L'érythropoïétine (EPO) est une cytokine ayant un rôle important dans l'homéostasie de l'oxygène (O2). Lors d'une hypoxie chronique, l'EPO stimule la maturation des progéniteurs érythroïdes en globules rouges augmentant ainsi le transport de l'O2 aux tissus. Outre cet effet érythropoïétique, l'EPO module la réponse ventilatoire à l'hypoxie (RVH) par une action directe sur la commande centrale respiratoire (CCR) et les chémorécepteurs périphériques. Cet effet a été principalement caractérisé chez des souris mutantes surexprimant l'EPO. Cependant, plusieurs aspects de l'effet de l'EPO sur l'activité du réseau respiratoire demeurent inconnus. Nos résultats montrent qu'une application aigüe d'EPO diminue la dépression centrale hypoxique mesurée in vitro chez le nouveau-né. En revanche, elle n'affecte pas la RVH mesurée in vivo au cours du développement postnatal mais diminue la fréquence des apnées survenant en hypoxie sévère à 6% d'O2. Aussi, chez la souris adulte, l'administration chronique d'EPO et de C-EPO augmente la sensibilité des chémorécepteurs périphériques à l'O2 et maintient la ventilation durant la phase tardive de la RVH. Enfin, l'EPO diminue la sensibilité ventilatoire à l'hypercapnie grâce à des effets périphériques et centraux. L'ensemble de nos résultats montrent que l'EPO module la respiration et contribue à l'homéostasie de l'O2 et du CO2 grâce à ses effets plasmatiques et centraux. Elle représente un candidat à fort potentiel thérapeutique pour les pathologies respiratoires où la sensibilité chimique à l'O2 et au CO2 sont altérés telles que l'apnée du nouveau-né ou le mal chronique des montagnes. / Erythropoietin (EPO) is a cytokine that plays a major role in O2 homeostasis. Upon chronic hypoxia, EPO stimulates the maturation of erythroid progenitors into red blood cells, contributing to increased O2 carrying to tissues. Besides this well-known erythropoietic effect, EPO also modulates the respiratory response to hypoxia by interacting with the central respiratory network in the brainstem and the peripheral chemoreceptors. This effect was mainly characterized in adult mutant mice that overexpress EPO. Several aspects regarding EPO's effect on breathing regulation remain unknown. Our results show that acute EPO treatment increases the O2 sensitivity of the central respiratory network in newborn mice in vitro. However, EPO does not impact the hypoxic ventilatory response to hypoxia in vivo, but decreases the apneic events during severe hypoxia in mice at postnatal day 7. In WT adults, chronic but not acute EPO and C-EPO treatment increases the O2 sensitivity by stimulating both peripheral chemoreceptor and central respiratory network. Finally, both cerebral and plasmatic EPO blunt the ventilatory response to increased CO2 levels in adult mice. Taken together, these results imply that EPO, by acting on the ventilatory system, plays a key role in the modulation of the chemical sensitivity to O2 and CO2.

