1,790 research outputs found
Tradução de uma disputa : Christophe versus Pétion em La tragédie du roi Christophe, de Aimé Césaire
No full textEsta contribuição propõe a tradução comentada da primeira cena da peça de teatro La Tragédie du roi Christophe (A tragédia do rei Christophe), de Aimé Césaire. O texto foi publicado pelo autor martinicano em 1963 e encenado a partir de 1964. A história aborda o embate histórico entre os dois líderes revolucionários haitianos, Alexandre Pétion (1770-1818) e Henry Christophe (1767-1820), após o estabelecimento da independência do Haiti, em 1804. O ex-escravizado Henry Christophe I, autoproclamado rei do Haiti em 1811, protagoniza, na peça, os impasses políticos decorrentes do processo de descolonização para a constituição de um novo estado haitiano livre e democrático. Optamos por traduzir a cena que anuncia e ilustra, na abertura do primeiro Ato, o caráter trágico que permeia o desenrolar dos eventos.This contribution proposes the commented translation of the first scene of the play La Tragédie du roi Christophe (The tragedy of King Christophe), by Aimé Césaire. The text was published by the Martinican author in 1963 and staged from 1964 onwards. The story addresses the historical clash between the two Haitian revolutionary leaders, Alexandre Pétion (1770-1818) and Henry Christophe (1767-1820), after the establishment of Haitian independence in 1804. The ex-enslaved Henry Christophe I, self-proclaimed King of Haiti in 1811, stars, in the play, political impasses resulting from the decolonization process, for the constitution of a new free and democratic Haitian state. We chose to translate the scene that announces and illustrates, in the opening of the first Act, the tragic character that permeates the unfolding of events
Analyse du fonctionnement synaptique du microcircuit de CA3 in vivo en utilisant des outils optogénétiques
No full textL'hippocampe est une région du cerveau située dans le lobe temporal médian. Avec d'autres structures limbiques, l'hippocampe est impliqué dans des processus d'apprentissage et de mémorisation et possède un rôle crucial dans le traitement spatial de l'information. Les synapses de l'hippocampe formées entre les fibres moussues (fm) originaires du gyrus denté et les neurones pyramidaux de CA3 ont reçu une attention particulière, compte tenu de la position stratégique occupée par le gyrus denté à l'entrée de l'hippocampe. En outre les synapses fm- CA3 sont distinctes de la plupart des autres synapses excitatrices du système nerveux central par leurs propriétés morphologiques et physiologiques uniques. Cela soulève la question de savoir si ces propriétés uniques reflètent aussi un rôle fonctionnel unique dans le traitement de l'information effectué par cette synapse au sein du microcircuit de l'hippocampe. Malheureusement nous ne savons que peu de choses sur la façon dont les cellules granulaires modulent l'activité des neurones de CA3 dans le réseau intact in vivo (Henze et al, 2002 ; Hagena et Manahan - Vaughan, 2010, 2011). Le manque d'information est dû au fait que la manipulation classique des circuits neuronaux par des approches électriques, pharmacologiques et génétiques manque de précisions spatiale et temporelle in vivo. L'utilisation de la stimulation extracellulaire de fibres moussues peut conduire à l'activation polysynaptique de cellules pyramidales de CA3, qui peuvent ensuite contaminer les réponses enregistrées. Par ailleurs, l'utilisation de critères trop conservateurs peut conduire à l'exclusion des réponses provenant des fibres moussues «purs» aux propriétés méconnues (Henze et al., 2000). Toutefois, le développement récent et rapide de l’optogénétique dans les neurosciences a fourni de nouveaux outils offrant une sélectivité spatiale élevée (activation optique spécifique de la cellule), et une grande précision temporelle (à l'échelle de la milliseconde), permettant la dissection et l'étude des circuits neuronaux in vivo. L'objectif de ma thèse était de mieux comprendre les mécanismes et les conséquences physiologiques de la plasticité synaptique à court terme se produisant à la synapse formée entre les fibres moussues et les neurones pyramidaux de CA3 dans le cerveau de souris intact. La présente thèse se compose de deux parties principales. Dans la première partie, j'ai exploré de nouveaux outils optogénétiques dans le but de contrôler l'activité des cellules granulaires à l’aide d’impulsions de lumière. La stimulation optogénétique repose sur l'activation du canal ionique channelrhodopsin - 2 - lumière fermée ( ChR2 ) par une lumière bleue et induit des potentiels d'action sur une large gamme de fréquences de stimulation. J'ai aussi observé que la stimulation optique peut être utilisée pour déclencher la plasticité à court terme au niveau des synapses fm-CA3.Dans la deuxième partie j'ai affiné la méthodologie de stimulation optogénétique in vivo pour la caractérisation non invasive du fonctionnement synaptique des synapses fm- CA3. La fiabilité de la stimulation optogénétique d'une population neuronale