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Orientador: Antonio Carlos Cicogna / Resumo: Introdução: A remodelação cardíaca compreende mudanças no fenótipo cardíaco, determinadas por alterações na expressão gênica em resposta a estresse biomecânico de etiologia fisiológica ou patológica. A RC, eventualmente, pode evoluir para um quadro de disfunção e posteriormente insuficiência cardíaca grave (ICG); alterações no sistema β-adrenérgico podem estar envolvidas nesse processo. Medidas não farmacológicas, como o treinamento físico (TF) são empregadas no manejo das cardiopatias, e entre os efeitos benéficos dessa terapia está a modulação do sistema β-adrenérgico. Apesar das evidencias de que o TF impacta positivamente a sinalização β-adrenérgica e a função cardíaca, não encontramos na literatura trabalhos que avaliaram a associação entre treinamento físico aeróbio de baixa intensidade, sistema β-adrenérgico e função cardíaca em ratos com sobrecarga pressórica e disfunção ventricular. Objetivo: Testar a hipótese de que o TF atenua as alterações β-adrenérgicas miocárdicas e a deterioração do desempenho cardíaco em ratos com sobrecarga pressórica e disfunção ventricular previamente instalada. Métodos: Ratos Wistar machos (70-90 g), submetidos à cirurgia de estenose aórtica supravalvar (EAo), foram inicialmente divididos em dois grupos: Controle operado (Sham) e estenose aórtica (EAo). Após 18 semanas do procedimento cirúrgico, foi realizada análise da função cardíaca para redistribuição dos grupos: não expostos ao treinamento físico (Sham, n= 18 e EAo, ... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Introduction: Cardiac remodeling comprises changes in the cardiac phenotype, determined by changes in genic expression in response to biomechanical stress of physiological or pathological etiology. CR can eventually evolve into a condition of dysfunction and subsequently severe heart failure (GCI); Alterations in the β- adrenergic system may be involved in this process. Non-pharmacological measures such as physical training (PT) are used in the management of heart diseases, and among the beneficial effects of this therapy is the β-adrenergic system modulation. Despite the evidence that TF has a positive effect on β-adrenergic signaling and cardiac function, we did not find in the literature studies that evaluate the association between low-intensity aerobic physical exercise, β-adrenergic system and cardiac function in rats with pressure overload and ventricular dysfunction. Objective: To test the hypothesis that PT attenuates myocardial β-adrenergic alterations and deterioration of cardiac performance in rats with pressure overload and previously installed ventricular dysfunction. METHODS: Male Wistar rats (70-90 g) submitted to supravalvular aortic stenosis surgery (AS), were initially divided into two groups: Operated control (Sham) and aortic stenosis (AS). After 18 weeks of the surgical procedure, cardiac function analysis was performed for redistribution of the groups: non-exposed to physical training (Sham, n = 18 and AS, n = 18) and trained (TFSham, ... (Complete abstract click electronic access below) / Mestre

Submitted by SÉRGIO LUIZ BORGES DE SOUZA null (enfeborges@gmail.com) on 2017-05-10T16:06:02Z No. of bitstreams: 1 SOUZA, SLB DISSERTAÇÃO DE MESTRADO.pdf: 3472837 bytes, checksum: 674f72f28688f4341f5c0af589d80f98 (MD5) / Approved for entry into archive by Luiz Galeffi (luizgaleffi@gmail.com) on 2017-05-10T16:34:14Z (GMT) No. of bitstreams: 1 souza_slb_me_bot.pdf: 3472837 bytes, checksum: 674f72f28688f4341f5c0af589d80f98 (MD5) / Made available in DSpace on 2017-05-10T16:34:14Z (GMT). No. of bitstreams: 1 souza_slb_me_bot.pdf: 3472837 bytes, checksum: 674f72f28688f4341f5c0af589d80f98 (MD5) Previous issue date: 2017-02-21 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Introdução: A remodelação cardíaca compreende mudanças no fenótipo cardíaco, determinadas por alterações na expressão gênica em resposta a estresse biomecânico de etiologia fisiológica ou patológica. A RC, eventualmente, pode evoluir para um quadro de disfunção e posteriormente insuficiência cardíaca grave (ICG); alterações no sistema β-adrenérgico