Quimioreceptores centrais (QC) e periféricos (QP) são células especializadas em detectar alterações de CO2, O2 e H+, e promover ajustes na ventilação e pressão arterial via sistema nervoso central. Avaliamos aqui a ação da sinalização purinérgica em áreas que apresentam essa propriedade (RTN, C1, NTScom e RPa) durante as respostas cardiorrespiratórias promovidas pela ativação dos quimiorreceptores, e a possível participação de astrócitos. Encontramos evidências que receptores P2 modulam a resposta de QC no RTN, enquanto que receptores P2Y1 e receptores glutamatérgicos, modulam a resposta de QP no C1, e que a sinalização purinérgica na região do NTScom ou na região RPa não contribui para resposta de QC. A manipulação farmacológica de astrócitos do RTN com fluorocitrato, mas não da região do NTScom e RPa, produz alterações respiratórias via receptores P2. Nossos achados evidenciam a importância e contribuem para descriminação dos mecanismos de ação da sinalização purinérgica na região bulbo ventrolateral rostral durante a ativação QC e QP. / Central (CC) and peripherals (PC) chemoreceptors are specialized cells to detect changes in CO2, O2 and H+, and produce adjustments in ventilation and blood pressure via the central nervous system. Here we evaluate the action of purinergic signaling in areas with this property (RTN, C1, commNTS, RPA) during the cardiorespiratory responses elicited by activation of chemoreceptors, and a possible role of astrocytes. We found evidence that P2 receptors modulate CC responses in RTN, while P2Y1 and glutamate receptors modulate PC responses in C1, and that the purinergic signaling in the RPa and commNTS region does not contribute to CC responses. The pharmacological manipulation of the RTN astrocytes, but not commNTS or RPa, with fluorocitrate produces respiratory changes via P2 receptors. Our findings show the importance and contribute to discrimination of the mechanisms of purinergic signaling in the rostral ventrolateral medulla during CC and PC activation.

The studies presented in this thesis were designed to investigate the effects of reduced sea water pH on the behaviour of intertidal decapod crustaceans, both within the context of the variations occurring naturally in the pH of rock pool habitats, and in relation to predicted changes to ocean pH resulting from ocean acidification and potential carbon dioxide (CO2) leaks from carbon capture storage (CCS) sites. Recent studies on marine fish have shown behavioural disruptions as a result of increased CO2 concentrations in sea water and reduced pH, but the effects on crustaceans are as yet unknown. The first two studies investigated the effects of reduced pH upon the olfactory behaviour of the prawn Palaemon elegans and the hermit crab Pagurus bernhardus, focussing on their responses to food odours. Short-term (five day) exposures to highly reduced pH (pHNBS = 6.60, 6.80) revealed disruptions to the chemo-sensory behaviour of both species with a reduction in their ‘sniffing’ response, and the inability of P. bernhardus to locate the chemical cue. This was also accompanied by elevated haemolymph chloride ions. in In a further study P. bernhardus was subjected to a longer exposure (60 days) and to a range of pH levels (pHNBS = 8.00, 7.90, 7.70, 7.35 and 6.80) in order to detect a threshold for the behavioural disruptions observed, and to determine if there would be any sign of acclimation over a longer period. A clear gradient in the disruptions to the chemo-sensory responses and survival rates of the hermit crabs, and disruption to a physiological marker (elevated haemolymph calcium ions), was found. Possible thresholds for disruption were also identified at levels that match predictions for ocean acidification and leaks from proposed CO2 CCS sites. Some of the crabs in the lower pH treatments exhibited a recovery in their responses by day 60, possibly indicating an acclimation effect. The presence of disruption to haemolymph ion concentrations in both the short and longer term hermit crab studies suggest a mechanism for behavioural disruption. In a final study the effects of reduced sea water pH on a more complex behaviour, involving decision making, was investigated. Reduced sea water pH was shown to disrupt the shell assessment and selection behaviour of P. bernhardus affecting its decision making processes, although not all crabs were affected in the same way. The work presented here therefore demonstrates that reduced sea water pH could have disruptive effects upon both information gathering, via chemo-sensory processes, and decision making in intertidal crustaceans. The mechanism responsible is unlikely to be due to changes in the odour molecule, or physical damage to receptor organs. Rather the observed disruptions could be due (a) to ionic changes, causing metabolic depression or interference with neurotransmitter function, or (b) to disruption to chemoreception per se. Such disturbances to key behavioural processes have implications for inter and intraspecific species interactions and population dynamics in the marine environment. Changes in pH are already experienced by intertidal animals for short periods when rock pools are emersed, but future anthropogencially-induced reductions in sea water pH are likely to cause more sustained and widespread disruptions with, as yet, unpredictable consequences. The differential responses observed between individuals in these studies may warrant further investigation as such differences may provide the basis for selection and adaptation to projected changes in ocean pH.

