Intranuclear rodlets (INRs) are poorly understood intranuclear bodies originally identified within neuronal nuclei on the basis of their unique morphology. Their mechanism of formation, biochemical composition and physiological significance are largely unknown. To gain insight into the molecular regulators of INR formation, mice with a conditional adult β cell-specific knockout of the master regulator of β-cell metabolism, Lkb1 protein kinase (LABKO mice) were studied. The proportion of beta cells containing INRs was significantly reduced in LABKO mice. Further examination ruled out mTOR and Mark2 as downstream effectors of Lkb1 knockout INR phenotype. Instead it identified the mTOR pathway as an independent regulator of INR formation. To investigate INR changes in a pathophysiological context, β cell INRs were examined in two models of human metabolic syndrome: (1) mice maintained on a high-fat diet and (2) leptin-deficient ob/ob mice. Significant INR reduction was observed in both models. Taken together, our results support the view that INR formation in pancreatic β cells is a dynamic and regulated process. The substantial depletion of INRs in LABKO and obese diabetic mice suggests their relationship to β cell function and potential involvement in diabetes pathogenesis. The significance of these findings was further underscored by the demonstration of INRs in human endocrine pancreas, suggesting their potential relevance to the development of metabolic syndrome in humans. The existence of biochemically distinct subtypes of INRs has been suggested by previous reports of differential immunological staining of INRs in neurochemically distinct neuronal populations. Here, a novel variant of INR has been identified that is immunoreactive for the 40kDa huntingtin associated protein and ubiquitin; and evidence was provided for the existence of additional INR subtypes sharing ubiquitin immunoreactivity as a common feature. Selective association of these INRs with melanin concentrating hormone and tyrosine hydroxylase immunoreactive neurons of the hypothalamus and the locus coeruleus was described. It was also demonstrated for the first time that biochemically distinct INR subtypes can co-exist within a single nucleus where they engage in non-random spatial interactions. These findings highlight the biochemical diversity and cell type specific expression of these enigmatic intranuclear structures. On the basis of these findings and previous literature a hypothesis is proposed as to the overall functional significance of INRs in the cell nucleus.

Orientador: Eduardo Rochete Ropelle / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Aplicadas / Made available in DSpace on 2018-08-24T08:41:08Z (GMT). No. of bitstreams: 1 Silva_VagnerRamonRodrigues_M.pdf: 4649552 bytes, checksum: 7f2da3d7826eb50aa18465d79242f6d6 (MD5) Previous issue date: 2013 / Resumo: A ingestão alimentar e o gasto energético são minuciosamente regulados por neurônios específicos localizados no hipotálamo. Durante as duas últimas décadas, a localização dos receptores da leptina em núcleos hipotalâmicos, bem como a descrição da via de transmissão intracelular disparado por este hormônio em neurônios hipotalâmicos, foi determinante para o entendimento do controle da ingestão alimentar e do gasto energético. Cada vez mais os distúrbios alimentares associados a doenças como obesidade são relacionados à disfunções na transmissão do sinal da leptina no hipotálamo. O processo inflamatório subclínica frequentemente observado em modelos experimentais de obesidade estão diretamente associados à distintos mecanismos de resistência à leptina no hipotálamo e resultam em aumento da ingestão alimentar e ganho de peso corporal. Por outro lado, estudos demonstram que o exercício físico é capaz de aumentar a sensibilidade da leptina no hipotálamo de animas obesos através de citocinas anti-imflamatórias, contudo, esses mecanismos permanecem apenas parcialmente conhecidos. Recentemente, a proteína S1PR1 (sphingosine-1-phosphate receptor-1) foi descrita como uma molécula com alta capacidade de exercer potentes efeitos sinérgicos sobre a via de sinalização da leptina, sustentando a ativação da via Jak2/STAT3 em algumas linhagens celulares. Assim, o presente estudo tem por objetivo