No description available.

Correlative light and electron microscopy (CLEM) is a powerful approach to investigate the molecular ultrastructure of labeled cell compartments. However, quantitative CLEM studies are rare, mainly due to small sample sizes and the sensitivity of fluorescent proteins to strong fixatives and contrasting reagents for EM. Here, we show that fusion of a self-labeling protein to insulin allows for the quantification of age-distinct insulin granule pools in pancreatic beta cells by a combination of super resolution and transmission electron microscopy on Tokuyasu cryosections. In contrast to fluorescent proteins like GFP organic dyes covalently bound to self-labeling proteins retain their fluorescence also in epoxy resin following high pressure freezing and freeze substitution, or remarkably even after strong chemical fixation. This enables for the assessment of age-defined granule morphology and degradation. Finally, we demonstrate that this CLEM protocol is highly versatile, being suitable for single and dual fluorescent labeling and detection of different proteins with optimal ultrastructure preservation and contrast.

Thèse numérisée par la Division de la gestion de documents et des archives de l'Université de Montréal.

Plus d’un millier d’exoplanètes ont été découvertes depuis une dizaine d’années. Plus incroyable encore, nous pouvons maintenant caractériser les atmosphères de ces mondes lointains. Des spectres de Jupiter-chauds tels que HD 189733b et HD 209458b et de planètes similaires à Neptune telles que GJ1214b sont déjà disponibles et ceux de planètes plus petites le seront bientôt. La plupart des observations caractérisent l’état moyen de l’atmosphère. Pour les cas les plus favorables, l’observation des courbes de phase et la technique de cartographie par éclipse secondaire permettent d’obtenir une résolution en longitude et en latitude. Les planètes les plus proches de leurs étoiles sont aussi les plus faciles à observer. Ces mondes chauds sont radicalement différents des exemples que nous avons dans le système solaire. Modéliser correctement leurs atmosphères est un défi à relever pour comprendre les observations présentes et à venir. Durant cette thèse, j’ai développé des modèles de différente complexité pour comprendre les interactions entre la structure thermique, la composition, la circulation atmosphérique et l’évolution à long terme des exoplanètes irradiées. La forte luminosité de leur étoile hôte détermine le climat de ces planètes. Elle engendre une circulation atmosphérique qui maintient l’atmosphère dans un état de déséquilibre thermique et chimique, affectant son évolution. Avec les futurs instruments de nombreuses autres planètes vont être découvertes et caractérisées. Nos modèles seront testés sur une large diversité de planètes, ouvrant les portes de la climatologie aux exoplanètes. / More than a thousand exoplanets have been discovered over the last decade. Perhaps more excitingly, probing their atmospheres has become possible. We now have spectra of hot Jupiters like HD 189733b and HD 209458b, of Neptune-like planets like GJ1214b and even smaller planets are within reach. Most exoplanet atmospheric observations are averaged spatially, often over a hemi- sphere (during secondary eclipse) or over the limb of the planet (during transit). For favorable targets, longitudinal and latitudinal resolution can also be obtained with phase curve and secondary eclipse mapping techniques respectively. The closer the planet orbits to its star, the easier it is to observe. These hot planets strongly differ from the examples we have in our solar-system. Proper models of their atmospheres are challenging yet necessary to understand current and future observations. In this thesis, I use a hierarchy of atmospheric models to understand the interactions between the thermal structure, the composition, the atmospheric circulation and the long-term evolution of irradiated planets. In these planets, the large stellar irradiation dominates the energy budget of the atmosphere. It powers a strong atmospheric circulation that transports heat and material around the planet, driving the atmosphere out of thermal and chemical equilibrium and affecting its long-term evolution. Future instruments (Gaia, SPIRou, CHEOPS, TESS, PLATO etc) will discover many more planets that the next generation of telescopes (GMT, TMT, E-ELT or JWST) will characterize with an unprecedented accuracy. Models will be tested on a large sample of planets, extending the study of climates to exoplanets.

