicity and discuss the possible protection mechanism. In view of the summarized studies, it is proposed that nitroxides attenuate tissue injury under inflammatory conditions mainly because of their ability to react rapidly with nitrogen dioxide and carbonate radical. In the process the nitroxides are oxidized to the corresponding oxammonium cation, which, in turn, can be recycled back to the nitroxides by reacting with upstream species, such as peroxynitrite and hydrogen peroxide, or with cellular reductants. An auxiliary protection mechanism may be down-regulation of inducible nitric oxide synthase expression. The possible therapeutic implications of these mechanisms are addressed.

Mecanismos de ação de compostos bioativos dos alimentos no contexto de processos inflamatórios relacionados à obesidade

É indiscutível o papel da dieta e dos alimentos na manutenção da saúde e na redução do risco de DCNT. Estudos epidemiológicos mostram que o aumento do consumo de alimentos de origem vegetal influencia positivamente a saúde, enquanto estudos in vitro e in vivo em modelo animal elucidam os mecanismos pelos quais compostos bioativos não nutrientes, presentes nos alimentos, atuam na manutenção da saúde e na redução do risco de doenças. A modulação da expressão de genes que codificam proteínas envolvidas em vias de sinalização celular ativadas em DCNT é um dos mecanismos de ação dos compostos bioativos, sugerindo que estes possam ser essenciais à manutenção da saúde. A biodisponibilidade dos compostos bioativos de alimentos, as suas rotas metabólicas e o modo de ação de seus metabólitos são importantes fatores no seu efeito nas DCNT. Todos esses aspectos são temas de investigações recentes, cujos resultados contribuem para a compreensão da ocorrência e desenvolvimento das DCNT e da sua relação com a dieta. Essa revisão visou discutir alguns dos mecanismos envolvidos na resposta inflamatória induzida pela obesidade, apresentar os compostos bioativos de alimentos que modulam essa resposta inflamatória e sua relação com o metabolismo desses compostos.

O uso da acupuntura no auxílio à terapia da doença idiopática do trato urinário inferior dos felinos

Diferentes afecções podem acometer o trato urinário inferior dos felinos, acarretando sinais clínicos inespecíficos como: hematúria, disúria, polaquiúria, estrangúria, periúria ou obstrução, caracterizando a doença do trato urinário inferior dos felinos (DTUIF). Entretanto, em até 65% dos felinos acometidos, a etiologia é indeterminada, denominada de doença idiopática do trato urinário inferior dos felinos (DTUIF idiopática), que tem se mostrado como um desafio ao clínico veterinário, uma vez que não há diagnóstico específico ou terapia efetiva. Atualmente, tem sido estudado o papel da inflamação neurogênica da bexiga urinária secundária ao estresse, como etiologia da DTUIF idiopática. Há evidências científicas de que a acupuntura restaure a homeostase, reduza o estresse e, pela estimulação neural periférica, ative mecanismos endógenos de antinocicepção, regulando a liberação de mediadores dos mecanismos da dor e do processo inflamatório, como a substância P. O objetivo desta revisão bibliográfica foi descrever como a acupuntura pode ser um recurso na terapia da DTUIF idiopática, tendo como base a modulação da inflamação neurogênica da bexiga urinária e o controle do estresse desses gatos.

Mechanisms of Vascular Damage by Hemorrhagic Snake Venom Metalloproteinases: Tissue Distribution and In Situ Hydrolysis

Background: Envenoming by viper snakes constitutes an important public health problem in Brazil and other developing countries. Local hemorrhage is an important symptom of these accidents and is correlated with the action of snake venom metalloproteinases (SVMPs). The degradation of vascular basement membrane has been proposed as a key event for the capillary vessel disruption. However, SVMPs that present similar catalytic activity towards extracellular matrix proteins differ in their hemorrhagic activity, suggesting that other mechanisms might be contributing to the accumulation of SVMPs at the snakebite area allowing capillary disruption. Methodology/Principal Findings: In this work, we compared the tissue distribution and degradation of extracellular matrix proteins induced by jararhagin (highly hemorrhagic SVMP) and BnP1 (weakly hemorrhagic SVMP) using the mouse skin as experimental model. Jararhagin induced strong hemorrhage accompanied by hydrolysis of collagen fibers in the hypodermis and a marked degradation of type IV collagen at the vascular basement membrane. In contrast, BnP1 induced only a mild hemorrhage and did not disrupt collagen fibers or type IV collagen. Injection of Alexa488-labeled jararhagin revealed fluorescent staining around capillary vessels and co-localization with basement membrane type IV collagen. The same distribution pattern was detected with jararhagin-C (disintegrin-like/cysteine-rich domains of jararhagin). In opposition, BnP1 did not accumulate in the tissues. Conclusions/Significance: These results show a particular tissue distribution of hemorrhagic toxins accumulating at the basement membrane. This probably occurs through binding to collagens, which are drastically hydrolyzed at the sites of hemorrhagic lesions. Toxin accumulation near blood vessels explains enhanced catalysis of basement membrane components, resulting in the strong hemorrhagic activity of SVMPs. This is a novel mechanism that underlies the difference between hemorrhagic and non-hemorrhagic SVMPs, improving the understanding of snakebite pathology.