A dispnéia na insuficiência cardíaca(IC) é complexa, com possível envolvimento de quimioreceptores periféricos(QP) e centrais(QC). Avaliamos a resposta de QP e QC no exercício submáximo em 15 pcts com IC e 7 ind. normais em testes ergoespirométricos de caminhada de 6 min: hipóxia isocápnica(HPX),hipercapnia hiperóxica(HPC) e ar ambiente. HPX aumentou ventilação (VE) com resposta aguda(RVA), freq. cardíaca(FC) e volume de O2 consumido;reduziu o espaço morto,distância caminhada(DC) e pressão arterial sistêmica(PAS). A HPC aumentou VE acima da HPX com RVA.Os QP têm ação maior sobre FC e PAS do que QC, apesar da maior ativação simpática.QP possuem estimulo rápido sobre VE,porém menor do que QC. / Heart failure(HF) dyspnea is complex with potential enrolment of central(CC) and peripheric chemoreceptors(PC).We investigated CC and CP behavior through submaximal exercise in 15 HF patients and 7 normal subjects in treadmill 6-minute cardiopulmonary walking tests:isocapnic hypoxia(HPO), hypercapnia hyperoxic(HCP) and room air.HPO increased:ventilation(VE) with acute ventilatory response(AVR), heart rate (HR) and O2 uptake and reduced dead space, distance walked (DW) and systemic blood pressure(SBP).The HPC improved VE above HPO level with AVR. PC have greater action on HR and SBP than CC,despite their largest sympathetic activation. PC have faster impulse on VE although be lowest than CC

The sympathetic nervous system plays a key role in the regulation of cardiovascular and ventilatory responses during exercise. The regulation of the heart and peripheral circulation by the autonomic nervous system is accomplished by control centers that receive input from mechanical and chemical receptors through the body. Therefore, the changes in sympathetic and parasympathetic activity allow for rapid responses. <p><p>Exercise is associated with increases of ventilation, heart rate and blood pressure. Ventilation increases adaptedly to increased oxygen uptake (VO2) and carbon dioxide output (VCO2) and eventually to limit metabolic acidosis occurring above the ventilatory threshold. Cardiac output increases to meet the contracting muscles’ requirement for flow. The increase in cardiac output occurs through increases in both heart rate and stroke volume and is regulated by feed-forward mechanisms: central command and exercise pressor reflex. <p><p>Skeletal muscle contraction elicits a reflex increase in sympathetic outflow which causes vasoconstriction contributing to the exercise induced rise in blood pressure. This reflex is triggered by stimulation of metabo- and chemoreceptors. Although the precise stimulus is not known, adrenergic receptor signaling is involved in the cardiovascular and respiratory alterations in response to exercise. <p><p>This thesis has been devoted to a better understanding of the functional aspects of sympathetic nervous system activation during dynamic and resistive exercise, with use of β blocker and β2 stimulant interventions The hypotheses were: 1) that β blocker interventions would decrease aerobic exercise capacity by a limitation of maximal cardiac output, but more so the ventilatory responses to exercise because of a decreased chemosensitivity, thereby decreasing dyspnea, and 2) β2 stimulant interventions would slightly increase aerobic exercise capacity by an increase in maximal cardiac output, but also the ventilatory responses because of an increased chemosensitivity, with possible decrease of the ventilatory reserve at exercise and increased dyspnea. Both interventions could affect maximal muscle strength through central effects.<p><p>Ventilatory responses to hyperoxic hypercapnia (central chemoreflex) and to isocapnic hypoxia (peripheral chemoreflex) were confronted to measurements of ventilatory equivalents for oxygen (O2) and carbon dioxide (CO2) during standard cardiopulmonary exercise test (CPET). Resting 5 measurements of muscle sympathetic nervous activity (MSNA) were obtained in different conditions with and without pharmacological interventions. Muscle metaboreflex and muscle stength measurements were also considered. Drugs with β blocker or β2 stimulant properties were administered in range of doses used in clinical practice for the teatment of cardiovascular or rerspiratory conditions. The results show that β blockade with bisoprolol slightly reduced maximal exercise capacity as assessed by a maximal oxygen uptake (VO2max) or maximal workload (Wmax), with a decreased maximal heart rate, without significant effect on ventilation (VE) or MSNA responses to hypercapnia, hyperoxia or to isometric muscle contraction or ischemia. Both VE/VO2 and VE/VCO2 slopes were decreased during CPET, which was attributable to β blockade-related hemodynamic changes. On the other hand, stimulation of β2 receptors with salbutamol did not affect exercise capacity as assessed by VO2max or Wmax in spite of increased peripheral chemosensitivity with increased VE/VCO2 slopes and early lactic acidosis. MSNA burst frequency, muscle metaboreflex and maximal isokinetic muscle strength were not affected by salbutamol. <p><p>Thus, aerobic exercise capacity in healthy subjects is sensitive to sympathetic nervous system modulation by β blocker or β2 stimulant interventions with drugs at doses prescribed in clinical practice. B blocker intervention has a slight limitation of aerobic exercise capacity and a hemodynamic decrease in ventilation, while β2 stimulant intervention has no change in exercise capacity with associated increased ventilatory responses because of increased chemosensitivity, partly related to early lactic acidosis. None of the studied phamacologic interventions affected MSNA or muscle strength measurements. <p><p>We hope that these results might be useful for the understanding of the effects of revalidation to exercise of patients treated with β blocker or β2 stimulant drugs, document the limited ergogenic properties and also side effects of the intake of these substances in healthy exercising subjects.<p> / Doctorat en Sciences de la motricité / info:eu-repo/semantics/nonPublished

