Melatonina é um hormônio produzido principalmente pela pineal, possui caráter temporizador do meio interno, ação antioxidante, neuroprotetora e neurotrófica. Outro fator que age nos processos de neuroplasticidade é o exercício físico. Este é responsável por alterar parâmetros metabólicos nos animais, e trabalhos do nosso grupo demonstraram que tais adaptações metabólicas em ratos dependem da melatonina pineal. Diante disso investigamos se as adaptações neuroplásticas em ratos submetidos ao treinamento físico também estariam relacionadas à presença deste neuro-hormônio. Nossos resultados demonstram que a melatonina não influenciou a expressão gênica e quantificação proteica de indicadores da neurogênese, somente a realização do treinamento físico alterou alguns dos parâmetros avaliados. Os níveis de corticosterona se mostraram alterados nos animais pinealectomizados. A avaliação da expressão gênica circadiana com oito pontos ao longo das 24 horas revelou o comportamento variável do mRNA de algumas proteínas atreladas ao processo de plasticidade hipocampal / Melatonin is a hormone produced mainly by pineal gland, has timer character of the internal medium, antioxidant action, neuroprotective and neurotrophic. Another factor that acts in the processes of neuroplasticity is physical exercise. This is liable to change metabolic parameters in animals, and work by our group have shown that such metabolic adaptations in rats depend on pineal melatonin. Thus we investigated whether the neuroplastic adaptations in rats submitted to physical training also would be related to the presence of this neurohormone. Our results demonstrate that the melatonin did not influence the gene expression and protein quantification of indicators of neurogenesis, only the execution of the physical training has changed some of the parameters evaluated. The levels of corticosterone were changed in pinealectomized animals. The assessment of gene expression circadian with eight points along the 24 hours showed the variable behavior of the mRNA of some proteins linked to the process of hippocampal plasticity.

A esquizofrenia (EZ) é uma das doenças neuropsiquiátricas mais prevalentes, afetando cerca de 1% da população ao redor do mundo, sendo caracterizada pela presença de delírios, alucinações, disfunção cognitiva e apatia, entre outros. Dentre as hipóteses que procuram explicar as bases biológicas da EZ, a que tem tido um maior embasamento e credibilidade é a Hipótese do Neurodesenvolvimento, que afirma que a EZ é uma desordem de desenvolvimento do sistema nervoso central, originada nos estágios intermediários da vida intra-uterina. Ao mesmo tempo o envolvimento de componentes genéticos em EZ é fortemente sugerido, principalmente por estudos envolvendo gêmeos e famílias. Uma série de estudos de ligação, realizados com famílias contendo indivíduos afetados, têm apontado para diferentes locos cromossômicos que devem estar associados à EZ. Enquanto tais estudos sugerem locos candidatos, a descoberta dos genes envolvidos com a doença, mapeados nestes locos, trará grande progresso ao estudo da genética da EZ. Dessa forma, o objetivo deste estudo foi analisar a possível associação entre polimorfismos em genes envolvidos com neurodesenvolvimento, mapeados em locos importantes sugeridos por estudos de ligação, e a EZ. Após uma série de análises in silico e validações in vitro, um total de 41 polimorfismos, mapeados em 39 genes candidatos, foram selecionados neste estudo. Estes polimorfismos foram genotipados em amostras de DNA, obtidas de 200 pacientes esquizofrênicos e 200 controles da população Brasileira. Uma análise individual das freqüências genotípicas e alélicas de cada polimorfismo identificou quatro polimorfismos como associados à EZ, mapeados nos genes AKT1, MAP1B, FZD3, e NUMBL. Em seguida, esses quatro polimorfismos foram analisados em um segundo set amostral, composto por amostras de DNA de casos e controles Dinamarqueses, de forma a tentar replicar os achados com a amostra Brasileira. Para dois destes polimorfismos (NUMBL e FZD3) observamos uma tendência de associação na amostra dinamarquesa, o que reforça o possível envolvimento destes polimorfismos com a EZ. Avançamos na caracterização haplotípica do SNP de