génétiquement ciblée ainsi que la résolution d'une seule cellule obtenue en utilisant des enregistrements de cellules entières sont des étapes importantes vers une meilleure compréhension du rôle fonctionnel des fibres moussues dans le réseau de l'hippocampe in vivo.The hippocampus is a brain region located in the medial temporal lobe. Along with other limbic structures, the hippocampus is involved in learning and memory processes and has a crucial role in spatial information processing. Within the hippocampus synapses made between mossy fibers (mf) originating from the dentate gyrus and CA3 pyramidal neurons have received particular attention, given the strategic position occupied by the dentate gyrus at the entrance of the hippocampus. Moreover mf-CA3 synapses are distinct from most of other excitatory synapses in the central nervous system for their unusual morphological and physiological properties. This raises the question if these unique properties reflect a unique functional role in information processing carried out by this synapse within the microcircuit of the hippocampus. Unfortunately very little is known on how granule cells modulate the activity of CA3 neurons in the intact network in vivo (Henze et al., 2002; Hagena and Manahan-Vaughan, 2010, 2011). The paucity of information is due to the fact that classical manipulation of neuronal circuits using electrical, pharmacological and genetic approaches lack spatial and temporal precision in vivo. The use of bulk extracellular stimulation may lead to polysynaptic activation of CA3 pyramidal cells, which can subsequently contaminate putative mossy fibers synaptic responses measured in CA3 pyramidal cells. The use of overly conservative criteria on the other side may lead to the exclusion of “pure” mossy fibers responses with unexpected properties (Henze et al., 2000).However the recent and fast growth of optogenetics in neuroscience has provided new tools with high spatial selectivity (cell specific optical activation) and temporal precision (at the millisecond scale), allowing the dissection and investigation of neuronal circuits in vivo. The aim of my thesis was to gain insight into the mechanisms and the physiological consequences of short-term synaptic plasticity occurring at mossy fibers to CA3 pyramidal neurons synapses in the intact mouse brain. The present thesis consists of two main parts. In the first part I explored new optogenetic tools to control the activity of granule cells with pulses of light. Optogenetic stimulation, which relies on the activation of the light-gated ion channel channelrhodopsin-2 (ChR2) by blue light reliably induced action potentials over a wide range of frequencies of stimulation. I also found that optical stimulation can be used to trigger short term plasticity at mf-CA3 synapses. In the second part I refined optogenetic stimulation methodology in vivo for non-invasive characterization of synaptic functioning of the mf-CA3 synapses. The reliability of optogenetic stimulation of a genetically targeted neuronal population together with the single cell resolution obtained using whole-cell recordings are important steps towards a better understanding of the functional role of the mossy fibers in the hippocampal network in vivo
Structure/function study and modulation of kainate receptors
No full textLes récepteurs de type kaïnate (rKA) appartiennent, avec les récepteurs de type NMDA (rNMDA) et les recepteurs de type AMPA (rAMPA), à la famille des récepteurs canaux glutamatergiques (iGluR). Les propriétés fonctionelles des rKA contenant la sous-unité GluK3 en font des récepteurs tout à fait singuliers. Une étude réalisée dans le laboratoire a montré que la faible sensibilité de ces récepteurs au glutamate est liée à une entrée très rapide dans l’état désensibilisé et que la fonction de ces récepteurs pourrait être amplifiée par des modulateurs endogènes.Parmi les modulateurs potentiels de la fonction des rKA pré-synaptiques, nous avons choisi d’étudier le zinc, en raison de sa concentration importante dans les vésicules des terminaisons des axones des cellules granulaires du gyrus denté (fibres moussues). En dépit du rôle proposé des rKA contenant la sous unité GluK3 dans la régulation pré-synaptique aux synapses MF-CA3, la modulation de ces récepteurs par le zinc n’a jamais été étudiée.Grâce à l’enregistrement électrophysiologique des courants GluK3 exprimés dans les cellules HEK-293, nous avons montré que le zinc facilite les courants des récepteurs contenant la sous-unité GluK3, activés par le glutamate. L’analyse des cinétiques, ainsi que la modélisation, montrent que l’effet facilitateur du zinc est dû à la réduction de l’entrée dans l’état désensibilisé des récepteurs GluK3. Grâce à la mutagénèse dirigée et l’étude cristallographique, nous avons pu déterminer le site de liaison du zinc, constitué de l’aspartate 759, de l’histidine 762 et de l’aspartate 730, et localisé dans l’interface de dimérisation du domaine de liaison de l’agoniste (LBD).Cette étude décrit pour la première fois un nouveau site de