Au cours d'une histoire évolutive singulière, les systèmes sensoriels des cétacés se sont adaptés à la vie en milieu aquatique. Aujourd’hui, alors que la littérature scientifique regorge de travaux sur leurs capacités acoustiques exceptionnelles, l’utilisation des sens chimiques par ces mammifères marins demeure encore largement méconnue. En effet, malgré quelques rares observations suggérant qu’ils pourraient détecter les composés sécrétés par leurs proies et leurs congénères, les arguments anatomiques et génétiques sont plutôt en faveur d’une régression voire d’une disparition de leurs capacités chémoréceptrices. Les cétacés auraient-ils perdu l’usage de ce canal sensoriel pourtant fondamental pour l’alimentation, la navigation et la reproduction chez les autres grands prédateurs marins ? L’objet principal de ma thèse était donc de déterminer si ces animaux sont capables de percevoir et d’utiliser les indices chimiques présents dans leur environnement en me basant principalement sur une approche comportementale. J’ai ainsi étudié les réactions des cétacés à dents (Odontocètes) et à fanons (Mysticètes) face à différents stimuli solubles ou volatiles, liés directement ou indirectement à leur alimentation. Après avoir mis en évidence, dans des conditions contrôlées, que le grand dauphin discrimine des extraits solubles de ses proies, j’ai développé un protocole permettant de mesurer la réponse des animaux (déplacements et comportements de surface) à des stimuli chimiques, dans leur milieu naturel. Des expériences en Méditerranée occidentale ont révélé que les grands dauphins et les globicéphales (Odontocètes) réagissent au sulfure de diméthyle (DMS), une molécule volatile émise dans les zones de forte productivité primaire. J’ai ensuite mesuré, chez la baleine à bosse (mysticète), la réaction au DMS et à des extraits de proies (krill) dans ses zones de reproduction (Océan Indien) et de nourrissage (Atlantique Nord et Antarctique). Les animaux exposés ont réagi par une augmentation de la fréquence respiratoire et, dans le cas de l’extrait de krill uniquement, par une attraction vers la source du signal (chemotaxis). Nos résultats comportementaux ayant mis en évidence une chémoréception fonctionnelle chez les cétacés, j’ai tenté dans un second temps d’identifier ses bases anatomiques et chimiques. J’ai ainsi initié l’exploration des muqueuses orales et nasales par des techniques d’immunohistochimie, ainsi que l’analyse chimique de leurs sécrétions (urines et fèces) à la recherche de potentielles phéromones. Cette approche innovante et multidisciplinaire a permis de dévoiler l’implication des signaux chimiques dans l’écologie des cétacés. Au-delà de leur aspect fondamental, ces résultats pourraient trouver des applications concrètes pour la gestion et la conservation de ces espèces emblématiques et menacées. / During the course of a unique evolutionary process, the sensory systems of cetaceans (whales, dolphins and porpoises) have secondarily adapted to life in an aquatic environment. While the extraordinary acoustic capacities of these animals have been widely studied, surprisingly little is known about their chemosensory abilities. The results of some sparse behavioural studies suggest that cetaceans can detect molecules secreted by, for example, a prey or congener. In contrast, anatomical and genetic investigations have concluded that cetacean chemosenses are greatly reduced or even absent. This poses the question; have cetaceans truly lost the use of the sensory channels that play a major role in the feeding, orientation and reproduction behaviours of other marine predators? The main objective of this thesis was, therefore, to establish whether these animals are capable of perceiving biologically-relevant chemical cues and exhibiting a behavioural response. For this project, I studied the reactions of both toothed (Odontoceti) and baleen (Mysticeti) whales to various soluble or volatile food-related stimuli. I first worked with captive bottlenose dolphins (Tursiops truncatus) and showed they could discriminate soluble extracts of their piscine prey. I then developed a novel protocol to measure the responses (surface behaviour and movements) of cetaceans to chemical cues in their natural habitat. Using this