O trabalho tem como objetivo central a análise de filmes feitos em Super-8 durante a década de 1970, pensando as suas inserções e vínculos com a grande vaga libertária da contracultura, o Tropicalismo, a poesia, o teatro, o happening e o desbunde, no quadro da ditadura civil-militar no Brasil: Terror da Vermelha (1972), de Torquato Neto; Céu sobre Água (1978), de José Agrippino de Paula e Maria Esther Stockler; O Rei do Cagaço (1977) e Exposed (1978), de Edgard Navarro. Priorizou-se o agenciamento do corpo na relação da câmera superoitista com o entorno imediato, em vista da compreensão de como foram tecidas as poéticas desses filmes. Procurou-se mostrar o modo particular e radical com que os realizadores tomaram esse \"brinquedo\" num âmbito de não separação entre vida e obra, que impregnava suas trajetórias tanto estéticas como existenciais. A contracultura deixou marcas indeléveis na subjetividade e no comportamento daqueles que souberam experimentar novas formas de vida. Micropoliticamente, ela inoculou a ideia de um corpo coletivo na sexualidade, na sociabilidade, influenciando as artes, a literatura, a poesia e o cinema, e teve na possibilidade da portabilidade da câmera superoitista um dos grandes exemplos de que o corpo era o centro dessa profunda transformação que se processava no sentido de uma recusa dos padrões socioculturais pré-estabelecidos. Se o AI-5 bloqueou o desejo brasileiro de transformação social e macropolítica, podia-se agora, em contrapartida, dar vazão ao corpo e à loucura antes que o lobo chegasse. / The main objective of this work is the analysis of films made in Super-8 during the 1970s, thinking about their insertions and links with the great libertarian wave of the counterculture, tropicalism, poetry, theater, happening and fading, in the framework of the civil-military dictatorship in Brazil: Terror of the Red (1972), Torquato Neto, Sky over Water (1978), José Agrippino de Paula and Maria Esther Stockler, The King of Cagaço (1977) and Exposed (1978) scored by Edgard Navarro. The agency of the body was prioritized in the relation of the superoitist camera to the immediate surroundings, in view of the understanding of how the poetics of these films were woven. It sought to show the particular and radical way in which the filmmakers took this \"toy\" in a framework of non-separation between life and work, which permeated both their aesthetic and existential trajectories. The counterculture left indelible marks on the subjectivity and behavior of those who knew how to try new ways of life. Micropolitically, she inoculated the idea of a collective body in sexuality, in sociability, influencing the arts, literature, poetry and cinema had in the portability possibility of the superoitist camera one of the great examples that the body was the center of this deep transformation which was proceeding towards a refusal of pre-established socio-cultural standards. If AI-5 definitively blocked the desire for social and macro-political transformation, one could now, on the other hand, give vent to body and madness before the wolf arrived.

Le cerveau est composé de neurones et de cellules gliales qui jouent un rôle de soutien et de protection du réseau cellulaire. L’espace extra-cellulaire (ECS) correspond à l’espace qui existe entre ces cellules. Les modifications de sa structure peuvent dépendre de plusieurs paramètres comme l’âge, l’apprentissage ou les maladies neuro-dégénératives. Le volume de l’ECS correspond à environ 20$%$ du volume total du cerveau et les neurotransmetteurs et autres molécules circulent dans cet espace pour assurer une communication neuronale optimale. Cependant, les dimensions et la viscosité locale de cet espace restent encore mal-connues. L’ECS est composé entre autres de protéoglycans, de glycoaminoglycans (acide hyaluronique…) et de fluide cérébrospinal. Nous avons proposé dans cette thèse une stratégie pour mesurer les dimensions et les propriétés rhéologiques de l’espace extra-cellulaire de tranches de cerveaux de rats maintenue en vie à l’aide du suivi de nanotubes de carbone individuels luminescents. Pour ces applications, nous avons étudier la biocompatibilité et le rapport signal sur bruit de nos échantillons de nanotubes afin de les détecter en profondeur dans les tranches de cerveaux et de pouvoir mesurer leurs propriétés de diffusion. / The brain is mainly composed of neurons which ensure neuronal communication and glialcells which play a role in supporting and protecting the neural network. The extracellular space corresponds to the space that exists between all these cells and represents around 20 %of the whole brain volume. In this space, neurotransmitters and other molecules circulate into ensure optimal neuronal functioning and communication. Its complex organization whichis important to ensure proper functioning of the brain changes during aging, learning or neurodegenerative diseases. However, its local dimensions and viscosity are still poorly known.To understand these key parameters, in this thesis, we developed a strategy based on the tracking of single luminescent carbon nanotubes. We applied this strategy to measure the structural and viscous properties of the extracellular space of living rodent brains slices at the nanoscale. The organization of the manuscript is as follows. After an introduction of the photoluminescence properties of carbon nanotubes, we present the study that allowed us to select the optimal nanotube encapsulation protocol to achieve our biological applications. We also present a quantitative study describing the temperature increase of the sample when laser irradiations at different wavelengths are used to detect single nanotubes in a brain slice.Thanks to a fine analysis of the singular diffusion properties of carbon nanotubes in complex environments, we then present the strategy set up to reconstruct super-resolved maps (i.e. with resolution below the diffraction limit) of the brain extracellular space morphology.We also show that two local properties of this space can be extracted : a structural complexity parameter (tortuosity) and the fluid’s in situ viscosity seen by the nanotubes. This led us to propose a methodology allowing to model the viscosity in situ that would be seen, not by the nanotubes,but by any molecule of arbitrary sizes to simulate those intrinsically present or administered in the brain for pharmacological treatments. Finally, we present a strategy to make luminescent ultra-short carbon nanotubes that are not intrinsically luminescent and whose use could be a complementary approach to measure the local viscosity of the extracellular space of the brain.