investigar o os efeitos do exercício físico sobre a atividade da SIPR1 e a sensibilidade à leptina em hipotálamo de roedores obesos. Acreditamos que a realização do presente estudo contribuirá para caracterizar a participação da S1PR1 na sinalização da leptina no hipotálamo, bem como determinar os efeitos do exercício físico sobre a atividade da S1PR1 neuronal / Abstract: The food intake and energy expenditure are closely regulated by specific neurons in the hypothalamus. During the last two decades, the location of the leptin receptor in hypothalamic nuclei as well as the description of the route of transmission Intracellular triggered by this hormone in hypothalamic neurons, were crucial to the understanding of the control of food intake and energy expenditure.Increasingly, eating disorders, diseases associated with obesity are related to signal transmission malfunction of leptin in the hypothalamus. The subclinical inflammatory process frequently observed in experimental models of obesity are directly associated with distinct mechanisms of leptin resistance in the hypothalamus and result in increased food intake and body weight gain. Furthermore, studies have shown that physical exercise can increase the sensitivity of leptin in the hypothalamus of obese animals, through of antiinflammatory cytokines, however, these mechanisms remain only partially understood. Recently, the protein S1PR1 (sphingosine-1-phosphate receptor-1) was described as a molecule with high ability to exert potent synergistic effects on the signaling pathway of leptin, supporting the activation of Jak2/STAT3 in some cell lines. Thus, this project aims to investigate the effects of exercise on the activity of SIPR1 and leptin sensitivity in hypothalamus of obese rodents. We believe that the completion of this project will contribute to characterize the involvement of S1PR1 in leptin signaling in the hypothalamus, and to determine the effects of exercise on the activity of neuronal S1PR1 / Mestrado / Metabolismo e Biologia Molecular / Mestre em Ciências da Nutrição e do Esporte e Metabolismo

Orientador: Gabriel Forato Anhê / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-27T14:26:05Z (GMT). No. of bitstreams: 1 Mesquita_CarolineCosta_D.pdf: 2692158 bytes, checksum: 5f07d43267a2e976f9b9818ec63ba452 (MD5) Previous issue date: 2015 / Resumo: Os ritmos circadianos de ingestão alimentar se estabelecem com objetivo de manter a homeostasia de nutrientes no meio celular, frente a variações intrínsecas ao ciclo claro/escuro. Neste sentido, a gliconeogênese de roedores é suprimida no período noturno, no qual há ocorrência de um surto alimentar bifásico que compreende aproximadamente 90% de todo o aporte calórico diário. Estes eventos apresentam, entre si, uma relação causal, onde o próprio aumento dos nutrientes circulantes, principalmente a glicose, controla a gliconeogênese. O padrão inverso é observado na fase clara do ciclo claro/escuro. Recentes estudos têm demonstrado que, a ativação farmacológica de vias da Unfolded Protein Response (UPR), no sistema nervoso central, resulta em resistência à ação anorexigênica da insulina e, consequentemente, aumento da ingestão alimentar através de um mecanismo não completamente esclarecido. A UPR é uma resposta celular adaptativa que atenua a taxa de tradução de mRNAs, aumenta a proteólise e, deste modo, recupera o fenótipo celular. Esta reposta, quando ativada cronicamente, pode resultar em morte celular programada e resistência à insulina. No entanto, ainda não estava claro se a via do ATF6 da UPR tem, de fato, uma relação com o ritmo alimentar. Para responder a esse questionamento, realizamos análises da variação circadiana das proteínas envolvidas na via da UPR, imunoprecipitações com animais adrenalectomizados, e aplicação de dexametasona subcutânea. Os resultados demonstram que o ATF6 teve um aumento noturno atingindo o máximo de transição da fase clara para fase escura. A partir desse fato, foram realizados ensaios de imunuprecipitações em ratos adrenalectomizados para evidenciar as associações dos complexos CRTC2/ATF6 e CRTC2/CREB1. Contudo, nossos dados suportam a hipótese que os níveis fisiológicos de glicocorticóides podem reprimir a expressão CRH e estimular a ingestão de alimentos, através de uma via