Investigations into the neural correlates of memory have found the hippocampus to be a crucial structure for long-term declarative memories, but the exact nature of this contribution remains under debate. This paper covers three theories concerned with how the hippocampus is involved in long-term memory, namely the Standard Consolidation Model, the Multiple-Trace Theory, and the Distributed Reinstatement Theory. According to the Standard Consolidation Model, long-term declarative memories (both episodic and semantic) are dependent on the hippocampus for a limited time during which the memories undergo a process of consolidation, after which they become dependent on the neocortex. In contrast, the Multiple-Trace Theory argues that detailed and context-specific episodic (but not semantic) memories remain dependent on the hippocampus indefinitely. While both the aforementioned theories posit that memories are initially dependent on the hippocampus, the Distributed Reinstatement Theory does not. Advocates of this theory propose that several memory systems compete for the encoding of a memory, and that the hippocampus usually is the dominant system. However, it is also suggested that the other (unspecified) memory systems can overcome the hippocampal dominance through extensive and distributed learning sessions. In this paper, findings from both human and rodent studies focusing on the hippocampus are reviewed and used to evaluate the claims made by each theory on a systems level.

Chez la levure et les mammifères le complexe rétromère est impliqué dans différentes étapes de trafic intracellulaire qui modulent divers processus cellulaires et développementaux. Chez les mammifères, le rétromère se compose des protéines Vacuolar Protein Sorting (VPS) 35, VPS26, VPS29 et d’un dimere de Sorting Nexins (SNX). Les composants du rétromère sont conservés chez les plantes et sont localisés dans le même endosome de tri. Dans ce travail, nous avons étudié le mécanisme d’assemblage et de recrutement du complexe à la membrane et analysé le rôle du rétromère dans le développement d’Arabidopsis. Nos resultsts montrent que chez les plantes, contrairement aux mammifères, VPS35 recrute le sous-complexe VPS à la membrane, indépendamment des SNXs. Ceci nous a permis de proposer un modèle d’assemblage du rétromère très original. L’analyse de mutants perte de fonction pour le rétromère révèle que VPS26, VPS29 et VPS35 n’ont pas de fonctions indépendantes, mais agissent ensemble dans la régulation de processus développementaux. De plus, par un crible double hybride chez la levure, nous avons isolé un interacteur de VPS35 : Ethylene Insensitive 2 (EIN2). EIN2 est une protéine localisée au Réticulum Endoplasmique et impliquée dans la voie de signalisation d’une hormone végétale : l’éthylène. Nous montrons que des mutants perte de fonction pour le rétromère présentent des défauts dans la réponse à l’éthylène, indiquant un rôle du rétromère dans la perception de cette hormone par les plantes. Ces résultats, combinés avec nos données antérieures montrant que le rétromère est impliqué dans la voie de signalisation de l’auxine, révèlent un lien entre signalisation de l’auxine et de l’éthylènevia le complexe rétromère. / In yeast and mammals, the retromer complex mediates various steps of intracellular trafficking and regulates a variety of cellular and developmental processes. In mammals, it is composed of Vacuolar Protein Sorting (VPS) 35, VPS26, VPS29 and a dimer of Sorting Nexins (SNX). Retromer components are conserved in plants and we showed that they colocalize to the same sorting endosome. In this work, we have investigated the mechanism of assembly and recruitment of the retromer to the endosomal membrane and studied its function in Arabidopsis development. We report that, unlike animals, plant VPS35 recruits the other VPS retromer components to the membrane of endosomes, independently of SNXs. This data allowed us to propose an original model of assembly of the plant retromer complex. By analyzing a series of retromer loss-of-function mutants, we show that VPS26, VPS29 and VPS35 always act together in modulating developmental processes. To identify retromer partners, we carried out a Yeast-2-Hybrid (Y2H) screen using VPS35 as a bait. We found that VPS35 can bind, among other proteins, Ethylene Insensitive 2 (EIN2). EIN2 is an Endoplasmic Reticulum-located protein involved in the signaling pathway of a plant hormone: ethylene. We demonstrate that retromer mutants are affected in ethylene signaling, which indicates that the retromer complex participates in a proper perception of ethylene by plants. Combined with our previous data in which we showed that the retromer is involved in the auxin signalling pathway, our present work reveals a link between auxin and ethylene signaling through the retromer complex.