protocol, experiments performed in the western Mediterranean Sea revealed that both bottlenose dolphins and pilot whales (Globicephala melas) react to dimethyl sulfate (DMS), a volatile molecule found in areas of high primary productivity. Using the same protocol, I subsequently measured the reactions of a mysticete species, the humpback whale (Megaptera novaeangliae) to DMS and to krill extract (their natural prey) in three different locations: in their reproduction zone in the Indian Ocean, and in their feeding grounds in the North Atlantic and Antarctic Ocean. I found that the humpback whales reacted with an increased respiratory rate to both stimuli and that the krill extract also attracted them (chemotaxis). As the results of the behavioural studies suggested the presence of functional chemical senses, I initiated two further studies; (1) a neuroanatomical and immunohistochemical exploration of tissues in the oral and nasal pathways, in order to identify potential chemosensory receptor cells in four species of odontocete, and (2) a chemical analysis of odontocete secretions (urine or feces) in search for potential semiochemicals. Overall, this innovative and multidisciplinary research project revealed that chemical signals are potentially very important in the behavioral ecology of cetaceans. Moreover, these results contribute not only to our fundamental knowledge of the sensory biology of these animals, but they could also have important practical implementations in the conservation of these iconic and threatened species.

Made available in DSpace on 2014-06-11T19:22:23Z (GMT). No. of bitstreams: 0 Previous issue date: 2008-07-25Bitstream added on 2014-06-13T18:08:10Z : No. of bitstreams: 1 clavijoayala_ja_me_jabo_prot.pdf: 2110673 bytes, checksum: 81a603896afe42aa218b5fa3b86d612c (MD5) / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O pacu Piaractus mesopotamicus é uma das espécies de maior relevância na piscicultura de águas quentes no Brasil, destacando-se pela qualidade de sua carne e desempenho em sistemas de cultivo. Apesar do avanço no estudo da biologia, anatomia e morfologia da espécie, os aspectos relacionados com o desenvolvimento inicial do sistema sensorial são praticamente desconhecidos. Este estudo teve por objetivo descrever o desenvolvimento inicial de pacu Piaractus mesopotamicus, com ênfase na ontogenia das estruturas sensoriais. Para isto, amostras seriadas de embriões e larvas em desenvolvimento foram coletadas desde as 9 horas pós fertilização (hpf) até os 25 dias pós eclosão (dpe), e destinadas para análises morfológicas, histológicas e de microscopia eletrônica de varredura. O desenvolvimento embrionário da espécie é rápido, e cerca das 19 hpf(a 26,S ± O,SOC), as larvas eclodem num estado altricial (3,19 ± 0,04 mm comprimento notocordal- CN; 0,477 ± 0,061 mm3 volume do saco+ vitelino¬VSV). Na eclosão, o desenvolvimento do sistema sensorial é limitado: no epitélio olfatório observam-se neurônios sensoriais ciliados e neurônios sensoriais com microvilosidades; a presença de mecanorreceptores é restrita ao neuromasto ótico imaturo; o desenvolvimento do ouvido interno é incipiente, observa-se a presença de dois otólitos em cada cápsula ótica, mas a larva ainda não apresenta estabilidade na coluna de água nem coordenação nos seus movimentos; e os olhos apresentam-se com pouco ou nenhum pigmento, lente diferenciada e separada da córnea, e retina não estratificada No entanto, o desenvolvimento pós-embrionário do sistema sensorial é acelerado: a partir de 1 dpe o epitélio olfativo se encontra coberto por uma densa camada de cilios não sensoriais que se distribuem. tanto na vesícula... / Pacu, Piaractus mesopotamicus, is one of the species of major importance for warmwater fish farming in Brazil due to its meat quality and performance in culture systems. Despite the advances in the study of biology, anatomy and morphology of the species, the aspects related to the early development of the sensorial system are unknown. The aim of this study was to describe the initial development of pacu Piaractus mesopotamicus, with emphasis on the ontogeny of the sensorial structures. Embryos and larvae were serially sampled during development, from the 9 hours after fertilization (haf) to the 25 days