dependente de ATF6. Estes eventos, por consequência, poderão reduzir a atividade transcricional do CREB1, sobre o CRH, corroborando com os dados da literatura que apontam que os glicocorticóides endógenos dos roedores (principalmente a corticosterona) exercem um conhecido papel no controle da ingestão alimentar, decorrente principalmente da modulação da expressão do neurotransmissor anorexigênico CRH / Abstract: The circadian cycles of food intake have an important role in the homeostasis of cellular environment, acting over intrinsic changes to the sleep/wake cycle. Therefore, mice gluconeogenesis is suppressed at night where a food outbreak, responsible by 90% of all daily caloric ingestion, occurs. These events are connected by a causal relation, where the current nutrient increasing, mostly glucose, controls gluconeogenesis. The oppose pattern is verified during the light stage of the sleep/wake cycle. Recent studies have indicated that pharmacological activation of Unfolded Protein Response (UPR) pathways, at central nervous system, implies in resistance to the anorectic insulin effect, and, consequently, increase of the food intake activity trough a not fully explained engine. UPR is an adaptive cellular response that reduces mRNA¿s transcriptional rate, increases proteolysis, and therefore, restores cellular phenotype. This response, when constantly enabled, may induce cellular death and insulin resistance. However, it was not clear yet if the UPR¿s ATF6 pathway has, indeed, a connection with the feed rhythm. To answer to this question, we did several analysis of the circadian variation of the proteins connected to the UPR¿s pathway, adrenalectomized animals immunopreciptations and subcutaneous dexamethasone applications. The results demonstrated that ATF6 has a nightly increase and has reached the maximum of transcription rate from the light to the dark cycle. From this point, it was made immunopreciptations tests in adrenalectomized mice to evidence the association between CRTC2/ATF6 and CRTC2/CREB1 complexes. However, our data support the hypothesis that the physiological levels of glucocorticoids may suppress CHR expression and stimulate food intake trough an ATF6 dependent pathway. Those events, therefore, may reduce CREB1¿s transcriptional activity, confirming other studies data that indicates that endogenous mice glucocorticoids (mainly corticosterone) play an well-known role in food intake control, mainly due from modulation of the expression of the anorectic neurotransmitter CRH / Doutorado / Farmacologia / Doutora em Farmacologia

A avicultura brasileira tem alcançado altos índices de desempenho na produção de carne e ovos, como resultado da atualização constante de tecnologias no setor. A biotecnologia vem contribuindo nesse sentido, atuando especialmente em programas de seleção de animais com maior potencial de desenvolvimento e crescimento. Como toda a fisiologia animal é controlada direta e/ou indiretamente pela hipófise e hipotálamo, este trabalho propôs identificar e analisar genes expressos nestas estruturas de galinhas de duas linhagens divergentes quanto ao potencial de crescimento e avaliar a expressão dos genes correspondentes a transcritos cuja identidade não pôde ser revelada (sem similar nos bacos de dados), uma vez que estes podem representar possíveis genes novos. Para isto, foram construídas e analisadas bibliotecas a partir da hipófise e hipotálamo de aves de 21 dias de idade de uma linha macho de corte (TT) e uma linhagem de postura (CC), provenientes da Embrapa Suínos e Aves. Um total de 4.286 ESTs válidas foi obtido (no mínimo150 pb com qualidade PHRED acima de 20), correspondendo a 2.133 ESTs da biblioteca da linhagem TT e 2.153 ESTs da biblioteca da linhagem CC. O exercício de montagem, via programa Cap3, revelou 3.074 seqüências únicas, sendo 1.643 da biblioteca da linhagem TT e 1.649 da CC. Estas seqüências únicas foram automaticamente classificadas, de acordo com as categorias do GeneOntology, sendo que as seqüências de ambas as bibliotecas apresentaram uma distribuição similar entre os termos. Após montagem das ESTs e análise do padrão de expressão digital das seqüências constituintes, dos 389 contigs obtidos, 194 foram compostos por seqüências diferencialmente expressas entre as