modulation positive de la fonction des rKA.Glutamate released at excitatory synapses acts on ligand-gated ionotropic receptors which fall into three classes: NMDA, AMPA and kainate receptors.At hippocampal mossy fiber synapses onto CA3 pyramidal cells, KARs are present both at the pre- and postsynaptic levels. Postsynaptic KARs are composed of the GluK2, GluK4 and GluK5 subunits, whereas presynaptic KARs are thought to comprise the GluK2 and GluK3 subunits. The functional properties of GluK3 (and GluK2/GluK3) receptors set it apart from the other ionotropic glutamate receptors. In particular, its sensitivity to glutamate is the lowest of all known ionotropic glutamate receptors, due in large part to fast desensitization of receptors with one or two bound glutamate molecules. The low agonist sensitivity of this receptor raises questions about its relevance for synaptic function. Therefore, it is possible that endogenous modulators may potentiate its function.Among potential endogenous modulators of KAR function, we chose to address the role of zinc, because of the large amounts contained in mossy fiber terminals. Zinc is thought to be accumulated into synaptic vesicles, and is co-released with glutamate in the extracellular milieu during neuronal activity. Zinc has been reported to inhibit most of native and recombinant KARs. Despite the proposed role of at hippocampal mossy fiber synapses, although modulation of GluK3-containing KARs by zinc has not yet been addressed.In this study, we show that zinc greatly potentiates recombinant GluK3 receptor currents evoked by glutamate. Zinc markedly slows receptor desensitization and increases apparent affinity for glutamate. Crystallographic studies and analysis of chimeric GluK2/GluK3 KARs and of GluK3 bearing selected point mutations, allowed us to identify the zinc binding domain defined by D759, H762, Q756 and D730, and localized in a region forming the interface between two GluK3 subunits in an LBD dimer assembly. Based on these structure-function studies and on modeling of KAR activity, we show that zinc plays a very distinct role on GluK3-KARs by stabilizing the interaction between dimers of LBD thereby reducing desensitization.Given the proposed localization of GluK3 close to zinc containing synaptic vesicles, zinc may be an endogenous allosteric modulator for native GluK3-KARs, and its binding site a new pharmacological target
Plasticité présynaptique et encodage de la mémoire dans les circuits de l'hippocampe
No full textCe projet étudie les changements dans la dynamique intracellulaire (propriétés synaptiques et intrinsèques) des neurones dans la région CA3 de l’hippocampe, pendant l’encodage d’une mémoire épisodique, chez la souris. L’architecture unique des circuits CA3 est censée permettre le stockage et le rappel rapide des mémoires associatives. Le réseau dense d’interconnexions entre les principales cellules de CA3 forme un réseau attracteur qui favoriserait ces processus. La capacité des circuits à générer des modèles d’activité rythmique essentiels pour différentes phases de la mémoire (encodage, consolidation, rappel) est basée sur la modulation des propriétés des neurones individuels en fonction des états cérébraux. À ce jour, aucune étude n’a examiné l’activité intracellulaire des neurones CA3 individuels.This project investigates the changes in the intracellular dynamics (synaptic and intrinsic properties) of neurons in the CA3 region of the hippocampus, during the encoding of an episodic-like memory, in mice.The unique architecture of CA3 circuits is thought to enable the storage and rapid recall of associative memories. The dense network of interconnections between the principal cells of CA3 forms an attractor network that would favor these processes. The ability of circuits to generate patterns of rhythmic activity essential for different phases of memory (encoding, consolidation, recall) is based on the modulation of the properties of individual neurons as a function of brain states. To date, no study has investigated the intracellular activity of individual CA3 neurons in awake animals. Ashley Kees and Meryl Malézieux in the laboratory have since 4 years developed very original techniques to record the intracellular dynamics of CA3 pyramidal cells in relation to the brain state of the animal. This relies on in vivo patch-clamp experiments in head-fixed awake mice, which provides rich information linking single cell activity to activity of neuronal ensembles. The great originality of the project consists in following the changes in the intracellular dynamics of CA3 cells, during memory processing of multimodal information. The thesis project is a follow-up to this work and will benefit from the methodological developments and analysis already carried out in the laboratory. These techniques will be supplemented by the development of behavioral memory protocols in a head-fixed animal, and the application of optogenetic techniques to modulate the activity of afferent neurons during the memory task