Chemoreception plays an important role in predator-prey interactions and feeding dynamics. While the chemoreception of attractant or pleasant tasting compounds has been well studied, aversive chemoreceptive signaling has been difficult to investigate behaviorally in an ecological context because these interactions are species- and context- specific and deterrent compounds vary among prey. Using the coral reef system, this thesis explores on a molecular level the deterrent mechanism underlying detection by fish predators of an aversive compound, in order to gain a greater understanding of predator-prey interactions in this community. Like other organisms that are sessile or slow-moving, marine sponges have special mechanisms for defense from predation, commonly containing aversive-tasting compounds that defend these organisms from predation. To this end, we sought to identify and characterize a fish chemoreceptor that detects one or more of these compounds. We isolated a single cDNA clone encoding RAMP-like triterpene glycoside receptor (RL-TGR), a novel co-receptor involved in the signaling of triterpene glycosides. This co-receptor appears to be structurally and functionally related to receptor activity-modifying proteins (RAMPs), a family of co-receptors that physically associate with and modify the activity of G protein-coupled receptors (GPCRs). Expression in Xenopus oocytes showed that it responds to triterpene glycosides in a receptor-mediated manner and requires co-expression of a GPCR to enable signaling in oocytes; both of these receptors may be components of a larger signaling complex. A 40 bp portion of the gene is conserved across multiple fish species, but is not found in any other organism with a sequenced genome, suggesting that the expression of this receptor is limited to fish species. RL-TGR is the first identified gene encoding a co-receptor that responds to a chemical defense. This finding may lead the way for the identification of many other receptors that mediate chemical defense signaling in both marine and terrestrial environments, as this protein has the potential to represent the first of an entire family of co-receptors that respond to aversive compounds.

Motile microorganisms navigate through their environment using special molecular machinery in order to sense gradients of various signals: chemotaxis (reactions to chemical compounds) and phototaxis (to light) sensory cascades. Transmembrane receptors play a central role in these cascades as they receive input signals and transmit them inside the cell, where they modulate activity of the kinases CheA, which are tightly bound to their cytoplasmic domains. CheA further phosphorylates the response regulator protein CheY, which regulates the flagella. At the same time, CheA phosphorylates and, by means of this, activates another response regulator, CheB, which, along with the constantly active CheR protein, catalyzes two opposite reactions: methylation and demethylation of the specific glutamic acid residues located at the cytoplasmic domain of the receptors. The latter reactions establish the adaptation mechanism, which allows microbes to sense in a very broad range of the input signal intensities. Many functional, structural and dynamical aspects of the signal propagation through the prokaryotic receptors as well as a mechanism of the signal amplification remain still unclear. In the present thesis we have used various techniques of computational biophysics, chiefly molecular dynamics (MD) simulations, in order to approach these problems. In Chapter 3, we have carried out MD simulations of the isolated linker domain (HAMP) from the E. coli Tsr chemoreceptor. The MD simulations revealed highly dynamical nature of this domain, which allows for interconversion between several metastable states. These metastable states feature a number of structural and dynamical properties, which were previously reported for HAMP domains of various receptors obtained from different organisms. It allowed us to reconcile numerous experimental data and to hypothesize that different HAMP domains share similar mechanism of their action. In Chapter 4, we have performed MD simulations of the whole cytoplasmic domain of the Tsr chemoreceptor. The simulations revealed a mechanism for the inter-domain coupling between the HAMP domain and a part of the cytoplasmic domain adjacent to the HAMP, the adaptation subdomain, by means of the regulated unfolding of a short linker region termed the stutter. Also, we have found that the reversible methylation/demethylation of the cytoplasmic domain affects its flexibility and symmetry. Altogether, these findings suggest a mechanism of signal propagation at the level of an individual chemoreceptor dimer. In Chapter 5, we have built a model of the trimer-of-dimers of the archaeal phototaxis receptor complex (NpSRII:NpHtrII). Subsequent MD simulations revealed an important role of dynamics in signal transduction and, potentially, in the kinase activation. In Chapter 6, we have reconstructed a whole transmembrane lattice formed by the NpSRII:NpHtrII complexes. The concave shape of the obtained lattice naturally explains polar localization of the receptor arrays in prokaryotic cells. At the same time, additional MD simulations of an individual unit of this lattice (a dimer of the photosensor) revealed global motional modes in its transmembrane region, which presumably co-occur with its activation and can spread across the tightly packed transmembrane arrays allowing for the signal amplification.