bibliotecas. Foram detectados 28 contigs contendo SNPs linhagem específicos, sendo 52 SNPs TT específicos e 25 CC específicos. Um total de 146 ESTs não pôde ter sua identidade revelada, porém destas, 133 puderam ser localizadas no genoma da galinha e apresentaram pelo menos uma fase de leitura aberta (ORF). Após análise de expressão gênica, os genes correspondentes a 78 destas ESTs sem identidade apresentaram-se diferencialmente expressos entre pelo menos dois dos tecidos de aves de uma linhagem comercial de corte de 21 dias de idade estudados: hipófise, hipotálamo, cérebro, fígado e músculo. Os resultados apresentados são promissores e merecem ser investigados em estudos futuros com o objetivo de identificar marcadores moleculares para programas de seleção e melhoramento animal. / The brazilian aviculture has reached a high development level in both meat and egg production as a result of constant technological atualization in the sector. Biotechnology can contribute in this way acting in selection programs of animals that show higher growth and development potential. The global animal physiology is direct or indirectly controlled by pituitary and hypothalamus. The aim of this work was to identify and analyse genes expressed in these structures of two divergent line chickens according to growth potential and evaluate the gene expression of corresponding transcripts classified as "no hit", once these ESTs could represent new genes. In this way, two pituitary and hypothalamus libraries of 21 days broiler (TT) and a layer (CC) chickens supplied by Embrapa Swine and Poultry National Research Center were constructed and analysed. A total of 4,286 valid ESTs was obtained (showing at least 150 bp with PHRED quality above 20) corresponding to 2,133 ESTs of TT line library and 2,153 ESTs of CC line library. The clustering process by Cap3 resulted in 3,074 unique sequences corresponding to 1,643 TT library sequences and 1,649 of CC sequences. These unique sequences were automatically annotated according GeneOntology categories and both library sequences showed a similar distribution between the terms. After ESTs clustering and northern digital analysis, 389 contigs were obtained – 194 of them showed differentially expressed sequences between the libraries. A total of 28 contigs showed line specific SNPs, corresponding to 52 TT specific SNPs and 25 CC specific SNPs. 146 ESTs were classified as "no hit" but 133 of these sequences were localized in chicken genome and showed open reading frame (ORF). After gene expression analysis, the genes of 78 "no hit" ESTs showed different expression level between the five tissues of commercial broilers with 21 days old: pituitary, hypothalamus, brain, liver and muscle. These results are promising and must be more investigated in future studies to identify molecular markers for use in animal selection and improvement programs.

Many signaling pathways which are classically understood for their roles in early development are also known to be involved in tissue maintenance and adult energy homeostasis. Furthermore, dysfunction of these signaling pathways results in human diseases such as cancer. An in depth understanding of how developmentally important signaling pathways function in the adult will provide mechanistic insights into disease and potential new therapeutic targets. Herein Chapter 1, the Wnt, fibroblast growth factor (FGF), and Hedgehog (Hh) signaling pathways are discussed and examples of their relevance in development, adult homeostasis, and disease are provided. Wnt signaling provides an example of this concept as it has well described roles during both development and adult metabolism.<div><br></div><div> Work included in Chapter 2, investigates the regulation of adult energy homeostasis by a member of the endocrine FGF family, FGF19. The three endocrine FGFs, FGF19 (FGF15 in mice), FGF21, and FGF23 have well described roles in the regulation of metabolic processes in adults. While FGF23 is primarily involved in the regulation of phosphate and vitamin D homeostasis,FGF19 and FGF21 have shown similar pharmacological effects on whole body metabolism. Here, the importance of adaptive thermogenesis for the pharmacological action of FGF19 is explored. UsingUCP1KO animals we show that whole-body thermogenesis is dispensable for body weight loss following FGF19 treatment.