Plasticité synaptique dans un circuit de la mémoire : rôle physiologique et physiopathologique de la protéine précurseur du peptide amyloïde
No full textLa protéine précurseur de l'amyloïde (APP), un acteur clé de la maladie d'Alzheimer, est exprimée de manière ubiquitaire dans le cerveau. L'APP est abondamment exprimée dans les compartiments présynaptiques où elle interagit avec les protéines de la machinerie de libération synaptique. Cependant, les fonctions physiologiques de l'APP au niveau des synapses restent en grande partie inconnues. Dans ce travail, nous étudions le rôle physiologique de l'APP dans la plasticité synaptique à court terme et dans le transfert d'informations dans les circuits de l'hippocampe. Notre travail se concentre sur la région CA3 de l'hippocampe et sur les synapses des fibres moussues (MF) établies entre les axones du gyrus denté (DG) et les cellules pyramidales de CA3 (CA3 PC).Nous avons supprimé l'APP et la protéine apparentée APLP2 dans les cellules granulaires du DG (DG-GC) en utilisant une stratégie de transfert viral de gène chez des souris doublement floxées APP/APLP2. En combinant l'optogénétique et l'électrophysiologie ex vivo, nous avons découvert que la suppression sélective de l'APP dans les DG-GC réduit fortement la plasticité présynaptique à court terme aux synapses MF-CA3 PC, exprimée par un niveau élevé de facilitation synaptique en réponse à une stimulation présynaptique répétée, ceci sans altérer les propriétés synaptiques de base ni l'excitabilité intrinsèque des DG-GC. De plus, l'absence d'APP altère l'évolution temporelle de la potentialisation post-tétanique, une forme de plasticité présynaptique qui dure plusieurs minutes après un train de stimulation à haute fréquence. Nous avons ensuite étudié les mécanismes moléculaires par lesquels l'APP contrôle la plasticité présynaptique à court terme en isolant des synaptosomes MF-CA3 marqués par la GFP et en effectuant un criblage protéomique comparatif pour identifier les protéines présynaptiques dérégulées en l'absence d'APP. Nous avons identifié des cibles potentiellement intéressantes qui sont dérégulées, notamment ZnT3, les complexines I et II et CSPalpha. Nous discutons du rôle causal potentiel de ces protéines dans les déficits fonctionnels observés, ainsi que des mécanismes possibles par lesquels l'APP contrôle la plasticité présynaptique.Parallèlement, nous avons supprimé l'APP et l'APLP2 sélectivement dans les CA3 PC post-synaptiques et nous avons enregistré les courants synaptiques MF-CA3. Nous avons découvert que l'absence d'APP dans les CA3 PC entraîne une diminution sélective des EPSCs MF-CA3 médiés par les récepteurs kaïnate (KAR), mais pas par les récepteurs AMPA ou NMDA. Ce phénomène a également été observé chez la souris APP/PS1, modèle de la maladie d'Alzheimer, et chez les souris KO conditionnelle pour la préséniline. Nous rapportons que la sous-unité GluK2 des KARs interagit avec l'APP et ses fragments biologiquement actifs, et que cette interaction est probablement responsable de la régulation des niveaux de KARs synaptiques. Nous interprétons ces résultats par un rôle transsynaptique précédemment inconnu de l'APP.Dans l'ensemble, nos données confirment le rôle central de l'APP dans la transmission synaptique et les mécanismes de plasticité au niveau de la synapse MF-CA3. Nous pensons que la mise en lumière de la contribution physiologique de l'APP dans l'activité des circuits hippocampiques nous permettra de mieux comprendre comment la perturbation des fonctions de l'APP contribue à la physiopathologie de la maladie d'Alzheimer.The full-length amyloid precursor protein (APP), a key player in Alzheimer’s disease (AD), is ubiquitously expressed throughout the brain. APP is abundantly expressed in presynaptic compartments where it interacts with proteins of the presynaptic release machinery. However, the physiological functions of APP at synapses remain in great part elusive. In this work, we study the physiological role of APP in short-term synaptic plasticity and information transfer within the hippocampus circuitry. Our work focuses on the CA3 region of the hippocampus and on mossy fiber (MF) synapses between the axons of the dentate gyrus (DG) and CA3 pyramidal cells (CA3 PCs).We deleted APP and the related protein APLP2 in DG granular cells (DG-GCs) using a viral gene transfer strategy in APP/APLP2 double floxed mice. By combining optogenetics and ex vivo electrophysiology, we found that the selective deletion of APP in DG-GCs strongly impairs presynaptic short term plasticity at MF-CA3 PC synapses, expressed as a high level of synaptic facilitation in response to repeated presynaptic stimulation, without altering nor basal synaptic properties nor intrinsic excitability of DG-GCs. Additionally, the lack of APP altered the time course of post-tetanic potentiation, a form of presynaptic plasticity lasting several minutes following a high frequency train of stimulation. We then investigated the molecular mechanisms by which APP controls presynaptic short-term plasticity by isolating GFP-labelled MF-CA3 synaptosomes, and