Les vésicules synaptiques (VSs) sont essentielles pour la neurotransmission. Les recherches actuelles se focalisent sur la caractérisation de leur contenu en neurotransmetteurs, leur cinétique de libération, leur distribution et leur mobilité. Les VS ne sont pas présentes exclusievement en paquet dans les boutons présynaptiques mais sont echangées de façon dynamique avec le reste de l’axone dans un super-contingent (super-pool). Notre laboratoire a précédement montré que le transporteur vésiculaire de glutamate de type 1 (VGLUT1) jouerait un rôle dans la régulation du super-pool. Mon projet de thèse se focalise sur la mobilité des VS dans les axones. En premier lieu, j’ai généré une souris gain de fonction VGLUT1mEos2 afin d'étudier la mobilité des VSs et de mieux caractériser le super-pool. Ensuite j’ai engagé une étude des relation entre la structure de VGLUT1 et ses fonctions afin d’identifier les signatures moléculaires responsable de la régulation de la taille du super-pool. J’ai identifié le second motif poly-proline à l’extremité C-terminale de VGLUT1 comme étant nécessaire et suffisante pour induire une diminution de la taille du super-pool des VSs. Pour conclure mes travaux de thèse ont contribué à la compréhension du rôle de VGLUT1 dans la régulation de la mobilité des VSs et à fournir les outils nécessaires pour de futures investigations concernant la physiologie du super-pool. / Synaptic vesicles (SVs) are essential for neurotransmission, and more efforts are needed for better understanding their neurotransmitter content, release kinetics, distribution and mobility. SVs are not only clustered in presynaptic boutons, but also dynamically shared among multiple en passant presynaptic boutons, a phenomenon named SV super‐pool. Previous work from our laboratory suggested that the Vesicular GLUtamate Transporter 1 (VGLUT1) may play a role in regulating SV super-­pool size beyond loading glutamate into SV. My Ph.D project is focused on SVs mobility in axons. Firstly, I generated a VGLUT1mEos2 knock-in (KI) mouse line, which provides extended possibilities to study the SV trafficking and characterize SV super‐pool. Secondly, I engaged in a thorough VGLUT1 structure‐function analysis. I identified that VGLUT1 tends to cluster SVs in the presynaptic boutons and reduce SVs exchange with the super‐pool via the second poly‐proline motif of its C­‐terminus. Overall, my Ph.D work contributes to the knowledge of the role of VGLUT1 in regulating SVs mobility and provides new tools for the further investigations on SV super-­pool physiology.

