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Efeito da luz e endotelina no mecanismo molecular do relógio em melanóforos de Xenopus laevis / Effect of light and endothelin on clock molecular mechanisms in Xenopus laevis melanophoresMoraes, Maria Nathália de Carvalho Magalhães 17 December 2014 (has links)
Os ciclos claro-escuro (CE) são considerados importantes pistas para o ajuste de relógios biológicos. Alças de retroalimentação positiva e negativa de transcrição e tradução de genes de relógio são a base molecular subjacente tanto a relógios centrais como periféricos. A opsina não visual, melanopsina (Opn4), expressa na retina de mamíferos, é considerada o fotopigmento circadiano pois é responsável pelo ajuste do relógio biológico endógeno. Este fotopigmento também está presente nos melanóforos de Xenopus laevis, onde ele foi descrito pela primeira vez, mas seu papel nestas células ainda não está completamente esclarecido. Espécies de vertebrados não mamíferos expressam duas ou mais melanopsinas e, no caso de X. laevis, há dois genes, Opn4m and Opn4x. Melanóforos de X. laevis respondem à luz com dispersão dos grânulos de melanina, a resposta máxima sendo atingida no comprimento de onda correspondente àquele de excitação máxima da melanopsina. Entre vários hormônios, endotelinas também dispersam os melanossomos em melanóforos de Xenopus através de via similar àquela evocada pela luz. Tendo esses fatos em mente, decidimos investigar se a luz e a endotelina modulam a expressão de genes de relógio em melanóforos de Xenopus, usando PCR quantitativo para avaliar os níveis relativos de RNAm de Per1, Per2, Clock e Bmal1. Ciclos CE promoveram alterações temporais na expressão de Per1, Per2 e Bmal1. Pulsos de 10 min de luz azul aumentaram a expressão de Per1 e Per2, diminuíram a expressão de Opn4x, mas não tiveram efeito sobre Opn4m. Ainda mais, diferentes localizações foram mostradas para cada melanopsina: imunorreatividade para OPN4x foi vista principalmente na membrana celular, enquanto OPN4m foi imuno-localizada no núcleo. Estes resultados em conjunto apontam para funções diferenciais das duas melanopsinas neste modelo. A translocação de grânulos de melanina foi maior quando um pulso de luz azul foi aplicado na presença de endotelina ET-3. E os níveis de RNAm de Clock exibiram variação temporal em melanóforos submetidos a CE após tratamento com ET-3 10-9M, enquanto a expressão de Per1 não foi afetada pelo tratamento hormonal. Em adição, ensaios farmacológicos indicaram que as respostas de Per1 e Per2 à luz azul são evocadas através da ativação da via de fosfoinositídeos, com crosstalks com GMPc/proteina quinase G (PKG) para ativar os genes de relógio. Estes dados sugerem a participação de melanopsina na foto-ativação de genes de relógio, e apontam para uma participação menor de endotelina como sincronizador desta linhagem celular. Nossos resultados constituem uma importante contribuição ao campo emergente dos relógios periféricos os quais, em espécies de não mamíferos têm sido mais extensivamente estudados em Drosophila melanogaster e Danio rerio. Dentro deste contexto, nós mostramos que os melanóforos de Xenopus laevis representam um modelo ideal para a compreensão da modulação de ritmos circadianos por luz e hormônios / Light-dark cycles (LD) are considered important cues to entrain biological clocks. Positive and negative feedback loops of clock gene transcription and translation are the molecular basis underlying the mechanism of both central and peripheral clocks. The non-visual opsin, melanopsin (Opn4), expressed in the mammalian retina, is considered a circadian photopigment because it is responsible of entraining the endogenous biological clock. This photopigment is also present in the melanophores of Xenopus laevis, where it was first described, but its role in these cells is not fully understood. Non-mammalian vertebrate species express two or more melanopsins, and in X. laevis there are two melanopsin genes, Opn4m and Opn4x. X. laevis melanophores respond to light with melanin granule dispersion, the maximal response being achieved at the wavelength of melanopsin maximal excitation. Among various hormones, endothelins also disperse melanosomes in Xenopus melanophores through a similar pathway as light does. Therefore, we decided to investigate whether light and endothelin modulate clock gene expression in Xenopus melanophores, using quantitative PCR to evaluate the relative mRNA levels of Per1, Per2, Clock and Bmal1. LD cycles elicited temporal changes in the expression of Per1, Per2 and Bmal1. A 10 min pulse of blue light increased the expression of Per1 and Per2, decreased Opn4x expression, but had no effect on Opn4m. In addition, a different localization was shown for each melanopsin: immunoreactivity for OPN4x was mainly seen in the cell membrane, whereas