by performing a comparative proteomic screening to identify presynaptic proteins dysregulated in the absence of APP. We identified prospectively interesting targets that are dysregulated, including ZnT3, complexins I and II, and CSPalpha. We discuss the potential causative role of these proteins in the observed functional deficits, as well as the possible mechanisms by which APP controls presynaptic plasticity.Concurrently, we virally deleted APP and APLP2 in postsynaptic CA3 PCs and performed ex vivo electrophysiology. We discovered that the lack of APP in CA3 pyramidal cells leads to a selective downregulation of MF-CA3 EPSCs mediated by kainate receptors (KARs), but not by AMPA or NMDA receptors. This was also observed in the APP/PS1 mouse model of AD and in conditional presenilin KO mice. We report that the GluK2 subunit of KARs interacts with APP and its biologically active fragments, and this interaction is likely responsible for regulating synaptic KARs levels. We interpret these results by a previously unknown transsynaptic role for APP.Altogether, our data supports a pivotal role of APP in synaptic transmission and plasticity mechanisms at the MF-CA3 synapse. We propose that shedding light on the physiological contribution of full-length APP in the activity of hippocampal circuits will enhance our understanding of how disruption of APP functions contributes to the pathophysiology of AD
Pathophysiology of Hippocampal CA3 Neurons in the APP/PS1 Mouse Model of Alzheimer’s Disease
No full textTese de doutoramento em Ciências da Saúde, no ramo de Ciências Biomédicas, apresentada à Faculdade de Medicina da Universidade de CoimbraAlzheimer’s Disease (AD) is a progressive neurodegenerative disease characterized clinically by progressive memory loss eventually resulting in dementia. Over the course of AD amyloid-beta (Aβ) deposition forms insoluble amyloid plaques but synapse loss is known be better correlated with the progression of the disease. Although the exact role of Aβ is not fully understood, recent evidence suggests that subtle alterations of synaptic transmission precede neuronal degeneration in the AD progression. Early disturbance of synaptic processes involved in learning and memory have been reported in several transgenic mouse models. In particular, the APP/PS1 transgenic mice display contextual memory impairments at 6 months, before accumulation of amyloid plaques. The Dentate Gyrus (DG) and the CA3 regions of the hippocampus are essential for the rapid encoding of contextual memories. Although this brain structure plays a key role in memory formation it has been poorly studied in AD mouse models. Thus, our project aimed to characterize the structural and functional age-dependent deficits of glutamatergic synaptic transmission in hippocampal CA3 pyramidal region. This region represents also an interesting model to study synaptic physiology and dysfunction since CA3 pyramidal cells receive different types of glutamatergic inputs, from dentate granule cells (mossy fibers), from other CA3 pyramidal cells (associative/commissural fibers), and from the entorhinal cortex (perforant path synapses). These inputs differ in terms of the position along the dendrites where they contact CA3 pyramidal cells, their spine structure, their glutamate receptor composition, their presynaptic properties, and the types of synaptic plasticities expressed (short and long term-synaptic plasticity). Combining patch-clamp electrophysiological recordings in acute hippocampal slices with high-resolution fluorescence microscopy of cells infected with new viral tools to study morphological changes we have characterized synapse specific alterations in APP/PS1 mice with 6 months.
We found that mossy fiber synapses (Mf) are relatively spared in transgenic animals as compared to associative-comisssural (A/C) synapses both functionally and morphologically. The prominent Mf short-term presynaptic plasticities showed not to be affected at 6 months, and their characteristic presynaptic long-term potentiation (LTP) was also normal. Morphology analysis also revealed no alterations in size of the Mf pre and postsynaptic compartments. There was although an alteration in the complexity of the Mf boutons (presynaptic) that was not matched with a postsynaptic alteration (thorny excrescences, display similar complexity index between genotypes). This result suggests alterations of the filopodia size and/or number, and putative impairments concerning the GABAergic network. In contrast to the lack of presynaptic glutamatergic deficits at Mf synapses, we found a striking loss of NMDA receptor (NMDAR)-dependent long-term plasticity (LTP) at A/C synapses. This loss of LTP was correlated with subtle alterations of the dendritic spine morphology accompanied by a reduced spine density. Importantly, we found that the loss of postsynaptic LTP was not caused by alterations in the amount of synaptic NMDARs, since we found no alteration in NMDAR/AMPAR ratio at A/C synapses. We further exclude alterations in extrasynaptic NMDARs signaling.