Orientador: Pedro Felício Estrada Bernabé / Banca: José Arlindo Otoboni Filho / Banca: Carlos Estrela / Banca: João Eduardo Gomes Filho / Banca: Rielson José Alves Cardoso / Resumo: O objetivo deste trabalho foi avaliar a capacidade seladora do MTA como material retrobturador, por via coronária, comparado ao cimento de óxido de zinco e eugenol consistente, assim como investigar os possíveis efeitos que esta via de contaminação provocaria nos tecidos periapicais de dentes de cães. Para tanto, foram utilizadas 48 raízes de 24 pré-molares superiores e inferiores de três cães adultos jovens. Constituíram-se 4 grupos experimentais: dentes com canais radiculares expostos ao meio bucal e a obturação retrógrada realizada com o cimento de óxido de zinco e eugenol consistente ou MTA e dentes com canais radiculares obturados e selados com guta-percha e amálgama e a obturação retrógrada realizada com o cimento de óxido de zinco e eugenol consistente ou MTA. Após 180 dias dos procedimentos cirúrgicos, os animais foram sacrificados, os maxilares removidos e fixados em solução de formol a 10%. Realizou-se então a descalcificação das peças e posterior processamento histológico, sendo os cortes corados pelas técnicas da hematoxilina e eosina e Brown & Brenn. A análise dos resultados permitiu concluir que o MTA, independentemente do tipo de tratamento executado, se canais obturados ou não, apresentou resultados mais favoráveis àqueles exibidos pelo OZE consistente (p=0,045). Os efeitos da infiltração marginal coronária são minimizados com o emprego de um material retrobturador eficiente do ponto de vista de selamento marginal, como é o MTA. Dos materiais retrobturadores utilizados, o único que estimulou a deposição de tecido cementário em íntimo contato com o material selador foi o MTA / Abstract: The aim of this study was to evaluate the sealing ability of MTA when used as root-end filling material to seal periapical tissues of the coronal microbial leakage, comparing it to the zinc oxid eugenol cement desiccated mixture. Forty-eight roots from 24 superior and inferior premolars of 3 young adult dogs were the sample. They were divided in four experimental groups. Two groups had the root canal exposed to the oral enviroment and the root-end cavities were filled with MTA or zinc oxid eugenol cement desiccated mixture. The other two groups (controls groups) had the root canal filled and the coronal acess was sealed with gutta-percha and amalgam and the root-end cavities also were filled with MTA or zinc oxid eugenol cement desiccated mixture. Animals were killed one hundred and eighty days after the treatment. The jaws were removed and fixed in 10% buffered formalin solution. After the block sections were demineralized, histological processing was done and the sections were stained with hematoxylin and eosin and Brown & Brenn techniques. Taking into consideration the experimental conditions under which this work was undertaken, as well as the results obtained, we can concluded that the MTA with or without root canal filling showed better results when compared to the zinc oxid eugenol cement desiccated mixture (p=0,045). The effect of coronal bacterial leakage can be minimized by using an efficient root-end filling material as MTA. From the root-end filling material investigated the MTA was the only one which stimulated the deposition of cement tissue in contact with the sealing material / Doutor