<br></div><div><br></div><div>Finally, the potential involvement of Hh signaling in mediating the hyperphagia driven obesity observed in certain ciliopathies is explored in Chapter 3. Emerging evidence suggests cilia play an important role in the regulation of feeding behavior. In mammals, the hedgehog pathway is dependent on the primary cilium as an organizing center and defects in hedgehog signaling share some clinical symptoms of ciliopathies. Here, we characterized the expression of core pathway components in the adult hypothalamus. We show that neurons within specific nuclei important for regulation of feeding behavior express Hh ligand and members of its signaling pathway. We also demonstrate that the Hh pathway is transcriptionally upregulated in response to an overnight fast. This work provides an important foundation for understanding the functional role of Hh signaling in regulation of energy homeostasis. In its entirety, this work highlights the emerging clinical relevance of developmentally critical pathways in diseases associated with dysfunction of adult tissue homeostasis, such as obesity.<br></div>

L’apparition de la puberté et la régulation de la fertilité chez les mammifères sont contrôlées par un réseau neuronal complexe, situé principalement dans l'hypothalamus, et qui converge vers les neurones synthétisant l'hormone de libération des gonadotrophines (GnRH). Ces neurones régulent la sécrétion des gonadotrophines, la croissance et le fonctionnement des gonades. Le développement correct du système à GnRH, incluant des changements rapides dans l'expression et la signalisation de l’hormone GnRH au sein de cette population clairsemée de quelques centaines de neurones, est essentiel pour la maturation sexuelle et le fonctionnement normal de l'axe hypothalamo-hypophyso-gonadique. Lors du développement embryonnaire, ces neurones migrent de la placode olfactive vers leur emplacement définitif, l’hypothalamus, pour y recevoir les connexions afférentes qui permettront une libération pulsatile de la GnRH et la libération subséquente des gonadotrophines (l'hormone de stimulation des follicules (FSH) et l'hormone lutéinisante (LH)). Dès les années 90, l'oxyde nitrique (NO) a été identifié comme molécule clé dans la décharge pré-ovulatoire de GnRH/LH. En effet, de nombreux travaux ont suggéré que des interactions entre les neurones exprimant la forme neuronale de l’enzyme de synthèse du NO (la nNOS) et le système GnRH étaient impliquées dans le contrôle central de la fonction de reproduction à l'âge adulte. De plus, si le NO est reconnu depuis longtemps comme un acteur majeur du contrôle central de l’ovulation à l’âge adulte, la possibilité qu’il soit aussi impliqué dans la maturation sexuelle en régulant l’activité des neurones à GnRH à des stades précoces précédant la puberté n’a pas été explorée auparavant. Cependant, même si nous avons progressé dans la connaissance des interactions entre les neurones à nNOS et des différents acteurs importants de l’axe gonadotrope, l’identité moléculaire de ces neurones reste mal connue. Au cours de cette étude, nous avons recherché 1) l'identité moléculaire des neurones á nNOS dans l'hypothalamus au cours de développement 2) si le NO régule la migration et l’intégration des neurones à GnRH dans l’hypothalamus et 3) si le NO régule la maturation sexuelle. Pendant ma thèse nous avons répertorié pour la première fois les différents neurotransmetteurs et les principaux récepteurs dans les neurones à nNOS au cours du développement post-natal. De plus, les résultats de ma thèse montrent pour la première fois une implication de la signalisation du NO dans la migration des neurones à GnRH vers l'hypothalamus et font échos à l'identification d'une série de mutations de la NOS1 chez des patients atteints du syndrome de Kallmann, une maladie génétique congénitale rare qui associe une carence en GnRH, due à un défaut de migration neuronale, et une anosmie. Enfin, mes travaux montrent que le NO est un nouveau protagoniste dans la maturation post-natale du système à GnRH, la survenue de la puberté et l’acquisition de la capacité à se reproduire. Plus généralement, les résultats de ce travail de thèse permettent d’identifier