after hatching (dah), and subjected to the morphology, histology and scanning electron microscopy analysis. The embryonic development of the species is fast: about 19 haf (at 26,5 ± 0,5°C), larvae hatch in an altricial state (3.19 ± 0.04 mm notochordal length - NL; 0.477 ± 0.061 mm3 yolk-sac volume - YSV). At hatching, the sensorial system development is limited: in the olfactory epithelium, there are ciliated sensorial neurons and microvillous sensorial neurons; the mechanoreceptors are restricted to the immature otic neuromast; the development of the inner ear is incipient, with two otoliths in each otic capsule, but larvae do not show neither stability in the water column or coordination of their movements; the eyes have no or few pigment, with lens differentiated and separated from the cornea, and non-stratified retina. However, the post-embryonic development of the sensorial system is accelerated: from the 1 st dah the olfactory epithelium is covered by a dense layer of non sensorial cilia, which are distributed in the olfactory vesicle and in the epithelium region around it; at 2,5-3 dah, the eyes are totally pigmented; from the 4th dah it is possible to identify solitary chemosensorial cells in the post-otical region;...(Complete abstract click electronic access below)

Made available in DSpace on 2016-06-02T19:23:01Z (GMT). No. of bitstreams: 1 6358.pdf: 1461364 bytes, checksum: 88df144bc77432e6c6026f8cd985fa7a (MD5) Previous issue date: 2014-08-29 / Universidade Federal de Minas Gerais / The hypocretin or orexin A plays an important role in the modulation of respiratory control in mammals, but there are no data available for the role of Orexins in the peripheral and central chemoreception of non-mammalian vertebrates. Thus, the present study was designed to evaluate the location of orexinergic neurons in toads (Rhinella schneideri). In addition, we investigate if the orexinergic system of this species is important to hypoxic (5% O2 and N2 for balance) and hypercarbic (5% CO2, 21% O2 and N2 for balance) drive to breath. We assessed the role of the orexinergic system on respiratory responses by using intracerebroventricular injection of SB-334867 (orexin A receptor antagonist) during the light and the dark phase. Our results demonstrated that orexinergic neurons of Rhinella schneideri are located in the suprachiasmatic nucleus of diencephalon. Additionally, the injection of the orexin antagonist attenuated the ventilatory response to hypercarbia during the dark phase by acting on tidal volume and breathing frequency, while in the light phase, there was an attenuation in the ventilatory response to hypoxia by acting just in the tidal volume. We conclude that central orexin A contributes to hypercarbic and to hypoxic chemoreflex in toads Rhinella schneideri. / A orexina ou hipocretina exerce uma importante modulação no controle respiratório em mamíferos, mas nenhum estudo verificou a participação das orexinas na quimiorrecepção central e periférica em vertebrados não-mamíferos. Em vista disso, o objetivo do presente estudo foi avaliar a imunorreatividade para orexina para localização dos neurônios orexinérgicos no encéfalo de sapos (Rhinella schneideri) e investigar se o sistema orexinérgico desta espécie participa nas respostas respiratórias à hipóxia (5% O2 e N2 balanço) e à hipercarbia (5% CO2, 21%O2 e N2 balanço). A participação da neurotransmissão orexinérgica na regulação respiratória em sapos (Rhinella schneideri) foi avaliada por meio de microinjeções intracerebroventriculares (i.c.v.) de SB-334867 (antagonista de receptores OX1R) em duas doses (5 mM e 10mM) no ventrículo lateral durante as fases clara e escura desses animais. Nossos resultados mostram que os neurônios orixinérgicos de sapos Rhinella schneideri estão localizados no núcleo supraquiasmático no diencéfalo. Adicionalmente, a injeção do antagonista orexinérgico atenua a resposta ventilatória à hipercarbia na fase escura, devido tanto à diminuição do volume corrente quanto da frequência respiratória, enquanto, na fase clara, atenua a resposta ventilatória à hipóxia devido a uma redução no volume corrente. Concluímos, portanto, que a Orexina A participa na modulação dos quimiorreflexos central e periférico em sapos Rhinella schneideri.