OPN4m was immunolocalized in the nucleus. These results taken together point to a differential role for each melanopsin in this model. Melanosome translocation was greater when a blue light pulse was applied in the presence of endothelin ET-3. And mRNA levels of Clock exhibited temporal variation in melanophores under LD cycles after 10-9 M ET-3 treatment, whereas Per1 expression was not affected by the hormone treatment. In addition, pharmacological assays indicated that Per1 and Per2 responses to blue light are evoked through the activation of the phosphoinositide pathway, which crosstalks with cGMP/protein kinase G (PKG) to activate the clock genes. These data suggest the participation of melanopsin in the photo-activation of clock genes and point to a minor role of endothelin as synchronizer for this cell line. Our results add an important contribution to the emerging field of peripheral clocks, which in non-mammalian species have been mostly studied in Drosophila melanogaster and Danio rerio. Within this context, we show that Xenopus laevis melanophores represent an ideal model to understanding circadian rhythms modulation by light and hormone
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Endokrin wirksame Stoffe (endocrine disruptors) und deren Wirkungen auf die Sexualdifferenzierung bei Amphib Xenopus laevisBögi, Christian 26 February 2003 (has links)
Die vorliegende Arbeit befasst sich mit der Erweiterung des etablierten Stu-dienmodells Xenopus laevis zur Untersuchung der Wirkung von endocrine disruptors auf die Reproduktionsbiologie von Amphibien. Um einen Einblick in die grundlegenden Mechanismen der sexuellen Differenzierung von Amphibien zu gewinnen, wurden die Konzentrationen bestimmt, mit denen androgene und estrogene Sexualsteroide während der larvalen Entwicklung in verschiedenen Stadien von Xenopus vorliegen. Parallel wurde das Auftreten der korrespondierenden Rezeptoren im Verlauf der Entwicklung untersucht, über welche die hormonelle Wirkung vermittelt wird. Auf der Basis der gewonnenen Erkenntnisse konnte eine neue Hypothese zur sexuellen Differenzierung von Amphibien entwickelt und vorgestellt werden. Sie stellt das Enzym 5alpha-Reduktase, das die Umwandlung von Testosteron in das potentere und nicht weiter aromatisierbare Androgen Dihydrotestosteron (DHT) bewerkstelligt, in den Mittelpunkt des Prozesses der Geschlechtsdifferenzierung. Abhängig von der genetisch bedingten Expression dieses Enzyms kommt es zu einem höheren oder niedrigeren Auftreten des DHT und damit zu Unterschieden im Verhältnis von DHT zu Estradiol (E2). Der Charakter dieses Verhältnisses scheint der entscheidende Auslöser für die Entwicklung eines weiblichen oder männlichen Phänotyps zu sein. In einem zweiten, anwendungsorientierten Teil wurde untersucht, in wie weit die bislang auf Laboruntersuchungen beschränkte Arbeit mit X. laevis auf Feldstudien erweiterbar ist und ob sich auf diese Weise gewonnene Daten auf die Situation heimischer Amphibien übertragen lassen. Parallele Expositionen des Krallenfrosches einerseits und des Grasfrosches (Rana temporaria) andererseits gegenüber realen Medien unter Freilandbedingungen bestätigten die hervorragende Eignung des Studienmodells X.laevis zur Beurteilung endokriner Belastungssituationen. Darüber hinaus konnte gezeigt werden, dass sich durch die Verwendung von Festphasenextrakten die endokrinen Wirkungen komplexer Matrizes unter standardisierten Laborbedingungen charakterisieren lassen. Rezeptorbindungsstudien sowie Untersuchungen zur Genexpression spezifischer Marker, histologische Betrachtungen von Gonadengewebe und die Bestimmung von Geschlechterverhältnissen ermöglichten Aussagen auf vielfältigen Nachweisebenen. Auf diese Weise konnte das Potenzial, mit dem Proben jeder Art, sowohl durch kurz- als auch durch langfristige Exposition, adverse Effekte auf das amphibische Hormonsystem hervorrufen können, umfassend und differenziert analysiert werden. / The presented work aims to contribute to the various opportunities of studying the effects of endocrine disruption on sexual differentiation in amphibians provided by the well established model Xenopus laevis. In order to gain insight into the basic mechanisms underlying the sexual differentiation in amphibians, the concentrations of androgen and estrogen sexual steroids during several stages of the larval development of Xenopus were determined. In parallel, the ocurrence of the corresponding receptors, which mediate the effects of the respective hormones, was observed. Based on the results of the studies described, a new hypothesis regarding sexual differentiation in amphibians is presented, which assignes the enzyme 5alpha-reductase as the central element of sexual development. This enzyme converts the androgen testosterone into dihydrotestosterone, which can not be aromatized