Cone Beam Computed Tomography (CBCT) is a medical imaging technique routinely employed for diagnosis and treatment of patients with cranio-maxillo-facial defects. CBCT 3D reconstruction and segmentation of bones such as mandible or maxilla are essential procedures in orthodontic treatments. However, CBCT images present characteristics that are not desirable for processing, including low contrast, inhomogeneity, noise, and artifacts. Besides, values assigned to voxels are relative Hounsfield Units (HU), unlike traditional Computed Tomography (CT). Such drawbacks render CBCT segmentation a difficult and time-consuming task, usually performed manually with tools designed for medical image processing. We introduce two interactive two-stage methods for 3D segmentation of CBCT data: i) we first reduce the CBCT image resolution by grouping similar voxels into super-voxels defining a graph representation; ii) next, seeds placed by users guide graph clustering algorithms, splitting the bones into mandible and skull. We have evaluated our segmentation methods intensively by comparing the results against ground truth data of the mandible and the skull, in various scenarios. Results show that our methods produce accurate segmentation and are robust to changes in parameter settings. We also compared our approach with a similar segmentation strategy and we showed that it produces more accurate segmentation of the mandible and skull. In addition, we have evaluated our proposal with CT data of patients with deformed or missing bones. We obtained more accurate segmentation in all cases. As for the efficiency of our implementation, a segmentation of a typical CBCT image of the human head takes about five minutes. Finally, we carried out a usability test with orthodontists. Results have shown that our proposal not only produces accurate segmentation, as it also delivers an effortless and intuitive user interaction. / Tomografia Computadorizada de Feixe Cônico (TCFC) é uma modalidade para obtenção de imagens médicas 3D do crânio usada para diagnóstico e tratamento de pacientes com defeitos crânio-maxilo-faciais. A segmentação tridimensional de ossos como a mandíbula e a maxila são procedimentos essências em tratamentos ortodônticos. No entanto, a TCFC apresenta características não desejáveis para processamento digital como, por exemplo, baixo contraste, inomogeneidade, ruído e artefatos. Além disso, os valores atribuídos aos voxels são unidades de Hounsfield (HU) relativas, diferentemente da Tomografia Computadorizada (TC) tradicional. Esses inconvenientes tornam a segmentação de TCFC uma tarefa difícil e demorada, a qual é normalmente realizada por meio de ferramentas desenvolvidas para processamento digital de imagens médicas. Esta tese introduz dois métodos interativos para a segmentação 3D de TCFC, os quais são divididos em duas etapas: i) redução da resolução da TCFC por meio da agrupamento de voxels em super-voxels, seguida da criação de um grafo no qual os vértices são super-voxels; ii) posicionamento de sementes pelo usuário e segmentação por algoritmos de agrupamento em grafos, o que permite separar os ossos rotulados. Os métodos foram intensamente avaliados por meio da comparação dos resultados com padrão ouro da mandíbula e do crânio, considerando diversos cenários. Os resultados mostraram que os métodos não apenas produzem segmentações precisas, como também são robustos a mudanças nos parâmetros. Foi ainda realizada uma comparação com um trabalho relacionado, gerando melhores resultados tanto na segmentação da mandíbula quanto a do crânio. Além disso, foram avaliadas TCs de pacientes com ossos faltantes e quebrados. A segmentação de uma TCFC é realizada em cerca de 5 minutos. Por fim, foram realizados testes com usuarios ortodontistas. Os resultados mostraram que nossa proposta não apenas produz segmentações precisas, como também é de fácil interação.

Apresentamos uma teoria para obtenção de seções de choque de espalhamento inelástico de luz via mecanismos de flutuações de densidade de carga e spin em um gás de elétrons não-uniforme formado por um sistema multi-camadas de GaAs periodicamente &#948-dopadas com concentração eletrônica relativamente alta. Os cálculos, onde estão inclusos efeitos da interação coulombiana entre os portadores, efeitos de correlação e troca dinâmicos e o acoplamento com fônons LO, foram efetuados em condições de extrema ressonância com o gap de split-off do GaAs. Em tais condições, a estrutura detalhada dos níveis de energia dos buracos de spin-split torna-se extremamente importante e é fundamental para o surgimento do espectro de partícula independente apresentado nos espectros polarizados. Este comportamento é revelado através da seção de choque de espalhamento que consiste da parte imaginária de uma função resposta constituída de um termo de caráter de partícula independente e outro de caráter coletivo. De forma a levar em conta o amortecimento das flutuações de densidade, propõe-se uma função espectral baseada na conservação da corrente local. Comparação com formas de linha experimentais disponíveis para o caso de espectros despolarizados mostram excelente concordância. No caso dos espectros polarizados a concordância se deu em nível semi-quantitativo, já que excitações de caráter coletivo obtidas experimentalmente apresentaram intensidade menor do que aquelas fornecidas pela teoria. Tal discrepância é atribuída a efeitos de desordem introduzidas no processo de dopagem e que implicam na quebra das regras de conservação de momentum. / We present a theory for the inelastic light scattering cross-section for the mechanisms of charge and spin-density fluctuations in the relatively high concentration of the non-uniform electron gas of a multi-layered &#948-doped GaAs system. The calculations are done in conditions of extreme resonance with the spin-split edge of GaAs and include the effects of Coulomb interactions between the carriers, dynamical exchange-correlations and coupling with LO phonons. In such conditions, the detailed energy level structure of the spin-split holes becomes extremely important and is responsible by the single-particle behavior presented in the polarized spectrum. This behavior revealed by the scattering cross-section derived from the imaginary part of a response function, consists of a term showing single particle character and another displaying collective character. To include the damping of the density fluctuations, a spectral function is proposed based on the foreknowledge that the local current must be conserved. Comparison with the available experimental line-shapes for the depolarized spectra show excellent agreement. In the case of polarized spectra the agreement was given in semi-quantitative terms as experimental collective excitations present less intensity than those calculated by the theory. Such difference is attributed to disorder effects produced during the doping process which results in break down of momentum conservation rules.