Submitted by Izabel Franco (izabel-franco@ufscar.br) on 2016-09-15T13:00:14Z No. of bitstreams: 1 DissLGAP.pdf: 2357885 bytes, checksum: dd8d8d69a2f2f83808b3f1561307aefd (MD5) / Approved for entry into archive by Marina Freitas (marinapf@ufscar.br) on 2016-09-16T19:24:00Z (GMT) No. of bitstreams: 1 DissLGAP.pdf: 2357885 bytes, checksum: dd8d8d69a2f2f83808b3f1561307aefd (MD5) / Approved for entry into archive by Marina Freitas (marinapf@ufscar.br) on 2016-09-16T19:24:13Z (GMT) No. of bitstreams: 1 DissLGAP.pdf: 2357885 bytes, checksum: dd8d8d69a2f2f83808b3f1561307aefd (MD5) / Made available in DSpace on 2016-09-16T19:24:24Z (GMT). No. of bitstreams: 1 DissLGAP.pdf: 2357885 bytes, checksum: dd8d8d69a2f2f83808b3f1561307aefd (MD5) Previous issue date: 2015-10-02 / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / It is well know that the respiratory network, undergoes significant development in the postnatal period. Among various processes, the maturing of the catecholaminergic (CA) system shows to be an important factor in the control and modulation of respiratory rhythmogenesis. Studies have also shown that these neurons are widely distributed in the Central Nervous System (CNS), including the A1/C1, A2/C2, C3, A5, A6 and A7 regions, having numerous projections to many regions of the brain. However, the development of respiratory network as well as its effect on the control of ventilation, is not yet fully understood. Thus, understanding the participation of CA neurons in the respiratory control during postnatal development is of most importance for a better understanding of some clinical disorders including Rett Syndrome, Sudden Infant Death Syndrome (SIDS) and Central Congenital Hypoventilation Syndrome (CCHS). Therefore, this study aimed to investigate the involvement of CA neurons in the brainstem on respiratory control in normoxic normocapnic, hypercapnic and hypoxic conditions during the postnatal period of male and female neonatal rats, through chemical injury with conjugated saporin anti-dopamine beta-hydroxylase (DBH-SAP). Thus, DBH-SAP (42 ng/100 nL – 1L), saporin (SAP – 1L) or phosphate buffered solution vehicle (PBS, 0.01M, pH 7.4 – 1L) were injected into the 4th ventricle in male and female neonates Wistar rats P0-1. Pulmonary ventilation ( EV ) was recorded in unanesthetized neonates (P7-8) by pressure plethysmography during normocapnia, hypercapnia (7% CO2) and hypoxia (10% O2) at 10 and 20 min after the start of exposure. Our data demonstrate that lesion of brainstem CA neurons increased ventilation in males and females newborn under room air conditions. In addition, the ventilatory response to hypercapnia was significantly reduced in male (57%) and female (55%) lesioned neonatal rats (Male – SAP group: 212.8 ± 7.0; PBS group: 203.9 ± 10.3; lesioned group: 151.1 ± 7.4; P < 0,001; Female – SAP group: 218.2 ± 10.4; PBS group: 200.0 ± 6.4; lesioned group: 154.0 ± 9.6; P < 0,001; all values relative to % of baseline). Also, a similar reduction was observed in the hypoxic condition (Male – SAP group: 185.2 ± 15.3; PBS group: 167.4 ± 5.0; lesioned group: 110.8 ± 9.2; P < 0,001; Female – SAP group: 197.3 ± 11.8; PBS group: 179.5 ± 13.7; lesioned: 129.4 ± 5.9; P < 0,001; all values relative to % of baseline). Additionally, the values for metabolic rate of control and lesioned groups, both