A sinalização retrógrada mitocondrial é uma via de comunicação entre a mitocôndria e o núcleo que regula a expressão de uma série de genes nucleares que codificam proteínas mitocondriais, em resposta a disfunções mitocondriais. Em Saccharomyces cerevisiae, a via depende de Rtg1p e Rtg3p, que juntos formam o fator de transcrição que regula a expressão gênica, e de Rtg2p, um ativador da via. Aqui, nós mostramos novos estudos direcionados à investigação do impacto da sinalização retrógrada RTG-dependente na fisiologia mitocondrial. Verificamos que mutantes incapazes de realizar sinalização retrógrada RTG-dependente apresentam consumo de oxigênio mais elevado e menor produção de peróxido de hidrogênio em fase estacionária quando comparados a células selvagens. Interessantemente, mutantes RTG são menos capazes de decompor peróxido de hidrogênio assim como manter-se viáveis quando desafiados com peróxido. Nossos resultados indicam que a sinalização por RTG está envolvida na indução hormética de defesas antioxidantes e de resistência a estresse, função ainda não descrita para este sistema. / Mitochondrial retrograde signaling is a communication pathway between the mitochondrion and the nucleus which regulates the expression of a subset of nuclear genes that codify mitochondrial proteins, mediating cell response to mitochondrial dysfunction. In Saccharomyces cerevisiae, the pathway depends on Rtg1p and Rtg3p, which together form the transcription factor that regulates gene expression, and Rtg2p, an activator of the pathway. Here, we provide novel studies aimed at assessing the functional impact of the lack of RTG-dependent signaling on mitochondrial activity. We show that mutants defective in RTG-dependent retrograde signaling present higher oxygen consumption and reduced hydrogen peroxide release in the stationary phase when compared to wild type cells. Interestingly, RTG mutants are less able to decompose hydrogen peroxide as well as maintain viability when challenged with hydrogen peroxide. Overall, our results indicate that RTG signaling is involved in the hormetic induction of antioxidant defenses and stress resistance, a function of this system not yet described.

Rtg2p é uma proteína que participa da sinalização retrógrada, uma via de comunicação da mitocôndria para o núcleo; também tem sido associada com a longevidade em S. cerevisiae. O objetivo deste trabalho foi identificar os determinantes estruturais de Rtg2p, envolvidos na sinalização retrógrada e no envelhecimento. Para isto foram produzidos treze mutantes pontuais a partir do desenho racional por decomposição de redes de correlação de aminoácidos (DRCN). Analisaram-se as cepas mutantes por ensaio de auxotrofia para glutamato, expressão do gene CIT2 e ensaio de longevidade replicativa. Em sua grande maioria as mutações realizadas causaram perturbações nas funções de Rtg2p, com destaque para as cepas E106A, R109E, E137A, T138A e D158A, que apresentaram longevidade igual à da cepa rtg2Δ, com apenas uma mutação pontual. Em conclusão, os resultados obtidos demonstram que o domínio N-terminal é muito importante para a função de Rtg2p, e indicam que existem determinantes estruturais que controlam a longevidade de forma dependente ou independente da resposta retrógrada. / Rtg2p is a protein involved in the retrograde signaling, a pathway of communcation from mitochondria to nucleus; also has been associated with longevity in S. cerevisiae. The goal of this study was to identify the structural determinants of Rtg2p, controlling the function of this protein in retrograde response and aging. For this purpose thirteen point mutants were produced by site-directed mutagenesis, using rational design by decomposition of residues correlation networks (DRCN). The strains was analyzed by glutamate auxotrophy, CIT2 gene expression and replicative life span assays. For the most of performed mutations, generated inactivation to Rtg2p functions, highlighting to R109E, E137A, T138A, and D158A showed longevity equal to rtg2Δ strain, even with a single amino acid change. In conclusion, our results demonstrate that the N-terminal domain is very important to the function of Rtg2p and also show there are structural determinants in Rtg2p that control longevity in both dependent or independent manner of the communication between mitochondria and nucleus.

京都大学 / 0048 / 新制・論文博士 / 博士(医学) / 乙第13363号 / 論医博第2205号 / 新制||医||1045(附属図書館) / (主査)教授 妹尾 浩, 教授 今中 雄一, 教授 川上 浩司 / 学位規則第4条第2項該当 / Doctor of Medical Science / Kyoto University / DFAM

The digastric muscle of the rabbit consists of a single anterior belly which inserts onto the lower jaw. Horseradish peroxidase was injected into the muscle and into subcutaneous regions overlying the lower jaw to determine the sites of origin of the motor innervation to both the digastric muscle and the platysma muscles. After digastric muscle injection, labelled cells were found in the ipsilateral retrotrigeminal nucleus as well as in the intermediate subnucleus of the main facial nucleus on both sides. Subcutaneous injections produced labelling which was found bilaterally in the intermediate subnucleus and in the ventromedial portion of the medial subnucleus. These results are interpreted in relation to the common embryological origin of these two muscles and their innervation.