FZD3 em amostras do Brasil e da Dinamarca e também investigamos as conseqüências funcionais deste SNP, utilizando ensaios de gene-repórter em duas diferentes linhagens celulares. Por fim, uma análise multilocus preliminar, utilizando o método set association, foi realizada para os marcadores bialélicos por nós avaliados (SNPs e indels), a fim de identificar a existência de um set de genes que estivessem contribuindo em conjunto para a etiologia da EZ. Os resultados dessa análise apontaram para o efeito aditivo de 5 genes para a etiologia da EZ: FZD3, AKT1, MAP1B, SEMA6C e ADAM28. Os genes encontrados em nosso estudo como associados à EZ, tanto pela análise individual quanto pela análise multilocus, se encontram envolvidos direta ou indiretamente com as vias sinalizadoras de WNT e NOTCH, sugerindo o envolvimento destas duas vias sinalizadoras para a etiologia da doença. / Schizophrenia (SZ) is one of the most prevalent neuropsychiatry disorders, affecting about 1% of the world\'s population. SZ is characterized by the presence of delirium, hallucinations, cognitive dysfunction, apathy, etc. Among the hypotheses that try to explain the biological basis of SZ, the Neurodevelopment Hypothesis is one of the most accepted. This hypothesis states that SZ is a neurodevelopment disorder, originated during intermediate stages of the intrauterine life. On the other hand, the involvement of genetic components in the etiology of SZ has been suggested by twin and family studies. A series of linkage studies pointed different genomic loci associated with the disease. While these studies suggest candidate loci, the determination of which genes/genetic alterations inside these loci are involved with the disease will bring great progress to the study of SZ genetics. The objective of this study was to analyze the possible associations between DNA polymorphisms in genes related to neurodevelopment and mapped to genomic loci previously associated with SZ. After a series of in silico and experimental analysis, a total of 41 polymorphisms, mapped to 39 candidate genes, were selected for analysis. These polymorphisms were genotyped in DNA samples obtained for 200 SZ patients and paired 200 controls from the Brazilian population. An individual analysis of the genotypic and allelic frequencies of each polymorphism identified 4 polymorphisms as associated with SZ, mapped in the AKT1, MAP1B, FZD3, and NUMBL genes. These four polymorphisms were then analyzed in a second set of samples, derived from Danish cases and controls. A trend for association was observed for two of these polymorphisms (FZD3 and NUMBL), reinforcing their putative association with EZ. Haplotipic analysis was performed for the FZD3 SNP and its functional consequences were confirmed in two different cell lines by using gene reporter assays. Finally, multilocus analysis, performed by the use of the set association method, pointed to an additive effect of 5 genes contributing to the etiology of SZ: FZD3, AKT1, MAP1B, SEMA6C e ADAM28. All the genes pointed by our study as associated with SZ, by both the individual and the multilocus analysis, are directly or indirectly involved in the WNT and NOTCH pathways, which strongly suggests the involvement of these pathways in the etiology of SZ.

Les pathologies dépressives se caractérisent par des symptômes hétérogènes impliquant de nombreuses régions cérébrales. L’une d’entre elles, l’hippocampe, est le siège d’un processus physiologique, appelé neurogenèse, qui, chez l’adulte, serait impliqué dans l’étiologie de la dépression et la réponse au traitement antidépresseur.L’objectif de ce travail a été d’étudier le rôle des 4 étapes du processus de neurogenèse hippocampique dans l’action des antidépresseurs dans un modèle physiopathologique de la dépression, chez la Souris adulte. / Depression is characterized by heterogeneous symptoms in several brain regions. One of them, the hippocampus, is related to a phenomenon called neurogenesis, which seems to be linked to the etiology of depression and to the response of antidepressants. The main goal of this work was to study the relationships between all the 4 steps of hippocampal neurogenesis and antidepressants action in a model of depression in adult mouse.