podem estar envolvidas nesse processo. Medidas não farmacológicas, como o treinamento físico (TF) são empregadas no manejo das cardiopatias, e entre os efeitos benéficos dessa terapia está a modulação do sistema β-adrenérgico. Apesar das evidencias de que o TF impacta positivamente a sinalização β-adrenérgica e a função cardíaca, não encontramos na literatura trabalhos que avaliaram a associação entre treinamento físico aeróbio de baixa intensidade, sistema β-adrenérgico e função cardíaca em ratos com sobrecarga pressórica e disfunção ventricular. Objetivo: Testar a hipótese de que o TF atenua as alterações β-adrenérgicas miocárdicas e a deterioração do desempenho cardíaco em ratos com sobrecarga pressórica e disfunção ventricular previamente instalada. Métodos: Ratos Wistar machos (70-90 g), submetidos à cirurgia de estenose aórtica supravalvar (EAo), foram inicialmente divididos em dois grupos: Controle operado (Sham) e estenose aórtica (EAo). Após 18 semanas do procedimento cirúrgico, foi realizada análise da função cardíaca para redistribuição dos grupos: não expostos ao treinamento físico (Sham, n= 18 e EAo, n= 18) e treinados (ShamTF, n= 23 e EAoTF, n= 26) durante 10 semanas. O protocolo de TF foi complementado com teste de esforço para ajuste de carga e avaliação da capacidade funcional, aplicado antes do início do programa de treinamento, após a 3ª e 7ª semana e ao fim do protocolo. A remodelação cardíaca foi caracterizada in vivo pela análise estrutural e funcional por ecocardiograma na 18ª e 28ª semana, in vitro, pelo músculo papilar isolado e pelo estudo macroscópico post mortem. O sistema β-adrenérgico foi avaliado pela expressão proteica dos β-adrenoceptores β-1 e β-2, adenilato ciclase e proteína quinase A, e pela manobra com agonista β-adrenérgico, isoproterenol, no músculo papilar isolado. Resultados: Na 18ª semana, antes de iniciar o TF, os animais EAo apresentavam hipertrofia ventricular esquerda concêntrica, disfunção diastólica e sistólica e prejuízo da capacidade funcional. Após o período de TF, os grupos ShamTF e EAoTF apresentaram aumento da capacidade funcional, visualizado pela tolerância e desempenho no exercício, e redução na ocorrência e intensidade dos sinais de insuficiência cardíaca. Na 28ª semana, o grupo EAo manteve a disfunção diastólica e sistólica, enquanto que o grupo EAoTF mostrou atenuação do prejuízo diastólico em ambas as análises, in vivo e in vitro. Não houve diferença entre os grupos na expressão proteica dos componentes do sistema β-adrenérgico; entretanto, os grupos EAo e EAoTF mostraram capacidade reduzida de resposta à estimulação por isoproterenol in vitro. Conclusão: Em conclusão, a exposição ao treinamento físico atenuou a disfunção diastólica e não alterou os os níveis proteicos dos componentes do sistema β-adrenérgico miocárdico, que não se modificaram com o processo de remodelação cardíaca. / Introduction: Cardiac remodeling comprises changes in the cardiac phenotype, determined by changes in genic expression in response to biomechanical stress of physiological or pathological etiology. CR can eventually evolve into a condition of dysfunction and subsequently severe heart failure (GCI); Alterations in the β- adrenergic system may be involved in this process. Non-pharmacological measures such as physical training (PT) are used in the management of heart diseases, and among the beneficial effects of this therapy is the β-adrenergic system modulation. Despite the evidence that TF has a positive effect on β-adrenergic signaling and cardiac function, we did not find in the literature studies that evaluate the association between low-intensity aerobic physical exercise, β-adrenergic system and cardiac function in rats with pressure overload and ventricular dysfunction. Objective: To test the hypothesis that PT attenuates myocardial β-adrenergic alterations and deterioration of cardiac performance in rats with pressure overload and previously installed ventricular dysfunction. METHODS: Male Wistar rats (70-90 g) submitted to supravalvular aortic stenosis surgery (AS), were initially divided into two groups: Operated control (Sham) and aortic stenosis (AS). After 18 weeks of the