At the cellular level processes that result in long-term modifications of synaptic strength such as LTP and LTD are thought to be key mechanisms underlying learning and memory. The results we obtained clearly highlight the importance of addressing the role individual synapses play in AD with the capability of spatially separating inputs and the high temporal resolution that just whole-cell patch-clamp recordings can provide. Our results also point out that, in this model, at an early phase of the disease the main synaptic deficits are post-synaptic, and not presynaptic as some of the previous work suggested. Furthermore, our data points out that the chronic deposition of Aβ disturbed LTP without altering NMDARs conductance (synaptically and extrasynaptically), suggesting that the loss of LTP might be due to alterations in intracellular mechanisms downstream NMDARs. A2A receptors (A2ARs) and metabotropic glutamate receptors 5 (mGluR5) are known to modulate NMDAR signaling and reported to be important targets for AD treatment. Furthermore, both caffeine (antagonist of A2AR) and MTEP (antagonist of mGluR5) have been reported to rescue behavioural deficits in AD mice models. We found that short application of both these drugs resulted in a partial rescue of the LTP levels in the A/C synapses of APP/PS1 animals. Our study provides novel insight into the mechanisms of early AD, examines the cellular deficits underlying learning and memory impairment and explores promising drugs for their potential to reverse those cellular deficits.A doença de Alzheimer (DA) é uma doença neurodegenerativa que, do ponto
de vista clínico, é caracterizada inicialmente pela perda da memória de curto prazo
que conduz, irreversivelmente, a perda progressiva de capacidades cognitivas. No
curso da DA há acumulação de péptidos β-amilóides (Aβ) que formam depósitos
insolúveis denominados placas amilóides. A evidência disponível indica que o início da
deterioração da memória observada na DA é devido a alterações na função sináptica e
não à acumulação de placas amilóides ou à perda de neurónios. Contudo, o
mecanismo de ação pelo qual os péptidos Aβ levam à alteração da transmissão
sináptica não é completamente conhecido. Ratinhos transgénicos, que apresentam
uma elevada taxa de produção de Aβ, têm sido utilizados como modelos da DA, dado
que sofrem uma degradação progressiva da função cognitiva. Em particular, o ratinho
transgénico APP/PS1 apresenta uma deterioração da memória de trabalho que tem
início antes de formação das placas amilóides. A comunicação entre o giro dentado
(GD) e a região CA3 do hipocampo, feita através dos axónios das células granulares
do GD, é essencial para este tipo de memória. Também a comunicação sináptica entre
os neurónios piramidais de CA3 é fundamental no processamento da informação no
hipocampo, em particular no que respeita à implementação de novas memórias. O
objetivo principal do trabalho desenvolvido nesta dissertação de doutoramento foi a
caracterização das alterações estruturais e funcionais das sinapses glutamatérgicas
na região CA3 de ratinhos transgénicos APP/PS1. Esta região do hipocampo, apesar
do seu papel na formação de traços mnemónicos, tem sido muito pouco estudada em
modelos animais da DA. Além disso, os circuitos neuronais de CA3 são modelos
interessantes no estudo da transmissão e plasticidade sináptica pois os neurónios de
CA3 recebem vários tipos de aferentes glutamatérgicos: das fibras musgosas (Mf)
provenientes do GD; das fibras associativas/comissurais (A/C) provenientes de outros
neurónios de CA3; das fibras da via perfurante (PP) diretamente do córtex
entorrinal. Estes três aferentes apresentam diferenças importantes em termos de
localização dos seus terminais nervosos ao longo da árvore dendrítica, mas também
em termos de morfologia da espinha dendrítica, da composição de recetores de
glutamato e das propriedades de plasticidade sináptica de curto e longo termo prazo.
Através de estudos de electrofisiologia e de imagiologia confocal de super-resolução, procedeu-se à caracterização de alterações de vários destes parâmetros nos ratinhos
APP/PS1 que modelam a DA.