La neurogenèse adulte du gyrus denté de l’hippocampe joue un rôle essentiel dans les processus mnésiques dépendants de l’hippocampe, mais son rôle dans des formes complexes de mémoire comme la mémoire épisodique n’a jamais été exploré. Le travail de cette thèse porte sur l’étude de l’implication des nouveaux neurones de l’hippocampe dans la mise en mémoire d’un souvenir épisodique à long terme. Nous avons développé une nouvelle tâche de mémoire épisodique reposant sur la présentation occasionnelle d’épisodes permettant d’encoder des informations de type « Quoi – Où – Dans quel contexte ». Nous montrons pour la première fois que les rats sont capables de se souvenir à très long terme de brefs épisodes de vie et d’utiliser cette mémoire d’une manière flexible. La caractérisation des profils de rétention permet d’accéder aux capacités individuelles de recollection des différents éléments du souvenir et montre que le rappel fiable de la mémoire épisodique nécessite l’intégrité de l’hippocampe et met en jeu un vaste réseau hippocampo-cortical dont l’activation est corrélée au rappel. Les performances de rats soumis à une irradiation focale de l’hippocampe montrent que la neurogenèse adulte hippocampique contribue de façon significative à la consolidation et au rappel fiable du souvenir épisodique. Ces résultats sont discutés dans le cadre d’une implication de la neurogenèse adulte dans la résolution de la mise en mémoire d’événements occasionnels dans le but de discriminer deux épisodes de vie proches, en lien avec les fonctions de séparation et de complétion de patterns de l’hippocampe. Par ailleurs, les mécanismes moléculaires qui sous-tendent le recrutement des nouveaux neurones lors d’un apprentissage restent inconnus. Nous avons analysé le rôle du gène immédiat précoce Zif268, acteur moléculaire essentiel dans les processus mnésiques, et montrons que ce gène joue un rôle crucial dans la sélection et le recrutement des nouveaux neurones lors de la mémorisation au cours de leur période critique d’intégration dans les réseaux neuronaux de l’hippocampe. Ce travail apporte des éléments nouveaux sur la participation des nouveaux neurones hippocampiques dans les processus mnésiques dans une situation à forte demande cognitive basée sur l’encodage d’une représentation intégrée et résolue d’événements occasionnels complexes, ainsi que sur les mécanismes qui sous-tendent leur recrutement. / Adult hippocampal neurogenesis plays a critical role in hippocampal-dependent memory, however its role in complex forms of memory such as episodic memory has not as yet been explored. The work presented in this thesis focuses on the issue of the involvement of newborn hippocampal neurons in long term episodic memory. We developed a new episodic memory task based on the presentation of occasional episodes allowing rats to encode “What – Where – In which context” information. We show for the first time that rats are able to remember on the long term brief past episodes of life and to use their episodic memory in a flexible manner. The characterization of retention profiles allows us to identify individual abilities in the recollection of the various elements of the memory and shows that episodic memory recall requires the integrity of the hippocampus and involves a hippocampo-cortical network, the activation of which correlates with recall performance. Performance of rats subjected to focal irradiation of the hippocampus shows that adult hippocampal neurogenesis contributes significantly to the consolidation and faithful recall of episodic memory. These results are discussed in the context of the implication of hippocampal newborn neurons in the resolution of memories of occasional events in order to discriminate different, but closely related episodes of life in relation to pattern separation and pattern completion functions of the hippocampus. Furthermore, the molecular mechanisms underlying the recruitment of newborn hippocampal neurons by learning remain to date unknown. We investigated the role of Zif268, an immediate early gene known to play an essential role in memory processes, and show that this gene plays a crucial role in the selection and recruitment of newborn hippocampal neurons by learning during their critical period of integration in hippocampal neural networks. Overall, this work brings new knowledge on the contribution of newborn hippocampal neurons to memory processes in a highly demanding cognitive situation based on the encoding of an integrated and high-resolution representation of complex occasional events, and on the mechanisms underlying their recruitment.