surgical procedure, cardiac function analysis was performed for redistribution of the groups: non-exposed to physical training (Sham, n = 18 and AS, n = 18) and trained (TFSham, n = 23 and PTAS, n = 26) for 10 weeks. The PT protocol was complemented with an exercise test for load adjustment and functional capacity assessment, applied before the start of the training program, after the 3rd and 7th week and at the end of the protocol. Cardiac remodeling was characterized in vivo by structural and functional analysis by echocardiogram at the 18th and 28th week, in vitro by the isolated papillary muscle and by the macroscopic post mortem study. The β-adrenergic system was evaluated by protein expression of β-1 and β-2 adrenoceptors, adenylate cyclase and protein kinase A, and by adding the isoproterenol β-adrenergic agonist in the isolated papillary muscle. Results: At the 18th week, before starting PT, the AS animals presented concentric left ventricular hypertrophy, diastolic and systolic dysfunction and impairment of functional capacity. After the PT period, the PTSham and PTAS groups showed increased functional capacity, visualized by tolerance and exercise performance, and reduction in the occurrence and intensity of the signs of heart failure. At week 28, the AS group maintained diastolic and systolic dysfunction, whereas the PTAS group showed attenuation of diastolic damage in both analyzes, both in vivo and in vitro. There was no difference between groups in the protein expression of β- adrenergic system components; However, the AS and PTAS groups showed reduced ability to respond to isoproterenol stimulation in vitro. Conclusion: In conclusion, exposure to physical training attenuated diastolic dysfunction and did not alter the protein levels of components of the myocardial β-adrenergic system, which did not change with the cardiac remodeling process.

Resumo: A ractopamina é um agonista β-adrenérgico que vem sendo utilizado como repartidor de energia em dietas de suínos na fase de terminação. Neste contexto, objetivou-se avaliar a influência de três níveis de inclusão deste aditivo (0, 5 e 10 ppm) na dieta de suínos (machos castrados e fêmeas, n=340) durante 28 dias pré abate, sobre o desempenho, características de carcaça e parâmetros econômicos. Não houve interação entre níveis de ractopamina e condição sexual para as variáveis avaliadas. Constatou-se que suínos que receberam ractopamina obtiveram ganho de peso diário superior e melhora na conversão alimentar. Com a inclusão de 10 ppm, foi verificada uma melhora na profundidade de músculo, redução na espessura de toucinho, aumento da porcentagem de carne magra e do rendimento de carcaça. A análise econômica aponta para aumento no valor da receita total e líquida com a venda dos suínos. A suplementação de ractopamina na dieta (5 e 10 ppm) promove melhorias no desempenho e nas características de carcaça de suínos em condições de produção comercial, e seu uso é economicamente viável na suinocultura / Abstract: The aim of this research was to evaluate the influence of three levels of inclusion of ractopamine (0, 5 and 10 ppm) in diet of finishing pigs (barrows and gilts, n=340) on performance, carcass traits and economical parameters. After 28 days on the feeding trial, pigs were slaughtered and the growth, carcass traits and economical analysis were measured. There was no interaction on levels of ractopamine and sexual condicion for variables evaluated. Ractopamine feeding increased (P<0.05) average daily gain and efficiency. Moreover, with inclusion of 10 ppm, there was an improvement in depth of muscle, reduced fat tissue, increased lean percentage and carcass yield. Economical analysis showed increase in total revenue value and net produce from swines. These results suggest that ractopamine will improve performance and carcass traits of finishing pigs breeding under commercial conditions supplemented with 5 and 10 ppm and it is economically viable / Orientador: Roberto de Oliveira Roça / Coorientador: Osmar Antonio Dalla Costa / Banca: Dirlei Antonio Berto / Banca: Jacinta Diva Ferrugem Gomes / Mestre