Este estudo permitiu identificar que, em ratinhos APP/PS1 com 6 meses de
idade, as sinapses das Mf estão relativamente preservadas em termos de alterações
fisiológicas e morfológicas, em comparação com as sinapses A/C. Diferentes
protocolos de estimulação pré-sináptica mostraram que a plasticidade sináptica
característica das Mf não sofreu alterações. Em linha com estes resultados não se
registaram também alterações no volume ou na área dos terminais pré-sinápticos ou
componentes pós-sinápticos. Contudo, observou-se uma alteração no índice de
complexidade dos terminais pré-sinápticos, sem alterações pós-sinápticas, que pode
ser indicativa de alterações no número ou tamanho dos contactos (filopodia) com
interneurónios GABAérgicos. Em contaste com a ausência de défices na comunicação
glutamatérgica nas Mf, observou-se uma total perda da capacidade de potenciação de
longa duração (LTP) nas sinapses A/C. Esta alteração electrofisiológica foi
acompanhada por uma subtil alteração da densidade sináptica, mas sem modificações
morfológicas de relevo. Surpreendentemente, concluiu-se que esta perda de LTP não
é devida a alterações no número ou nas características dos recetores de glutamato
presentes na membrana, uma vez que os rácios de recetores AMPA e NMDA na
sinapse não se encontravam alterados. Também o número e/ou condutância dos
recetores NMDA extrasinápticos, importantes para a plasticidade sináptica, não estava
alterado neste modelo de AD.
As modificações da plasticidade sináptica como LTP e LTD (depressão de
longa duração) são considerados os principais mecanismos neurofisiológicos que
estão na base da aprendizagem e memória. Os resultados obtidos neste estudo
ilustram o potencial e a importância que a investigação de sinapses individuais,
através de técnicas de patch-clamp, oferece para compreender os mecanismos da DA,
com vista a melhorar as terapêuticas disponíveis. Por exemplo, no modelo de DA
utilisado neste estudo, a produção anormal de péptidos Aβ causou défices sinápticos
que foram maioritariamente pós-sinápticos, sendo estas alterações independentes do
número ou da composição molecular dos recetores NMDA. Estes dados sugerem que
a perda de LTP se deve à alteração de mecanismos intracelulares de controlo dos
recetores NMDA e não a defeitos na sua ativação. Os receptores de adenosina A2A e
os receptores metabotrópicos de glutamato do tipo 5 (mGluR5) tem a capacidade de modular processos de plasticidade sináptica dependentes de recetores NMDA. Estes
recetores A2A e mGluR5 estão presentes na membrana pós-sináptica de várias
sinapses do hipocampo, sendo ambos alvo de fármacos (cafeína, antagonista de
receptores A2A e MTEP, antagonista de mGluR5) responsáveis pela atenuação de
sintomas da DA em diversos modelos animais da DA. Conformemente, observou-se
que uma curta incubação de fatias de cérebro de ratinhos APP/PS1 com um
antagonista de recetores A2A ou de recetores mGluR5 foi suficiente para recuperar
parcialmente os níveis de LTP registados nas sinapses A/C. Em conclusão, ao
analisar as células de CA3, este estudo permitiu detalhar alterações sinápticas que
estão subjacentes à aprendizagem e memória, bem como documentar efeitos
sinápticos de fármacos capazes de reverter os sintomas da DA.Fundação para a Ciência e Tecnologia - SFRH/BD/33468/200
Morpho-functional maturation of hippocampal mossy fiber synapses
No full textLes synapses se forment selon plusieurs étapes comprenant la stabilisation des contacts nouvellement formés et leur maturation. Ces différentes étapes dépendent d’une mise en place coordonnée entre la terminaison pré- et postsynaptique. Les protéines composant la présynapse et les récepteurs ionotropiques du glutamate ont des rôles clés dans ces processus. Lors de ma thèse, je me suis intéressé à l’implication de la protéine présynaptique Bassoon lors de la maturation des synapses glutamatergiques entre les fibres moussues et les cellules pyramidales de CA3 dans l’hippocampe. Cette synapse constitue un modèle attractif pour l’étude de la maturation synaptique car elle suit des étapes de maturation morphologique et fonctionnelle bien définies. Bassoon est une des premières protéines se mettant en place au niveau des contacts synaptiques nouvellement formés. Par des approches électrophysiologiques, nous avons montré que la protéine Bassoon était importante pour l’organisation du site de libération de neurotransmetteur durant les deux premières semaines de vie post-natale chez la souris.Les récepteurs kaïnate jouent un rôle important dans la régulation de l’activité de réseau au cours du développement post-natal. Cependant l’impact de l’activation de ces récepteurs sur la maturation synaptique est peu connu. J’ai pu mettre en évidence un délai dans la maturation fonctionnelle de la synapse fibre moussue/cellule pyramidale de CA3 chez les souris déficientes