No description available.

Molécules du goût et odeurs se fixent sur des récepteurs dits gustatifs et olfactifs, présents dans la bouche et le nez. Ils sont donc en contact avec le monde environnant. Toutefois, on les trouve également dans des organes isolés de l’extérieur, comme le pancréas ou le cerveau, où ils ne sont plus impliqués dans la détection du non-soi. Ils y régulent la glycémie ou l’activation du système immunitaire. Dans le cerveau, leurs rôles demeurent mystérieux. Mon travail a consisté à déterminer: 1) si, et où les récepteurs gustatifs et olfactifs sont présents dans le cerveau humain, 2) quand, où et pourquoi les récepteurs olfactifs sont présents dans le cerveau de souris et 3) si une maladie comme Alzheimer peut modifier leur expression. Mes résultats montrent qu’ils sont présents dans l’ensemble du cerveau humain et particulièrement dans le «cerveau émotionnel». De plus, le cerveau de souris «Alzheimer» surexprime des récepteurs olfactifs, notamment dans les neurones. Le cerveau est donc capable de goûter et sentir son monde intérieur. On peut imaginer que ces récepteurs jouent un rôle dans la détection de la maladie et, qui sait, qu’ils participent à la lutte contre ses effets néfastes. / Taste molecules and odours bind to so-called gustatory and olfactory receptors present in the mouth and nose. They are therefore in contact with the surrounding world. However, they are also found in organs isolated from the outside, such as the pancreas or brain, where they are no longer involved in the detection of non-self. They regulate blood sugar levels or the activation of the immune system. In the brain, their roles remain mysterious. My work consisted in determining: 1) if, and where, taste and smell receptors are present in the human brain, 2) when, where and why smell receptors are present in the mouse brain, and 3) whether a disease like Alzheimer's can change their expression. My results show that they are present in the entire human brain and particularly in the "emotional brain". In addition, the brains of "Alzheimer" mice overexpress olfactory receptors, particularly in neurons. The brain is therefore able to taste and feel its inner world. It is conceivable that these receptors play a role in detecting the disease and, who knows, in combating its harmful effects.

The aim of this thesis was to elucidate the role of carbonic anhydrase (CA) in the modulation of central pH/CO2-sensitive fictive breathing (measured using in vitro brainstem-spinal cord preparations) in leopard frogs (Rana pipiens) following exposure to chronic hypercapnia (CHC) and chronic hypoxia (CH). CHC caused an augmentation in fictive breathing compared to the controls (normoxic normocapnic). Addition of acetazolamide (ACTZ), a cell-permeant CA inhibitor, to the superfusate reduced fictive breathing in the controls and abolished the CHC-induced augmentation of fictive breathing. ACTZ had no effect on preparations taken from frogs exposed to CH. Addition of bovine CA to the superfusate did not alter fictive breathing in any group, suggesting that the effects of ACTZ were due to inhibition of intracellular CA. Taken together, these results indicate that CA is involved in central pH/CO2 chemoreception and the CHC-induced increase in fictive breathing in the leopard frog.