into estradiol. Depending on the genetic sex of the indivual, genexpression of 5a-reductase may differ and therefore lead to a characteristic ratio of androgens and estrogens. We suggest, that this ratio might be the essential trigger for amphibians to develop into a male or a female. A second part aimed to enlarge the Xenopus model to the use in field studies and to proof the transferability of such data to the situation of endemic amphibians. Exposure in parallel of Xenopus on one hand and the green frog Rana temporaria on the other to the effluent of a bavarian wastewater treatment plant revealed the exceeding suitability of the model to asess the endocrine charge of the environment. Furthermore, the use of solid phase extracts derived from natural samples allowed the characterization of the respective endocrine potential under standardized laboratory conditions. Rezeptor binding studies, detection of genexpression of specific biomarkers, histological examination of gonadal tissue and the determination of sex ratios provided the evaluation of effects on several levels of investigation. By this means the Xenopus model offers the opportunity to assess the ability of any kind of sample to cause endocrine impacts on amphibians after short time as well as after long time exposure in a broad and at the same time differentiated way.
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Efeito da luz e endotelina no mecanismo molecular do relógio em melanóforos de Xenopus laevis / Effect of light and endothelin on clock molecular mechanisms in Xenopus laevis melanophoresMaria Nathália de Carvalho Magalhães Moraes 17 December 2014 (has links)
Os ciclos claro-escuro (CE) são considerados importantes pistas para o ajuste de relógios biológicos. Alças de retroalimentação positiva e negativa de transcrição e tradução de genes de relógio são a base molecular subjacente tanto a relógios centrais como periféricos. A opsina não visual, melanopsina (Opn4), expressa na retina de mamíferos, é considerada o fotopigmento circadiano pois é responsável pelo ajuste do relógio biológico endógeno. Este fotopigmento também está presente nos melanóforos de Xenopus laevis, onde ele foi descrito pela primeira vez, mas seu papel nestas células ainda não está completamente esclarecido. Espécies de vertebrados não mamíferos expressam duas ou mais melanopsinas e, no caso de X. laevis, há dois genes, Opn4m and Opn4x. Melanóforos de X. laevis respondem à luz com dispersão dos grânulos de melanina, a resposta máxima sendo atingida no comprimento de onda correspondente àquele de excitação máxima da melanopsina. Entre vários hormônios, endotelinas também dispersam os melanossomos em melanóforos de Xenopus através de via similar àquela evocada pela luz. Tendo esses fatos em mente, decidimos investigar se a luz e a endotelina modulam a expressão de genes de relógio em melanóforos de Xenopus, usando PCR quantitativo para avaliar os níveis relativos de RNAm de Per1, Per2, Clock e Bmal1. Ciclos CE promoveram alterações temporais na expressão de Per1, Per2 e Bmal1. Pulsos de 10 min de luz azul aumentaram a expressão de Per1 e Per2, diminuíram a expressão de Opn4x, mas não tiveram efeito sobre Opn4m. Ainda mais, diferentes localizações foram mostradas para cada melanopsina: imunorreatividade para OPN4x foi vista principalmente na membrana celular, enquanto OPN4m foi imuno-localizada no núcleo. Estes resultados em conjunto apontam para funções diferenciais das duas melanopsinas neste modelo. A translocação de grânulos de melanina foi maior quando um pulso de luz azul foi aplicado na presença de endotelina ET-3. E os níveis de RNAm de Clock exibiram variação temporal em melanóforos submetidos a CE após tratamento com ET-3 10-9M, enquanto a expressão de Per1 não foi afetada pelo tratamento hormonal. Em adição, ensaios farmacológicos indicaram que as respostas de Per1 e Per2 à luz azul são evocadas através da ativação da via de fosfoinositídeos, com crosstalks com GMPc/proteina quinase G (PKG) para ativar os genes de relógio. Estes dados sugerem a participação de melanopsina na foto-ativação de genes de relógio, e apontam para uma participação menor de endotelina como sincronizador desta linhagem celular. Nossos resultados constituem uma importante contribuição ao campo emergente dos relógios periféricos os quais, em espécies de não mamíferos têm sido mais extensivamente estudados em Drosophila melanogaster e Danio rerio. Dentro deste contexto, nós mostramos que os melanóforos de Xenopus laevis representam um modelo ideal para a compreensão da modulação de ritmos circadianos por luz e hormônios / Light-dark cycles (LD) are considered important cues to entrain biological clocks. Positive and negative feedback loops of clock gene transcription and translation are the molecular basis underlying the mechanism of both