males and females, did not differ significantly, whether in normoxic normocapnic, hypercapnic or hypoxic conditions. These results suggest that brainstem CA neurons exert a tonic inhibitory role in neonatal ventilation and promote an important excitatory modulation in CO2 and O2 chemosensitivity in unanesthetized males and females neonatal rats (P7-8). / Sabe-se que o sistema respiratório, bem como suas vias de controle, sofrem significativo desenvolvimento no período pós-natal. Dentre vários processos, o amadurecimento do sistema catecolaminérgico (CA) mostra-se como um importante fator no controle e modulação da ritmogênese respiratória. Estudos demonstram que esses neurônios estão amplamente distribuídos pelo Sistema Nervo Central (SNC), incluindo as regiões A1/C1, A2/C2, C3, A5, A6 e A7, e que apresentam inúmeras projeções para várias regiões do encéfalo. No entanto, a participação dos neurônios CA no controle respiratório durante o desenvolvimento pós-natal não está bem esclarecido, e esse entendimento é de extrema importância para uma melhor compreensão de alguns problemas clínicos que inclui a Síndrome de Rett, Síndrome da Morte Súbita Infantil (SIDS) e a Síndrome da Hipoventilação Central Congênita (CCHS). Sendo assim, o presente estudo teve por objetivo investigar a participação dos neurônios CA do tronco encefálico no controle respiratório em situações normóxica normocápnicas, hipercápnicas e hipóxicas durante o período pós-natal de ratas e ratos (P7-8), por meio de lesão química com saporina conjugada com anti-dopamina beta-hidroxilase (DBHSAP). Assim, DBH-SAP (42 ng/100 nL – 1L), Saporina (SAP – 1 L) ou veículo solução fosfato tamponado (PBS 0,01 M, pH 7,4 – 1 L) foram injetados no 4° ventrículo de ratas e ratos neonatos Wistar P0-1. A ventilação pulmonar ( EV ) foi registrada em neonatos não anestesiados (P7-8) por pletismografia de pressão, durante normóxia normocápnica, hipercapnia (7% CO2) e hipóxia (10% O2) aos 10 e 20 min após o início da exposição. Nossos dados demonstram que a lesão dos neurônios catecolaminérgicos do tronco encefálico promove um aumento da ventilação em neonatos machos e fêmeas durante a normóxia normocápnica. A resposta ventilatória à hipercapnia foi significativamente reduzida em ratos neonatos lesados (57%) e ratas (55%) (Machos – grupo SAP: 212,8 ± 7,0; grupo PBS: 203,9 ± 10,3; grupo lesado: 151,1 ± 7,4; P < 0,001; Fêmeas – grupo SAP: 218,2 ± 10,4; grupo PBS: 200,0 ± 6,4; grupo lesado: 154,0 ± 9,6; P < 0,001; todos os valores relativos à % do basal). Uma redução similar foi observada na resposta ventilatória à hipóxia (Machos – grupo SAP: 185,2 ± 15,3; grupo PBS: 167,4 ± 5,0; grupo lesado: 110,8 ± 9,2; P < 0,001; Fêmeas – grupo SAP: 197,3 ± 11,8; grupo PBS: 179,5 ± 13,7; grupo lesado: 129,4 ± 5,9; P < 0,001; todos os valores relativos à % do basal). Adicionalmente, os valores referentes às taxas metabólicas de neonatos machos e fêmeas lesados e controles não diferiram significativamente, seja em condição de normóxia normocápnica, hipercapnia ou hipóxia. Esses resultados sugerem que os neurônios catecolaminérgicos localizados no tronco encefálico exercem um papel inibitório tônico sobre ventilação em neonatos P7-8 e apresentam uma importante modulação excitatória na resposta ventilatória ao CO2 e hipóxia em ratas e ratos neonatos (P7-8) não anestesiados.