The aim of this thesis was to elucidate the role of carbonic anhydrase (CA) in the modulation of central pH/CO2-sensitive fictive breathing (measured using in vitro brainstem-spinal cord preparations) in leopard frogs (Rana pipiens) following exposure to chronic hypercapnia (CHC) and chronic hypoxia (CH). CHC caused an augmentation in fictive breathing compared to the controls (normoxic normocapnic). Addition of acetazolamide (ACTZ), a cell-permeant CA inhibitor, to the superfusate reduced fictive breathing in the controls and abolished the CHC-induced augmentation of fictive breathing. ACTZ had no effect on preparations taken from frogs exposed to CH. Addition of bovine CA to the superfusate did not alter fictive breathing in any group, suggesting that the effects of ACTZ were due to inhibition of intracellular CA. Taken together, these results indicate that CA is involved in central pH/CO2 chemoreception and the CHC-induced increase in fictive breathing in the leopard frog.

Submitted by Daniele Amaral (daniee_ni@hotmail.com) on 2017-10-16T18:31:31Z No. of bitstreams: 1 TeseWIF.pdf: 2113715 bytes, checksum: 6432f11b933c0c20bd1a2e05f8ba6252 (MD5) / Approved for entry into archive by Ronildo Prado (producaointelectual.bco@ufscar.br) on 2017-10-31T11:05:59Z (GMT) No. of bitstreams: 1 TeseWIF.pdf: 2113715 bytes, checksum: 6432f11b933c0c20bd1a2e05f8ba6252 (MD5) / Approved for entry into archive by Ronildo Prado (producaointelectual.bco@ufscar.br) on 2017-10-31T11:06:09Z (GMT) No. of bitstreams: 1 TeseWIF.pdf: 2113715 bytes, checksum: 6432f11b933c0c20bd1a2e05f8ba6252 (MD5) / Made available in DSpace on 2017-10-31T11:11:20Z (GMT). No. of bitstreams: 1 TeseWIF.pdf: 2113715 bytes, checksum: 6432f11b933c0c20bd1a2e05f8ba6252 (MD5) Previous issue date: 2017-08-11 / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / The nucleus of the solitary tract (NTS) is the primary site of visceral afferents, such as baroreceptors and arterial chemoreceptors. Recent data from our laboratory have shown that the microinjection of acetylcholine (ACh) into the commissural moiety of the NTS (cNTS) of decorticated arterially-perfused in situ preparations of male juvenile rats did not change the sympathetic nerve activity (SNA), but increased the phrenic nerve activity (PNA). Furthermore, we demonstrated that the ACh-induced responses in cNTS involve the activation of both nicotinic and muscarinic receptors. However, nicotinic receptors seem to play a more relevant role in the control of breathing, especially considering that such receptor antagonism promotes a decrease in the KCN- activated peripheral chemoreflex tachypneic response. However, the effects of specific nicotinic and muscarinic agonists in the cNTS on respiratory and sympathetic responses have not been studied yet. Once established the involvement of nicotinic receptors in the cNTS on peripheral chemoreflex ventilatory responses activated by cytotoxic hypoxia (KCN), we also evaluated the involvement of the cholinergic system in the cNTS on respiratory and sympathetic responses induced by hypercapnia or 24 h sustained hypoxia. Therefore, this project proposed to study the effects of selective activation of distinct cholinergic receptors in the cNTS on respiratory and sympathetic activities and the role of the cholinergic system in cNTS on sympathetic and respiratory activities reflex changes in response to hypercapnia or sustained hypoxia. We observed that the injection of both nicotinic and muscarinic agonists in the cNTS induces an increase in SNA and changes in the respiratory modulation pattern. The nicotinic agonist induces a decrease in respiratory frequency, as well as the blockade of the enzyme acetylcholinesterase. It was also observed that the cholinergic agonists promote an increase in the amplitude and duration of the pre-inspiratory (pre-I) period of the hypoglossal nerve and also increased the amplitude of the vagus nerve. When it comes on the protocols involving hypoxia, we observed that the cholinergic antagonists injected into the cNTS of rats previously exposed to hypoxia promoted a decrease in sympathetic activity, increased respiratory frequency, decreased hypoglossal nerve amplitude, and decreased post-inspiratory peak amplitude of the vagus nerve, but only the