Este estudo pretende investigar a pluralidade observada no trabalho do artista contemporâneo japonês Takashi Murakami (1962-), discutindo como a sua atuação em diversas áreas está diretamente ligada ao seu desenvolvimento artístico e fazer poético. No primeiro capítulo, construiremos um panorama sobre o mercado de arte, apresentando alguns dos principais agentes e eventos que o constituem e o movimentam. A compreensão deste cenário possibilita adentrarmos com mais clareza sobre as variadas ações de Murakami durante o segundo capítulo. Tomando o conceito de artista-etc do artista e crítico brasileiro Ricardo Basbaum (1961-) como ponto de partida, começaremos a perceber que parcerias em moda e design, assim como trabalho de galerista, colecionador e empreendedor, são papéis que Murakami desempenha em busca de respostas ligadas à sua indagação sobre a arte. A complexidade do artista também é vista no seu desenvolvimento estético, que constitui o terceiro capítulo deste texto. Por meio do mapeamento de suas obras realizadas e artistas que o instigaram desde a sua formação na faculdade, encontraremos um repertório diverso, iniciado com a prática de nihonga. A insatisfação com essa linguagem levou Murakami por uma série de experimentações, antes de desenvolver o super flat conceito pelo qual ele é reconhecido no âmbito mundial. A projeção internacional conquistada proporcionou ao artista seguir expandindo a sua pesquisa, por meio da sua parceria com o professor e historiador de arte Nobuo Tsuji (1932-) na qual trabalha o resgate de alguns nomes da arte japonesa ainda pouco estudados fora (e até mesmo dentro) do Japão. Espera-se que, a partir da leitura sobre Murakami apresentada neste trabalho, possa-se contribuir para as discussões sobre arte contemporânea e japonesa. / This study aims to investigate the plurality observed in the work of contemporary Japanese artist Takashi Murakami (1962-), discussing how his work in various areas is directly linked to his artistic and to make poetic development. In the first chapter, we will construct a panorama on the art market, presenting some of the main agents and events that constitute and move it. Understanding this scenario makes it possible to delve more clearly into Murakami\'s various actions during the second chapter. Taking the \"artista-etc\" [artist-etc] concept of the Brazilian artist and critic Ricardo Basbaum (1961-) as a starting point, we will begin to realize that partnerships in fashion and design, as well as the work of gallerist, collector and entrepreneur, are roles that Murakami plays in search of answers linked to his inquiry about art. The complexity of the artist is also seen in his aesthetic development, which is the third chapter of this text. Through the mapping of his accomplished works and artists who have instigated him since his formation in college, we will find a diverse repertoire, started with the practice of nihonga. The dissatisfaction with this language led Murakami through a series of experiments, before developing the super flat concept by which he is recognized worldwide. The international projection gained allowed the artist to continue to expand his research, through his partnership with Professor and Art Historian Nobuo Tsuji (1932-) in which he works to rescue some Japanese names of Japanese art that have yet been little-studied outside (and even within) Japan. It is hoped that, from the reading about Murakami presented in this work, one can contribute to the discussions on contemporary and Japanese art.