Orientadora: Maria Célia Portella / Banca: Irene Bastos Franceschini Vicentini / Banca: Marcos Antonio Cestarolli / Resumo: O pacu Piaractus mesopotamicus é uma das espécies de maior relevância na piscicultura de águas quentes no Brasil, destacando-se pela qualidade de sua carne e desempenho em sistemas de cultivo. Apesar do avanço no estudo da biologia, anatomia e morfologia da espécie, os aspectos relacionados com o desenvolvimento inicial do sistema sensorial são praticamente desconhecidos. Este estudo teve por objetivo descrever o desenvolvimento inicial de pacu Piaractus mesopotamicus, com ênfase na ontogenia das estruturas sensoriais. Para isto, amostras seriadas de embriões e larvas em desenvolvimento foram coletadas desde as 9 horas pós fertilização (hpf) até os 25 dias pós eclosão (dpe), e destinadas para análises morfológicas, histológicas e de microscopia eletrônica de varredura. O desenvolvimento embrionário da espécie é rápido, e cerca das 19 hpf(a 26,S ± O,SOC), as larvas eclodem num estado altricial (3,19 ± 0,04 mm comprimento notocordal- CN; 0,477 ± 0,061 mm3 volume do saco+ vitelino¬VSV). Na eclosão, o desenvolvimento do sistema sensorial é limitado: no epitélio olfatório observam-se neurônios sensoriais ciliados e neurônios sensoriais com microvilosidades; a presença de mecanorreceptores é restrita ao neuromasto ótico imaturo; o desenvolvimento do ouvido interno é incipiente, observa-se a presença de dois otólitos em cada cápsula ótica, mas a larva ainda não apresenta estabilidade na coluna de água nem coordenação nos seus movimentos; e os olhos apresentam-se com pouco ou nenhum pigmento, lente diferenciada e separada da córnea, e retina não estratificada No entanto, o desenvolvimento pós-embrionário do sistema sensorial é acelerado: a partir de 1 dpe o epitélio olfativo se encontra coberto por uma densa camada de cilios não sensoriais que se distribuem. tanto na vesícula...(Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Pacu, Piaractus mesopotamicus, is one of the species of major importance for warmwater fish farming in Brazil due to its meat quality and performance in culture systems. Despite the advances in the study of biology, anatomy and morphology of the species, the aspects related to the early development of the sensorial system are unknown. The aim of this study was to describe the initial development of pacu Piaractus mesopotamicus, with emphasis on the ontogeny of the sensorial structures. Embryos and larvae were serially sampled during development, from the 9 hours after fertilization (haf) to the 25 days after hatching (dah), and subjected to the morphology, histology and scanning electron microscopy analysis. The embryonic development of the species is fast: about 19 haf (at 26,5 ± 0,5°C), larvae hatch in an altricial state (3.19 ± 0.04 mm notochordal length - NL; 0.477 ± 0.061 mm3 yolk-sac volume - YSV). At hatching, the sensorial system development is limited: in the olfactory epithelium, there are ciliated sensorial neurons and microvillous sensorial neurons; the mechanoreceptors are restricted to the immature otic neuromast; the development of the inner ear is incipient, with two otoliths in each otic capsule, but larvae do not show neither stability in the water column or coordination of their movements; the eyes have no or few pigment, with lens differentiated and separated from the cornea, and non-stratified retina. However, the post-embryonic development of the sensorial system is accelerated: from the 1 st dah the olfactory epithelium is covered by a dense layer of non sensorial cilia, which are distributed in the olfactory vesicle and in the epithelium region around it; at 2,5-3 dah, the eyes are totally pigmented; from the 4th dah it is possible to identify solitary chemosensorial cells in the post-otical region;...(Complete abstract click electronic access below) / Mestre

Endogenously produced carbon monoxide (CO) is an important gaseous signalling molecule which regulates a variety of cardiorespiratory functions. CO is produced in cells by the heme oxygenase (HO) family of proteins by the breakdown of heme into equimolar amounts of CO, bilirubin and Fe2+. My thesis focuses on the hypoxia- and hyperoxia-inducible HO-1/CO system exclusively and aims to provide the first evidence that the HO-1/CO system is involved in cardiorespiratory control in the zebrafish (Danio rerio) and goldfish (Carassius auratus). Overall, I hypothesise that the HO-1/CO system acts as a negative regulator of cardiorespiratory function in fish. Using immunohistochemistry, I was able to characterise the distribution of HO-1 and thus reveal the potential for endogenous CO production (from heme breakdown) in branchial and skin neuroepithelial cells (NECs; putative O2 chemoreceptors) and associated innervation as well as the heart of the developing zebrafish larva. The presence of HO-1 in these structures suggests the likelihood of specific and localized production of CO in fish. To assess the functional significance of the HO-1/CO system in control of cardiorespiratory function, I used pharmacological and gene knock down approaches to diminish HO-1 activity, and presumably endogenous CO production, in adult and larval fish, respectively. The results from these experiments provided evidence that 1) CO has an inhibitory influence on ventilation in goldfish and zebrafish but that its function is temperature- and species-dependent and 2) showed that the HO-1/CO system tonically inhibits cardiac activity in larval zebrafish.