de nouveaux mécanismes potentiellement responsables de troubles développementaux dans la mise en place des circuits neuronaux contrôlant l’axe gonadotrope chez les mammifères en général et l’homme en particulier. Nous espérons que ces résultats élargiront notre compréhension de la régulation de l'axe reproducteur, offrant ainsi des possibilités nouvelles de stratégies thérapeutiques contre les troubles de la fertilité. / The onset of puberty and the regulation of fertility in mammals are governed by a complex neural network, primarily in the hypothalamus, that converges onto gonadotropin-releasing hormone (GnRH)-producing neurons, the master regulators of gonadotropin secretion and postnatal gonadal growth and function. The proper development of the GnRH system, including timely changes in GnRH expression and signaling by this sparse population of a few hundred neurons, is essential for sexual maturation and the normal functioning of the hypothalamic-pituitary-gonadal axis. As the brain develops during embryogenesis, these neurons should move from the olfactory placode into the correct brain location in adequate numbers, and then establish the afferent connections that will allow the pulsatile release of GnRH peptide, and the subsequent release of the gonadotropins (follicle stimulating hormone, i.e FSH and luteinizing hormone, ie. LH). As early as in the 90’s NO was presented as a key molecule in the preovulatory GnRH/LH surge, and results from different groups, have suggested the interaction of NOS-containing neurons with the GnRH system, and their involvement in the regulation of reproductive capacity. Even though nitric oxide has now been long recognized as a key player in the central hormonal regulation of ovulation during adulthood, no one has considered the possibility that it could act in an earlier stage as the master regulator of GnRH neurons before puberty, hence participating in the actual maturation of the neuroendocrine axis. The relationship of nNOS-expressing neurons with other important molecules of the hypothalamic axis has been well studied, whilst the molecular identity of this neuronal NOS-expressing population is poorly documented. . To this end, we address the hitherto unaddressed questions concerning 1) the molecular identity of nNOS-expressing neurons in the developing hypothalamus, 2) the putative involvement of the NO molecule in the migration of GnRH neurons and the proper establishment of their afferent connections in the hypothalamic region and 3) the plausible determinant role of NO signaling in the maturation of the reproductive system. During this study we identified for the first time the cohort of the principal neurotransmitters and important receptors expressed by these cells in the hypothalamic region during development. Additionally, our results reveal for the first time an involvement of NO signaling in the migration of GnRH neurons in the hypothalamus and are in line with the identification of a series of NOS1 mutations in Kallmann syndrome (KS), a rare congenital genetic condition presenting a unique combination of GnRH deficiency, arising from a faulty migration of the neuronal population, and anosmia. Lastly, our study identifies NO as a novel protagonist during postnatal development, in the regulation of the onset of puberty and the acquisition of reproductive competence. Overall, the results of my Phd thesis identify putative new targets causing alterations of developmental programming under pathophysiological gestational environment in mammals in general, and in humans in particular. Here we thus provide new insights into the mechanisms by which the alteration of GnRH neuronal function leads to hypogonadotropic hypogonadism and infertility. We are hopeful that our results will expand our understanding of how the neuroendocrine axis is regulated and will possibly provide opportunities for therapeutic strategies against debilitating conditions.