Introduction: Mini percutaneous nephrolithotomy (mini-PCNL) and retrograde intrarenal surgery (RIRS) are well-established techniques with little morbidity. The combined use of standard PCNL and the mini-PCNL or the RIRS technique was evaluated and compared to investigate their own role in the management of staghorn calculi in solitary kidney. Materials and Methods: 23 patients received combined standard PCNL and mini-PCNL (group 1), and 22 patients received combined standard PCNL and RIRS (group 2). The treatment results and complications were evaluated and compared. Results: The mean operation time was 128.8 ± 9.1 min in group 1 and 109.8 ± 10.7 min in group 2 (p < 0.001). The decrease in hemoglobin level in group 1 was significantly higher than that in group 2 (3.5 ± 0.6 vs. 2.1 ± 0.5 g/dl, p < 0.001). The final stone-free rate was significantly higher (p = 0.038) in group 2 (90.9%) than in group 1 (65.2%). Conclusions: Combined standard PCNL and RIRS technique can extract the majority of staghorn calculi quickly by PCNL with EMS Lithoclast, and RIRS used simultaneously can reduce the need for multiple tracts and therefore reduce blood loss and potential morbidity related to multiple tracts, shorten the operation time and achieve a high stone-free rate.

The vagal postganglionic control of cardiac rate is mediated by two intracardiac ganglia, i.e., the sinoatrial (SA) and posterior atrial (PA) ganglia. Nothing is known about the vagal preganglionic neurons (VPNs) that innervate the PA ganglion or about the neurochemical anatomy of central afferents that innervate these VPNs. These issues were examined using light microscopic retrograde labeling methods and dual-labeling electron microscopic histochemical and immunocytochemical methods. VPNs projecting to the PA ganglion are found in a narrow column exclusively in the ventrolateral nucleus ambiguus (NA-VL). These neurons are relatively large (37.6 ± 2.7 μm by 21.3 ± 3.4 μm) with abundant cytoplasm and intracellular organelles, rare somatic and dendritic spines, round uninvaginated nuclei, and myelinated axons. Previous physiological data indicated that microinjections of neuropeptide Y (NPY) into the NA-VL cause negative chronotropic effects. The present morphological data demonstrate that NPY-immunoreactive nerve terminals formed 18 ± 4% of the axodendritic or axosomatic synapses and close appositions on VPNs projecting to the PA ganglion. Three approximately equal populations of VPNs in the NA-VL were retrogradely labeled from the SA and PA ganglia. One population each projects to the SA ganglion, the PA ganglion, or to both the SA and PA ganglia. Therefore, there are both shared and independent pathways involved in the vagal preganglionic controls of cardiac rate. These data are consistent with the hypothesis that the central and peripheral parasympathetic controls of cardiac rate are coordinated by multiple potentially redundant and/or interacting pathways and mechanisms.

Intracardiac pathways mediating the parasympathetic control of various cardiac functions are incompletely understood. Several intracardiac ganglia have been demonstrated to potently influence cardiac rate [the sinoatrial (SA) ganglion], atrioventricular (AV) conduction (the AV ganglion), or left ventricular contractility (the cranioventricular ganglion). However, there are numerous ganglia found throughout the heart whose functions are poorly characterized. One such ganglion, the posterior atrial (PA) ganglion, is found in a fat pad on the rostral dorsal surface of the right atrium. We have investigated the potential impact of this ganglion on cardiac rate and AV conduction. We report that microinjections of a ganglionic blocker into the PA ganglion significantly attenuates the negative chronotropic effects of vagal stimulation without significantly influencing negative dromotropic effects. Because prior evidence indicates that the PA ganglion does not project to the SA node, we neuroanatomically tested the hypothesis that the PA ganglion mediates its effect on cardiac rate through an interganglionic projection to the SA ganglion. Subsequent to micro-injections of the retrograde tracer fast blue into the SA ganglion, >70% of the retrogradely labeled neurons found within five intracardiac ganglia throughout the heart were observed in the PA ganglion. The neuroanatomic data further indicate that intraganglionic neuronal circuits are found within the SA ganglion. The present data support the hypothesis that two interacting cardiac centers, i.e., the SA and PA ganglia, mediate the peripheral parasympathetic control of cardiac rate. These data further support the emerging concept of an intrinsic cardiac nervous system.