A odontologia moderna, mesmo usando as suas técnicas mais primorosas, na prática, ainda recupera a perda dental com implantes metálicos recobertos por coroas protéticas. Há um empenho coletivo dos cientistas em criar técnicas de desenvolvimento dental in vitro na busca por maneiras de recuperar, de maneira biológica, a ausência dental. Já é possível criar estruturas similares a dentes a partir de células-tronco de origem dental (polpa de dentes permanentes e decíduos) e não dental (células-tronco embrionárias, células-tronco da medula óssea e da crista neural) por meio de recombinação dos tecidos epiteliais e mesenquimais de germes dentais. As técnicas de reconstrução tecidual nunca estiveram tão perto do desenvolvimento da terceira dentição mas a ciência ainda tem muito a aprender no que concerne o estudo da biologia dental e engenharia de tecidos. Não basta saber como um dente se desenvolve; há de se entender como ele interage com o organismo do qual faz ou fará parte. É com esta preocupação que nos propomos a estudar se pode haver uma correlação entre o desenvolvimento do sistema nervoso e vascular de um germe dental transplantado com a posição que ele é implantado e/ou com a presença de tecido ósseo que no leito receptor. Ademais, buscamos saber se o estágio de desenvolvimento do germe dental a ser transplantado pode influenciar a formação de tecido nervoso e vascular na polpa dental ou não. Nossos resultados revelaram que o local do sítio do implante influencia diretamente o desenvolvimento dental e que isto é tempo dependente. A vascularização e a reinervação da polpa dental nos espécimes implantados nas tíbias é mais semelhante ao grupo controle que os implantados nos rins e isto independe da posição de implantação dental. Entretanto, a polpa dental dos germes implantados nos rins parece estar comumente mais sadia, conter mais odontoblastos viáveis e ser capaz de produzir tecidos mineralizados como a osteodentina. / Contemporary dentistry, even using modern techniques, still deal with missing teeth using metal implants coated by prosthetic crowns. However, there is a worldwide effort to develop a biotooth using in vitro techniques. In this way it is already possible to generate structures similar to teeth using recombination of odontogenic and non odontogenic cells in tissue engineering experiments. The transplant of the recombined cells into a host is a necessary and major step to obtain the biotooth. In this context, at the same time that the development of an appropriate sensorial and vascular system in the biotooh is required, there are many unclear questions about it. Therefore, herein we intend to analyze (I) whether may exist a correlation between the stage of development and vascular and nervous re-growth in the dental pulp after tooth transplantation; (II) if the absence or presence of bone could influence this processes or (III) if the position of implantation could change the vascular and/or nervous development in the transplanted tooth. Our results showed that the site of implantation directly alter tooth development modifying morphogenesis and expression of different vascular, perivascular and neural markers in a time dependant way. The re-growth of the vascular and neural tissue on samples transplanted to the tibia is more similar to the control group than the kidney ones and it is non dependant of the position of implantation. However, the pulp tissue of the samples transplanted under the kidney capsule seemed to be healthier as they were capable of producing mineralized tissue such as osteodentin and still had live odontoblasts.

No description available.

Eventos estressantes durante a infância promovem alterações comportamentais e encefálicas persistentes, aumentando a predisposição para transtornos psiquiátricos. A separação materna tem sido utilizada como modelo de estresse pós-natal. Animais submetidos à separação materna apresentam uma resposta exacerbada do eixo hipotálamo-hipófise-adrenal ao estresse. Ao contrário, estudos sugerem que o ambiente enriquecido, por aumentar a neurogênese no giro denteado do hipocampo, pode ter efeitos benéficos sobre doenças de distúrbio comportamental. No presente projeto questionamos se o enriquecimento ambiental interfere com as alterações plásticas promovidas pela separação materna no hipocampo de ratos jovens. Utilizamos imunofluorescência, microscopia confocal, microscopia eletrônica e qRT- PCR de amostras coletadas por microdissecção a laser. A separação materna reduziu a neurogênese hipocampal, bem como a expressão de mRNA para os genes Nr3c1, codificador de receptores glicocorticóides, e Htr1a, codificador de receptores serotoninérgicos (5TH-1A). O enriquecimento ambiental reduziu a expressão de Htr1a. Além disso, aumentou a proporção de sinapses sobre espinhos dendríticos, sugerindo maior plasticidade sináptica. O enriquecimento ambiental, nos animais previamente submetidos à separação materna, aumentou a sobrevivência celular e a expressão de Nr3c1, mas não a diferenciação neuronal hipocampal. As alterações promovidas pela separação materna parecem ser persistentes, mas podem ser parcialmente revertidas pelo enriquecimento do ambiente. / Stressful events during childhood promote persistent behavioral and brain changes, increasing the predisposition to psychiatric disorders. The maternal separation has been used as postnatal stress model. Animals subjected to maternal separation exhibit an exaggerated response of the hypothalamus-pituitary-adrenal axis to stress. Instead, studies suggest that environmental enrichment, by increasing neurogenesis in the dentate gyrus of the hippocampus, has beneficial effects on behavioral disorders. In this project, we discuss whether the environmental enrichment interferes with plastic changes promoted by maternal separation in the hippocampus of young rats. We used immunofluorescence, confocal microscopy, electron microscopy and qRT-PCR of samples collected by a laser microdissection system. The maternal separation reduced hippocampal neurogenesis, as well as the mRNA expression for the genes Nr3c1, that codify glycocorticoid receptors, and Htr1a, that codify serotonin receptors (5HT-1A). Environmental enrichment reduced the expression of Htr1a. Furthermore, increases the proportion of synapses on dendritic spines, suggesting greater synaptic plasticity. The environment enrichment of the animals subjected to maternal separation increased cell survival and the expression of Nr3c1 mRNA, but not the neuronal differentiation in the hippocampus. The changes promoted by maternal separation are persistent, however may be partially reversed by the environmental enrichment.