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.

Submitted by Bruna Rodrigues (bruna92rodrigues@yahoo.com.br) on 2016-09-21T12:17:25Z No. of bitstreams: 1 DissMCV.pdf: 1396676 bytes, checksum: d3fa0ce7f0f0504dfe6926528b1e7366 (MD5) / Approved for entry into archive by Marina Freitas (marinapf@ufscar.br) on 2016-09-21T18:23:20Z (GMT) No. of bitstreams: 1 DissMCV.pdf: 1396676 bytes, checksum: d3fa0ce7f0f0504dfe6926528b1e7366 (MD5) / Approved for entry into archive by Marina Freitas (marinapf@ufscar.br) on 2016-09-21T18:23:27Z (GMT) No. of bitstreams: 1 DissMCV.pdf: 1396676 bytes, checksum: d3fa0ce7f0f0504dfe6926528b1e7366 (MD5) / Made available in DSpace on 2016-09-21T18:23:33Z (GMT). No. of bitstreams: 1 DissMCV.pdf: 1396676 bytes, checksum: d3fa0ce7f0f0504dfe6926528b1e7366 (MD5) Previous issue date: 2015-10-16 / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / The orexins are hypothalamic neuropeptides involved in an array of functions such as regulation of sleep/wake states and chemoreception to CO2/pH. The Locus coeruleus (LC) is a chemosensitive site and expresses an extensive population of orexin receptor 1 (OX1R). Here we tested the hypothesis that OX1Rs located in the LC participate in the ventilatory response to hypercapnia in a vigilance state and diurnal cycle-dependent manner. To this end, we performed unilateral injections of SB-334867 (OX1R antagonist, 5 mM) into the LC of male Wistar rats and evaluated the ventilatory response to 7% CO2 during wakefulness and sleep in the dark and light phase of the diurnal cycle. Hypercapnia induced an increase in ventilation in all groups compared to normocapnic values. However, the injection of (SB-334867) promoted an attenuation of the hypercapnic chemorreflex during wakefulness, due to changes in VT. In addition, microinjection of SB-334867 decreases the wakefulness time during dark phase. We suggest that projections of orexin-containing neurons to the LC contribute, via OX1Rs, to the hypercapnic chemoreflex during wakefulness in the dark phase. / As orexinas são neuropeptídeos hipotalâmicos envolvidos em uma variedade de funções, tais como, na regulação do ciclo sono-vigília e na quimiorecepção ao CO2 / pH. O locus coeruleus (LC) é um núcleo quimiossensível e expressa uma extensa população de receptor de orexina 1 (OX1R). Portanto, nós testamos a hipótese de que OX1Rs localizados no LC participam da resposta ventilatória à hipercapnia no estado de vigília de maneira dependente do ciclo claro e escuro. A participação da neurotransmissão orexinérgica do LC na resposta ventilatória à hipercapnia foi avaliada por meio da microinjeção do antagonista de receptor-1 OXR- 1 (SB-334867, 5 mM) no LC de ratos Wistar não anestesiados durante o sono e a vigília nas fases clara e escura. Nossos resultados demonstraram que a hipercapnia induz um aumento significativo da ventilação em todos os grupos comparado aos valores de ventilação na normocapnia. No entanto, na fase escura, a microinjeção do SB-334867 promoveu uma atenuação do quimiorreflexo hipercápnico durante a vigília, mas não durante o sono devido à diminuição do volume corrente VT. Adicionalmente, a microinjeção do SB-334867 diminui o tempo que os animais passam acordados em normocapnia na fase escura. Portanto, nossos dados sugerem que as projeções orexinérgicas para o LC atuando em OXR-1 exercem uma modulação excitatória na resposta ventilatória ao CO2 durante a vigília na fase escura. / 2014/00330-3