Succinate is a dicarboxylic acid that accumulates due to succinate dehydrogenase inhibition. This dicarboxylic acid has a biphasic effect on neural activity in vitro that seems to be mediated by N-methyl-D-aspartate (NMDA) receptors. The NMDA receptor is distributed throughout the central nervous system and mediates synaptic plasticity-related events, such as learning and memory. Although it has been described that succinate modulates NMDA conductance, it is not known if this organic acid modulates learning and me mory. Therefore, in the present study we investigated whether the immediate post-training systemic or intrahippocampal administration of succinate affects the memory of fear conditioning in rats. In addition, we investigated whether the NMDA and β-adrenergic receptors are involved in the facilitatory effect of succinate on memory. Systemic (0,00005, 0,0005, 0,005, 0,05 and 0,5 mg/kg i.p) or bilateral intra-hippocampal (0.21 pmol i.h.) immediate post-training administration of succinate biphasically facilitated contextual fear conditioning and had no effect on conditioning to tone. Systemic or intra-hippocampal administrations of MK-801 (0,001 mg/kg i.p. or 0,21 pmol i.h. respectively), a noncompetitive NMDA receptor antagonist, at a dose that had no effect per se, reversed the facilitatory effect of succinate on contextual fear conditioning. The systemic administration of propranolol (10 mg/kg i.p.), a β-adrenergic antagonist, at a dose that had no effect per se, also reversed the facilitatory effect of succinate on contextual fear conditioning. The administration of succinate (0,005 mg/kg i.p.) immediately after training and 15 minutes before the test did not affect the performance of the animals on fear conditioning. These results suggest that the facilitatory effect of succinate on the memory of fear conditioning involves NMDA and β-adrenergic receptors. The results also indicate that the facilitatory effect of succinate on memory is not related to state-dependence / O sucinato é um intermediário do ciclo de Kreks e também da cadeia respiratória, mas que também parece ter funções não relacionadas ao metabolismo energético. De fato, este ácido dicarboxílico tem efeito bifásico sobre a atividade neural in vitro, que parece ser mediado pelo receptor glutamatérgico N-metil-D-aspartato (NMDA). O receptor NMDA está presente em todo o sistema nervo central (SNC) e os processos mediados por este receptor incluem plasticidade sináptica e formação de circuitos neurais. Embora tenha sido descrito que o sucinato modula a atividade neural in vitro, não se sabe se este ácido orgânico modula processos fisiológicos ligados ao receptor NMDA, como o aprendizado e memória. Consequentemente, neste estudo foi investigado o efeito da administração sistêmica e intrahipocampal de sucinato, MK-801 e propranolol sobre a consolidação da memória em ratos, utilizando a tarefa de medo condicionado clássico. Posteriormente se investigou se os efeitos do sucinato sobre a memória envolvem dependência de estado. O teste do medo condicionado consistiu na apresentação de estímulos pareados, condicionado (tom 2000 Hz 90 dB/10 s) e incondicionado (choque 0.6 mA/1 s). O estado de imobilidade do animal foi utilizado como um indicativo de memória nas sessões de avaliação de memória ao contexto e ao tom. A administração sistêmica de sucinato nas doses de 0,00005, 0,0005, 0,005, 0,05 e 0,5 mg/kg i.p. imediatamente após o treino melhorou a memória. Contudo, a dose de 5 mg/kg de sucinato não alterou a memória dos animais, caracterizando um efeito bifásico sobre a memória. Essa melhora da memória induzida por sucinato (0,005 mg/kg i.p.) foi revertido pela administração pós-treino de um antagonista do receptor NMDA, MK-801 (0,001 mg/kg i.p.) e pela administração imediatamente após o treino de um bloqueador adrenérgico, propranolol (10 mg/kg i.p.). A administração de sucinato (0,005 mg/kg i.p.) imediatamente após o treino e 15 minutos antes do teste não afetou a performance dos animais, não caracterizando dependência de estado. Nos experimentos que visaram identificar se a administração central de sucinato alterava a memória, os animais foram canulados bilateralmente no hipocampo e após a recuperação cirúrgica, recebiam injeções bilaterais imediatamente após o treino, de sucinato (0,21 pmol i.h.) que melhorou a memória. A administração imediatamente após o treino de MK- 801 (0,22 nmol i.h.) foi capaz de reverter esse efeito facilitador do sucinato sobre a memória. Estes resultados sugerem que o efeito facilitador da memória induzida pela administração de sucinato é mediado pelo receptor NMDA e envolve de alguma maneira o sistema adrenérgico