Les perturbateurs endocriniens (PE) sont une préoccupation majeure de par leurs effets sur les grandes fonctions physiologiques des organismes etparticulièrement sur la fonction de reproduction. Chez les vertébrés, la reproduction est sous le contrôle d’un axe neuroendocrinien nommé axegonadotrope. Celui-ci comprend différents acteurs cérébraux (gonadolibérines et kisspeptines), hypophysaires (gonadotropines) et gonadiques(stéroïdes sexuels) dont la mise en place et le fonctionnement sont régulés finement. Parmi les mécanismes de régulation, les stéroïdes vont jouer unrôle important en effectuant des boucles de rétrocontrôle. Les PE mimant ces stéroïdes sont donc potentiellement capables d’impacter ces régulations.L’objectif de ce travail était d’évaluer les effets d’un xénooestrogène sur les mises en place des acteurs cérébraux impliqués dans l’axe gonadotrope chezle bar européen (Dicentrarchus labrax). Nous avons d’abord décrit l’ontogénèse des systèmes à kisspeptine, à gonadolibérine et à gonadotropine aucours du développement larvaire. Nous avons mis en évidence une régulation différentielle de ces acteurs suggérant un rôle durant ce processusdéveloppemental. De plus, nous montrons que le 17α-éthinylestradiol (EE2) est capable de perturber l'ontogenèse des systèmes à GnRH2 et à GnRH3au cours des stades précoces du développement larvaire.Ces données constituent une base permettant l’évaluation des effets des PE sur l’ontogénèse des systèmes neuroendocriniens. / Endocrine disruptors (EDs) are a major concern because of their effects on the main physiological functions, particularly on the reproductive function.In vertebrates, reproduction is under the control of a neuroendocrine axis called hypothalamic-pituitary-gonadal (HPG) axis. It includes various actors atthe brain (gonadotropin releasing hormones (GnRH) and kisspeptins), pituitary (gonadotropic hormones) and gonadal (sex steroids) levels. Theirdévelopment and functioning are finely regulated. Among the regulators, steroids play an important role by performing feedback loops. EDs mimickingsteroids are therefore potentially able to impact these regulations. The objective of this work was to evaluate the effects of xenoestrogens on theimplementation of cerebral actors involved in the HPG axis in the European sea bass (Dicentrarchus labrax). We first described the ontogeny of thekisspeptin, GnRH and gonadotropin systems during larval development. We have highlighted a differential regulation of these actors suggesting a roleduring this developmental process. In addition, we show that 17α-ethinylestradiol (EE2) is able to disrupt the ontogeny of GnRH2 and GnRH3 systemsduring the early stages of larval development.These data provide a basis for evaluation of the EDs effects on the ontogeny of neuroendocrine systems.