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Die Hyperaktivität der HHN-Achse ist bei der Multiplen Sklerose umfassend untersucht und mehrfach bestätigt; der genaue Pathomechanismus bleibt jedoch ungeklärt. Das serotonerge System ist eng mit der HHN-Achse verbunden und besitzt die Eigenschaft, die HHN-Achse herabzuregulieren. Daraus ergab sich im Umkehrschluss, dass die HHN-Achsen-Hyperaktivierung bei Patienten mit Multipler Sklerose im Zusammenhang mit einer reduzierten serotonergen Verfügbarkeit stehen könnte. Um die HHN-Achse und das serotonerge System zu untersuchen, schlossen wir 17 Patienten mit Multipler Sklerose und 13 gesunde Kontrollpersonen ein. Die HHN-Achse wurde mittels des Dex-CRH-Tests untersucht und die zentrale Verfügbarkeit des präsynaptischen Serotonintransporters mittels einer [11C]DASB-PET in 31 Hirnregionen gemessen. Die Patientengruppe hatte insgesamt höhere Kortisolwerte, dementsprechend erniedrigt war die ACTH-Kortisol-Ratio, so dass wir das Vorliegen einer HHN-Achsen-Hyperaktivität bestätigen konnten. Am ausgeprägtesten waren die Unterschiede in der Gruppe der primär chronisch progredienten MS. Eine signifikant reduzierte Verfügbarkeit des Serotonintransporters fand sich im Hypothalamus, den Raphe-Kernen, limbischen Strukturen, dem linken Temporallappen und Thalamus, auch hier war die Gruppe der PPMS am stärksten betroffen. Zusätzlich erhobene Fragebögen hinsichtlich Depression, Fatigue und Lebensqualität (BDI, WEIMuS, WEIMuS 1, WEIMuS 2, EuroQol, EuroQolScale und VAS) zeigten durchgängig signifikant unterschiedliche Ergebnisse bei den Patienten im Vergleich zur Kontrollgruppe. Zusammenfassend, konnten wir darstellen, dass die HHN-Achsen-Aktivierung bei der Multiplen Sklerose in enger Verbindung mit der serotonergen Aktivität steht, eine direkte Korrelation fand sich jedoch nicht. Dennoch kann davon ausgegangen werden, dass die serotonergen Neurone durch ihre enge anatomische Verbindung zu einer negativen Modulation der HHN-Achse beitragen. Der Einsatz von SSRI könnte therapeutisches Potential bieten, indem die herabregulierte serotonerge Aktivität sowie HHN-Achse wieder normalisiert werden könnten.:Inhaltsverzeichnis I Abkürzungsverzeichnis III 1. Einleitung 1 1.1 Einführung in die Fragestellung 1 1.2 Multiple Sklerose 1 1.2.1 Definition, Epidemiologie, Ätiologie und Pathogenese 1 1.2.2 Diagnosekriterien, Verlaufsformen, Klinik und Therapie 6 1.2.3 Fatigue und Multiple Sklerose 9 1.3 Die Hypothalamus-Hypophysen-Nebennierenrinden-Achse 10 1.4 Das serotonerge System und die Hypothalamus-Hypophysen-Nebennierenrinden-Achse 13 1.5 Aufgabenstellung 16 2. Materialien und Methoden 17 2.1 Durchführung 17 2.2 Einschlusskriterien 17 2.3 Ausschlusskriterien 18 2.4 Allgemeiner Ablauf 19 2.5 Dexamethason-CRH-Test 19 2.6 Positronen-Emissions-Tomographie 20 2.7 Magnetresonanztomographie 22 2.8 Genotypisierung 22 2.9 Fragebögen zu Depression, Fatigue und Lebensqualität 22 2.9.1 Beck-Depressions-Inventar 22 2.9.2 Würzburger Erschöpfungsinventar bei Multipler Sklerose 23 2.9.3 Europäischer Lebensqualitäts-Fragebogen 23 2.10 Statistische Datenanalyse 24 I 3. Ergebnisse 25 3.1 Klinische und demographische Daten 25 3.2 Dexamethason-CRH-Test 27 3.3 Verfügbarkeit des Serotonintransporters 29 (mittlere Distribution Volume Ratios) 3.4 Lateralität des Serotonintransporters 32 3.5 Beziehung der neuroendokrinen Funktion zur Serotonintransporter-Verfügbarkeit 33 3.6 Fatigue und Depression 34 4. Diskussion 35 4.1 Dexamethason-CRH-Test und Hypothalamus-Hypophysen-Nebennierenrinden-Achse 35 4.2 Das serotonerge System 37 4.3 Fatigue 41 4.4 Lateralität 42 4.5 Methodenkritik 44 4.6 Therapeutischer Ausblick 45 5. Zusammenfassung der Arbeit 48 6. Literaturverzeichnis 50 7. Thematisch assoziierte Publikation als Zweitautorin 69 8. Anlagen 79 8.1 Fragebögen 79 8.2 Selbständigkeitserklärung 87 8.3 Lebenslauf 88 8.4 Danksagung 90 II 3. Ergebnisse 25 3.1 Klinische und demographische Daten 25 3.2 Dexamethason-CRH-Test 27 3.3 Verfügbarkeit des Serotonintransporters 29 (mittlere Distribution Volume Ratios) 3.4 Lateralität des Serotonintransporters 32 3.5 Beziehung der neuroendokrinen Funktion zur Serotonintransporter-Verfügbarkeit 33 3.6 Fatigue und Depression 34 4. Diskussion 35 4.1 Dexamethason-CRH-Test und Hypothalamus-Hypophysen-Nebennierenrinden-Achse 35 4.2 Das serotonerge System 37 4.3 Fatigue 41 4.4 Lateralität 42 4.5 Methodenkritik 44 4.6 Therapeutischer Ausblick 45 5. Zusammenfassung der Arbeit 48 6. Literaturverzeichnis 50 7. Thematisch assoziierte Publikation als Zweitautorin 69 8. Anlagen 79 8.1 Fragebögen 79 8.2 Selbständigkeitserklärung 87 8.3 Lebenslauf 88 8.4 Danksagung