central and peripheral clocks. The non-visual opsin, melanopsin (Opn4), expressed in the mammalian retina, is considered a circadian photopigment because it is responsible of entraining the endogenous biological clock. This photopigment is also present in the melanophores of Xenopus laevis, where it was first described, but its role in these cells is not fully understood. Non-mammalian vertebrate species express two or more melanopsins, and in X. laevis there are two melanopsin genes, Opn4m and Opn4x. X. laevis melanophores respond to light with melanin granule dispersion, the maximal response being achieved at the wavelength of melanopsin maximal excitation. Among various hormones, endothelins also disperse melanosomes in Xenopus melanophores through a similar pathway as light does. Therefore, we decided to investigate whether light and endothelin modulate clock gene expression in Xenopus melanophores, using quantitative PCR to evaluate the relative mRNA levels of Per1, Per2, Clock and Bmal1. LD cycles elicited temporal changes in the expression of Per1, Per2 and Bmal1. A 10 min pulse of blue light increased the expression of Per1 and Per2, decreased Opn4x expression, but had no effect on Opn4m. In addition, a different localization was shown for each melanopsin: immunoreactivity for OPN4x was mainly seen in the cell membrane, whereas OPN4m was immunolocalized in the nucleus. These results taken together point to a differential role for each melanopsin in this model. Melanosome translocation was greater when a blue light pulse was applied in the presence of endothelin ET-3. And mRNA levels of Clock exhibited temporal variation in melanophores under LD cycles after 10-9 M ET-3 treatment, whereas Per1 expression was not affected by the hormone treatment. In addition, pharmacological assays indicated that Per1 and Per2 responses to blue light are evoked through the activation of the phosphoinositide pathway, which crosstalks with cGMP/protein kinase G (PKG) to activate the clock genes. These data suggest the participation of melanopsin in the photo-activation of clock genes and point to a minor role of endothelin as synchronizer for this cell line. Our results add an important contribution to the emerging field of peripheral clocks, which in non-mammalian species have been mostly studied in Drosophila melanogaster and Danio rerio. Within this context, we show that Xenopus laevis melanophores represent an ideal model to understanding circadian rhythms modulation by light and hormone
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Die Regulation der Pankreasentwicklung von Xenopus laevis durch Transkriptionsfaktornetzwerke / Transcription factor networks directing pancreas development in Xenopus laevisJäckh, Christine 29 October 2008 (has links)
No description available.
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Identifikation und funktionelle Analyse von Xdach1 und Xeya3 als morphogenetische Faktoren der Kopfentwicklung von Xenopus laevis / Identification and functional analysis of Xdach1 and Xeya3 as morphogenetic factors of head development in Xenopus laevisKriebel, Martin 26 January 2005 (has links)
No description available.
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Elektrophysiologische Untersuchungen von Transporteigenschaften des Natrium-Dikarboxylat-Kotransporters aus der Flunderniere / Electrophysiological characterization the sodium-dicarboxylate cotransporter of the flounder kidneyHerbst, Christine 12 May 2010 (has links)
No description available.
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Zur Kodierung der Konzentration von Odoranzien in Zellen des Bulbus olfactorius von larvalen Xenopus-laevis-Fröschen / Coding of concentrations of odours in the olfactory bulb of the tadpole of the Xenopus laevisRöttger, Johannes 20 June 2011 (has links)
No description available.
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Die Funktion des Wnt Antagonisten XsFRP5 während der frühembryonalen Musterbildung des Entoderms in Xenopus laevis / The role of the secreted Wnt antagonist XsFRP5 in endodermal organogenesis in Xenopus embryosDamianitsch, Katharina 29 April 2008 (has links)
No description available.
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Imaging-Analyse dopaminerger Wirkungen am olfaktorischen Nerven von Xenopus-laevis-Larven / Imaging analysis of dopaminergic effects on the olfactory nerv of xenopus laevis tadpoles.Baßfeld, Eiko 07 November 2013 (has links)
No description available.
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An investigation into the critical domains and function of XMI-ER1 during xenopus development /Teplitsky, Yoella, January 2003 (has links)
Thesis (M.Sc.)--Memorial University of Newfoundland, 2004. / Bibliography: leaves 130-141.
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