Acute cold exposure increases the risk of sudden cardiac events, similarly exposure to constant light negatively affects the cardiovascular system. However, the individual effects of these factors and the effect of their combination on cardiomyocytes are not yet known. The thesis deals with the influence of a 3 day cold exposure and constant light on the expression of β-adrenergic receptors and associated G-proteins in association with apoptotic signals in the left ventricle of the Wistar rat heart. In this work apoptotic proteins BAX, BCL2, caspase 8 and important components of β-adrenergic signalization - β1, β2, G-proteins, Gas, Gi1/2 and Gi3 were determined. The relative expression of the proteins was analyzed by Western blotting. The results confirm the detrimental effect of cold and light exposure. However, the synergistic effect of these two stressors shows surprising results.

It has recently been discovered that the effect of morphine can significantly reduce the tissue damage that occurs during myocardial ischemia. The molecular mechanisms by which morphine acts on the heart are still little understood. The aim of this thesis was to monitor the effect of chronic 27-day and 10-day administration of low (1 mg/kg/day) and high (10 mg/kg/ day) doses of morphine on the expression of selected G-protein-coupled receptors (GPCR) and on the expression and activity of adenylyl cyclase (AC). Chronic (27 days) morphine treatment reduced the expression of к-opioids receptors, but 10-day morphine exposure did not influence the expression of these receptors. Assessment of β1- and β2-AR by immunoblot technique did not show any significant change in the expression, but the more accurate determination of β-AR expression using the saturation binding studies revealed that 27-day treatment with high doses of morphine appreciable increased the total number of these receptors. Administration of high doses of morphine led to marked up-regulation of adenylyl cyclase (AC) isoforms V/VI, and the amount of AC decreased proportionally with the time of discontinuation of morphine administration. Low doses of morphine up- regulated AC only during 27-day administration. Chronic morphine exposure did...

Les catécholamines induisent la synthèse d’AMPc par une stimulation des récepteurs β-adrénergiques et contrôlent ainsi la fonction cardiaque en activant une pléiade de voies de signalisation intracellulaires. Les protéines Epac sont des facteurs d’échange pour les petites protéines G et sont directement activés par l’AMPc. Devant l’importance de la voie β-adrénergique dans la physiopathologie cardiaque et dans le but de mieux comprendre la régulation des processus cellulaires dépendants de l’AMPc dans le cœur, il apparaît essentiel de caractériser le rôle des facteurs d’échange Epac dans le myocarde. Dans une première partie, cette étude démontre que les effets de Epac sur l’hypertrophie des cardiomyocytes ventriculaires de rats nouveaux nés requièrent les GTPases H-Ras et Rap2B. Epac active la voie PLC/IP3/Ca2+ qui est nécessaire pour l’activation de H-Ras. Au niveau transcriptionnel, Epac induit l’export nucléaire de HDAC4 permettant l’activation d’un programme génique d’hypertrophie. Dans une deuxième partie, cette étude révèle l’implication de Epac1 dans l’hypertrophie des cardiomyocytes in vivo, chez la souris. La délétion de Epac1 protège du remodelage cardiaque induit par l’activation prolongée des récepteurs β-adrénergiques et améliore la fonction cardiaque. La surexpression de Epac1 spécifiquement dans le myocarde entraîne une hypertrophie des cardiomyocytes. Par ailleurs, la voie β-AR/Epac1 induit l’accumulation de protéines ubiquitinylées et provoque l’activation du processus d’autophagie in vitro et in vivo. L’autophagie protège des