muscarinic antagonist decreased phrenic nerve amplitude and hypoxia-induced hypoglossal nerve pre-I increase. Regarding to the experiments with hypercapnia, we verified that the nicotinic antagonist in the cNTS inhibited the hypercapnia-induced increase in pre-I of the hypoglossal nerve. In addition, the nicotinic antagonist injected into the cNTS also potentiated the recruitment of late-E activity from the abdominal nerve. Taken together, the responses observed with the cholinergic agonists and injected into the cNTS, as well as the antagonists upon hypoxia, suggest the involvement of cholinergic pathways in the cNTS in the modulation of sympathetic and respiratory responses to sustained hypoxia. On the other hand, it seems that only nicotinic receptors in the cNTS are involved in hypercapnia-induced increase in pre-inspiratory activity and active expiration. / O núcleo do trato solitário (NTS) é o sítio primário de aferências viscerais, como barorreceptores e quimiorreceptores arteriais. Estudos recentes do nosso laboratório demonstraram que, em preparações in situ, decorticadas e perfundidas intra-arterialmente, a microinjeção de acetilcolina (ACh) na porção comissural do NTS (NTSc) não alterou a atividade simpática (SNA), mas promoveu aumento da atividade do nervo frênico (PNA). Além disso, evidenciamos que as respostas induzidas pela ACh no NTSc envolvem a ativação dos receptores nicotínicos e muscarínicos. Contudo, os receptores nicotínicos parecem desempenhar um papel mais relevante no controle da respiração, principalmente considerando que o antagonismo de tais receptores promove uma redução da resposta taquipneica do quimiorreflexo periférico ativado pelo KCN. Entretanto, os efeitos de agonistas específicos nicotínicos e muscarínicos, bem como a inibição da inibição da degradação de ACh no NTSc sobre as respostas respiratórias e sobre a atividade simpática ainda não foram estudados. Sabendo-se da participação dos receptores nicotínicos do NTSc sobre as respostas ventilatórias dos quimiorreceptores periféricos ativados por hipóxia citotóxica (KCN), avaliamos também a participação do sistema colinérgico do NTSc sobre as respostas simpática e respiratória induzidas por hipercapnia ou hipóxia sustentada por 24 h. Portanto, este projeto se propôs a estudar o efeito da ativação seletiva de diferentes receptores colinérgicos no NTSc sobre as atividades simpática e respiratória e o papel do sistema colinérgico no NTSc sobre as alterações reflexas nas atividades simpática e respiratória em resposta à hipercapnia ou hipóxia sustentada por 24 h. Observamos que a injeção de agonistas tanto nicotínico quanto muscarínico no NTSc promovem aumento da SNA e modifica o seu padrão de modulação respiratória. O agonista nicotínico induz uma diminuição da frequência respiratória, assim como o bloqueio da enzima acetilcolinesterase. Também foi observado que os agonistas colinérgicos promovem um aumento na amplitude e duração do período préinspiratório (pre-I) do nervo hipoglosso e também aumento na amplitude do nervo vago. Com relação aos protocolos envolvendo hipóxia, observamos os antagonistas colinérgicos injetados no NTSc de ratos previamente expostos à hipóxia, promoveu diminuição da atividade simpática, aumento da frequência respiratória, diminuição da amplitude do nervo hipoglosso e diminuição da amplitude do pico pós-inspiratório do nervo vago, mas somente o antagonista muscarínico diminuiu a amplitude do nervo frênico e o aumento do pre-I do nervo hipoglosso induzido pela hipóxia. Com relação aos experimentos com hipercapnia, verificamos que o antagonista nicotínico no NTSc inibiu o aumento do pre-I do nervo hipoglosso induzido pela hipercapnia. Além disso, o antagonista nicotínico injetado no NTSc também potencializou o recrutamento de atividade late-E do nervo abdominal. Tomados em conjunto, as respostas observadas com os agonistas colinérgicos injetados no NTSc, bem como com os antagonistas mediante a hipóxia, sugerem a participação de vias colinérgica do NTSc na modulação das respostas simpática e respiratória à hipóxia sustentada. Por outro lado, apenas os receptores nicotínicos do NTSc parecem estar envolvidos com o aumento da atividade pré-inspiratória e da expiração ativa induzidos por hipercapnia. / FAPESP: 2013/22526-4