Als Kopf der Hypothalamus-Hypophysen-Nebennierenrinden-Achse spielt der Hypothalamus eine Schlüsselrolle für die depressive Symptomatik und bei pathogenetischen Modellen affektiver Störungen. Für nahezu alle Ebenen dieser Hormonachse lassen sich Funktions- und Strukturveränderungen, insbesondere Volumenveränderungen bei uni- oder bipolaren affektiven Störungen nachweisen. Zum Hypothalamus existiert hingegen, neben histochemischen Analysen, nur ein explorativer post mortem Befund einer Volumenreduktion von bis zu 15.5% bei uni- oder bipolar affektiv Erkrankten. Die vorliegende Arbeit verfolgt das Ziel, Volumenveränderungen des Hypothalamus bei uni- und bipolaren affektiven Störungen in vivo nachzuweisen. Mittels der Hochfeld-MRT lässt sich der Hypothalamus seit einigen Jahren mit einer Auflösung von weniger als 1 mm detailgetreu abbil-den. Zur Beurteilung, ob diese Genauigkeit dem Studienziel gerecht wird, wird eingangs in einem vorab publizierten Literaturüberblick der aktuelle Forschungstand zu Strukturveränderungen des Hypothalamus bei uni- und bipolaren affektiven Störungen zusammengefasst und diskutiert. Die Überblicksarbeit kommt zu dem Urteil, dass auch in vivo Volumenreduktionen des Hypothalamus solcher Größenordnungen zu erwarten sind, dass sie mittels Hochfeldbildgebung nachgewiesen werden können. Zur präzisen Vermessung des Hypothalamusvolumens stellen anschließend zwei ebenfalls publizierte Methodenstudien die Entwicklung und Evaluation einer geeigneten Messmethodik anhand hochaufgelöster, T1-gewichteter 7 Tesla MRT-Aufnahmen vor. Sie umfasst eine Intensitätsstandardisierung sowie einen falschfarbengestützten Segmentierungsalgorithmus. Aufbauend auf diesen theoretischen und methodischen Vorarbeiten präsentiert die vierte publizierte Arbeit die weltweit ersten in vivo Daten zu Volumenveränderungen des Hypothalamus bei uni- und bipolaren affektiven Störungen. Im querschnittlichen Vergleich mit gesunden Probanden und unter Kontrolle des intrakraniellen Volumens und psychotroper Medikation konnten bei beiden Störungsbildern linksseitige Volumenvergrößerungen des Hypothalamus nachgewiesen werden. Diese sind möglicherweise ein strukturelles Korrelat histochemisch nachweisbarer Aktivitätssteigerungen hypothalamischer Kerngebiete. Alternativ können sie eine Aktivierung und Vermehrung der Gliazellen anzeigen. Schließlich kann eine Volumenzunahme des Hypothalamus auch auf eine Vergrößerung der Zellzwischenräume zurückgehen. Der relative Mangel an Gerüststrukturen könnte, infolge mechanischer Krafteinwirkungen bei der histologischen Gewebeaufbereitung, zu einer verstärkten Stauchung bei den Patienten führen und so den früheren, gegenteiligen post mortem Befund erklären. Zur Untersuchung der mikrostrukturellen Gewebeeigenschaften des Hypothalamus bei uni- und bipolaren affektiven Störungen soll daher in der Folge diffusionsgewichtete Bildgebung zum Einsatz kommen.:Kapitel 1 1.1 Der Hypothalamus als Vermittler zwischen Gehirn und Körper 1.2 Theoretische Einordnung und empirischer Kenntnisstand 1.2.1 Die HPA-Achse als Bindeglied zwischen Diathese und Stress. 1.2.2 Hirnstrukturelle und -funktionelle Korrelate affektiver Störungen. 1.3 Forschungsthema 1.3.1 Problemstellung. 1.3.2 Forschungsziele. 1.4 Fragestellungen und Hypothesen 1.4.1 In vivo Strukturveränderungen des Hypothalamus bei affektiven Störungen. 1.4.2 Wie gestaltet sich eine reliable Messmethode? 1.4.3 Welche Intensitätsstandardisierung optimiert die Bilddatenqualität? 1.4.4 Verringertes in vivo Hypothalamusvolumen bei affektiven Störungen. Kapitel 2 2.1 Review Artikel 2.2 Segmentierungsalgorithmus 2.3 Intensitätsstandardisierung 2.4 Patientenstudie Kapitel 3 3.1 Hauptergebnisse 3.1.1 Theoretische und methodische Vorarbeiten. 3.1.2 Patientenstudie. 3.2 Wissenschaftliche Bewertung und Einordnung der Hauptergebnisse 3.2.1 Fundierung der Hypothesen. 3.2.2 Stärken und Schwächen der Patientenstudie. 3.2.3 Bewertung der Messmethodik. 3.2.4 Inhaltliche Interpretation des explorativen Befunds. 3.2.5 Ausblick. Anhang 4.1 Literaturverzeichnis 4.2 Abkürzungsverzeichnis 4.3 Zusammenfassung 4.4 Summary 4.5 Publikationsverzeichnis 4.6 Selbsständigkeitserklärung 4.7 Nachweise über Anteile der Co-Autoren

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