La neurogenèse adulte est cruciale pour certaines fonctions mnésiques dépendantes de l'hippocampe. La mise en évidence d'une altération de la neurogenèse dans le cerveau de souris transgéniques modèles de la maladie d'Alzheimer (MA), en parallèle d'une réduction du contenu mitochondrial de leurs nouveaux neurones ouvre une nouvelle piste de recherche ciblant les mitochondries. Aujourd'hui, l'hypothèse d'un rôle causal des dysfonctionnements mitochondriaux dans l'étiologie des pathologies neurodégénératives est particulièrement pertinente dans la MA. Les mitochondries, " centrales électriques " et régulateurs du métabolisme oxydatif, forment un réseau dynamique qui s'adapte aux différents types et contextes cellulaires, via des événements antagonistes de fusion et de fission de leurs membranes. Les protéines clés ont été identifiées, dont OPA1 qui permet la fusion. Les dysfonctionnements de cette dynamique influent non seulement sur la forme et la distribution des mitochondries dans les neurones, mais affectent aussi leurs principales activités que sont respiration, régulation calcique, production de ROS et apoptose. Dans les neurones, cellules excitables à l'architecture complexe, les dysfonctionnements mitochondriaux ont des conséquences particulièrement cruciales pour la transmission synaptique. Au cours de cette thèse, nous avons étudié parallèlement des souris modèles de la MA, les souris Tg2576 (mutation d'APP) et des souris OPA1+/-, porteuses d'une mutation d'OPA1, modèles de l'Atrophie Optique Dominante. Nous avons observé chez ces deux lignées de souris une altération précoce des performances dans des tests comportementaux mettant en jeu le gyrus denté et les nouveaux neurones (tests de localisation d'objet et de séparation de patron). Nous avons démontré chez les souris Tg2576 et OPA1+/- que ces déficits cognitifs sont associés à des perturbations de la neurogenèse hippocampique adulte.[...] / Adult neurogenesis is crucial for some hippocampus-dependent memory functions. Both the demonstration of an alteration of neurogenesis in the brain of transgenic mouse models of Alzheimer's disease (AD), in parallel with a reduction in the mitochondrial content of their new neurons, open a new research avenue targeting the mitochondria. Today, the hypothesis of a causal role of mitochondrial dysfunctions in the etiology of neurodegenerative pathologies is particularly relevant in AD. Mitochondria, "power plants" and regulators of oxidative metabolism, form a dynamic network that adapts to different cell types and contexts, via antagonistic events of fusion and fission of their membranes. Key proteins have been identified, including OPA1 that allows fusion. Dysfunctions of this dynamics affect not only the shape and distribution of mitochondria in neurons, but also alter their main activities: respiration, calcium regulation, ROS production and apoptosis. In neurons, excitable cells with complex architecture, mitochondrial dysfunctions have particularly crucial consequences for synaptic transmission. In this thesis, we studied in parallel an AD mouse model, the Tg2576 mice (APP mutation) and the OPA1 +/- mice, carrying a mutation of OPA1, a Dominant Optic Atrophy model. In both mouse lines, we observed precocious performance alterations in behavioral tests involving the dentate gyrus and new neurons (object location, pattern separation tests). We demonstrated in Tg2576 and OPA1 +/- mice that these cognitive deficits are associated with disturbances of adult hippocampal neurogenesis. [...]

Alzheimer's disease (AD) is the most common neurodegenerative disease, characterized by progressive amyloid plaque aggregation, neurofibrillary tangles, and cortical tissue death. As the prevalence of AD is projected to climb in coming years, there is a vested interest in identifying endophenotypes by which to improve diagnostics and direct clinical interventions. The risk for complex disorders, such as AD, is influenced by multiple genetic, environmental, and lifestyle factors. Significant strides have been made in identifying genetic variants linked to AD through the genome-wide association study (GWAS). It has been estimated in more recent years, however, that GWAS-identified variants account for limited AD heritability, suggesting the role of non-sequence genetic mechanisms, such as epigenetic moderators. By influencing gene expression, epigenetic markers have been linked to age- associated decline through modulation of chromatin architecture and global genome instability, though such mechanisms are also involved with a number of normal biological processes, including neurogenesis. As the strategies of clinical genetics shift to include a heavier focus on epigenetic contributors, altered adult neurogenesis presents itself as a strong candidate for an endophenotype of AD development. This thesis proposes that, due to neuropathological dysfunction of epigenetic mechanisms in AD, new generations of neurons fail to proliferate, differentiate, and mature correctly, resulting in the larger loss of neurons and cognitive deficits characteristic to neurodegenerative disease. The plasticity of the epigenome and the role of epigenetic factors as mediators of the genome and the environment make such alterations attractive in AD research and implies the potential for therapeutic interventions. The present review submits neurogenesis as a viable target of epigenetic research in AD, highlights shared loci between neurogenesis and AD in the epigenome, and considers the promises and limitations of the neurogenic endophenotype.