effets délétères de la voie β-adrénergique/Epac en participant à l’élimination des agrégats protéiques et en contrant les effets hypertrophiques de Epac1. Ces résultats ouvrent de nouvelles perspectives pour le traitement de l’hypertrophie et de l’insuffisance cardiaque. / Catecholamines regulate cardiac function by stimulating β-adrenergic receptors (β-AR), leading to cAMP production and activation of a multiplicity of signaling pathways. Epac proteins are exchange factors for small G proteins which are directly activated by cAMP. Given the importance of the β-adrenergic pathway in cardiac physiopathology, it becomes essential to characterize functions of Epac protein in myocardium. In a first part, this study shows that H-Ras and Rap2B GTPases are involved in Epac-induced neonatal rat cardiac myocytes hypertrophy. Epac induces activation of the PLC/IP3/Ca2+ pathway which is necessary for H-Ras activation. At the transcriptional level, Epac causes HDAC4 nuclear export leading to activation of a hypertrophic gene program. In a second part, this study reveals implication of Epac1 in cardiac hypertrophy in vivo. Deletion of Epac1 in mice protects from cardiac remodeling induced by chronic isoproterenol infusion and enhances cardiac function. Cardiac specific overexpression of Epac1 in mice induces cardiac myocytes hypertrophy. Interestingly, β-AR/Epac1 pathway triggers ubiquitinated proteins accumulation and activation of autophagy both in vitro and in vivo. By eliminating aggregates and by counteracting hypertrophic effects of Epac, autophagy protects from deleterious effects of the β-AR/Epac pathway. These results open news insights into the treatment of cardiac hypertrophy and heart failure.

The cost of lamb mortality caused by cold exposure has been estimated at approximately 40 million dollars per year. This value is probably conservative as it does not include the cost due to the reduction in productivity in hypothermic lambs that manage to survive or the cost of reduced selection potential incurred by fewer lambs surviving until selection. The objectives of this research was to investigate whether sire-line variation exists in neonatal lamb cold tolerance and whether polymorphism in the β₃ adrenergic receptor gene can be used as a genetic marker for lamb cold tolerance and lean muscle growth. The influence of the climate, birthweight, age of dam at lambing, gender and birth rank on neonatal lamb cold tolerance was also analysed. Neonatal lamb mortality due to cold exposure was analysed in four field trials that used neonatal lamb morality from cold exposure as a predictor of neonatal lamb cold tolerance. Sire-line variation in neonatal lamb morality was observed in all trials, though it appeared that this effect was largely mediated through sire-line variation in lamb birth weight. Variation in lamb birth weight between birth rank classed was also found to be responsible for the influence of birth rank on neonatal lamb mortality due to cold exposure. The age of dam at lambing and the lamb gender was not observed to influence neonatal lamb mortality due to cold exposure. The sires from the cold tolerance study and the progeny of the lean muscle growth study were genotyped for the β₃ adrenergic receptor locus. Other studies have found evidence that a major gene exists in the catecholamine stimulation of brown adipose thermogenesis and evidence that the β₃-AR gene is a likely candidate. However, this hypothesis and the hypothesis that polymorphism in the β₃-AR gene is also linked to lean muscle growth in lambs was not confirmed in this study. So while it appears that the results were confounded by experimental design, there is evidence that influence of polymorphism in the ovine β₃ AR gene on neonatal lamb mortality and/or lean muscle growth is not sufficient to be considered a major gene effect. The implications of this experiment on the sheep industry and sheep farmers in general are huge. While completely eliminating lamb deaths due to inadequate cold tolerance is impossible, this study shows that large gains in lamb survival could be possible through selective breeding.

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