pour la sous-unité GluK2 des récepteurs kaïnate (GluK2-/-). Afin de comprendre si ce délai de maturation fonctionnelle est corrélé à un retard dans la maturation morphologique de cette synapse, nous avons mis en place des infections de lentivirus codant pour une protéine membranaire fluorescente (YFP) chez le souriceau nouveau-né (P1-P2). A l’aide de microscopie confocale et de reconstruction en 3D, nous avons ainsi pu décrire la maturation morphologique de la synapse fibre moussue/cellule pyramidale de CA3. Cela m’a également permis de corréler la maturation fonctionnelle à la maturation morphologique et mes résultats montrent également un retard dans la mise en place des synapses chez les souris GluK2-/-. L’ensemble de cette étude révèle l’importance de l’activité synaptique et de la coordination entre mise en place de la pré- et de la postsynapse au cours de la maturation synaptique.The formation of synapses follows different steps including synaptogenesis and maturation. These different steps depend on coordinated pre- and post-synaptic assembly. Pre-synaptic proteins and ionotropic glutamate receptors play a central role in these processes. During my thesis, I have been interested in the implication of the presynaptic protein Bassoon in the maturation of the hippocampal mossy fiber to CA3 pyramidal cell glutamatergic synapses. This synapse constitutes an attractive model for the study of synaptic maturation because it follows several steps of defined morphological and functional maturation. Bassoon in one of the first protein present at newly formed synaptic contacts. By electrophysiological approaches, we showed that Bassoon is important for the organization of the active zone during the first two postnatal weeks.Kainate receptors play an important role in the regulation of network activity during postnatal development. However, the impact of kainate receptors activation on synaptic maturation is less known. I showed a delay in functional maturation of mossy fiber synapses in mice deficient for the GluK2 subunit of kainate receptors (GluK2-/-). To know if this delay is correlated to morphological alterations of this synapse, we setup in vivo lentiviral infections of membrane fluorescent protein (YFP) in mouse pups (P1-P2). Using confocal microscopy and 3D reconstruction, we described the morphological maturation of mossy fiber synapses. We were able to correlate functional and morphological maturation and our results also showed an impairment in the formation of mossy fiber synapses in GluK2-/-. Together, these data reveal the importance of synaptic activity and of the coordination of pre- and post-synaptic assembly during synaptic maturation
Morisset/PyNeb_devel: 1.1.9
No full textcommit ab6cdb8a4af2ff75b82fc1422004934945c82530
Author: morisset [email protected]
Date: Sat Jul 6 11:25:45 2019 -0700
1.1.9
V 1.1.9
commit f7e6a418d2affb8db163c8c6c2428173b5f4aacf
Author: morisset [email protected]
Date: Fri Jun 7 07:15:42 2019 -0700
Create _chianti_tools_9.py
commit 81213b885e2b359a62e3d6e73515e39dc7056561
Author: Christophe Morisset [email protected]
Date: Wed Jun 5 16:19:33 2019 -0700
Adding Chianti 9
commit 5a9a7dfc9a4d922a216ac11f39b76c2d63097d3b
Author: Christophe Morisset [email protected]
Date: Wed Jun 5 16:19:05 2019 -0700
1.1.9b4
commit 152f9192847cd7f3194e0a778caca66f966392c1
Author: morisset [email protected]
Date: Wed May 22 14:13:30 2019 -0700
T_min ans max were not used
commit 22aeec4db4efc7bcc377b54a92292e157335825d
Author: morisset [email protected]
Date: Sun May 19 10:18:52 2019 -0700
1.1.9b3
commit 4dc721eecdd2511fcde69f835fc6181f7be1b74d
Author: morisset [email protected]
Date: Sun May 19 10:18:34 2019 -0700
manage wl<Lyalpha master
commit 44192233b57ef46cc479e6bfc2a9020cbfc3d5f6
Author: morisset [email protected]
Date: Fri May 17 13:07:55 2019 -0700
1.1.9b2
commit 95e07d78fb38cc6267f38e11d3051738814a4dfd
Author: morisset [email protected]
Date: Fri May 17 13:07:47 2019 -0700
light wl array
commit 9a159290e931fbdbbff8b13968d7540f8b0cef18
Author: morisset [email protected]
Date: Fri May 17 13:07:27 2019 -0700
add continuum to init
commit da0e0b6ac19bc6808bc7366217ad4a0885e1ca0e
Author: morisset [email protected]
Date: Thu Mar 21 11:48:08 2019 -0300
1.1.9b1
before branch to Regresor developemen
Christophe Colomb, le caoutchouc et les tétines
No full textChristoph Kolumbus, Kautschuk und die Gummisauger.
Der Verfasser schildert kurz die Geschichte des Kautschuks und erwänt die apotheker welche daran beteiligt sind. Er legt anschliessend die Geschichte der Gummisauger und der Milchflaschen dar, deren eine der meistgängisten die « Robert'sche Milchflasche » gewesen ist.Christopher Columbus, rubber, and rubber nipples.
The author quickly traces the history of rubber, making note of the pharmacists who therein played a role. He exhibits the history of rubber nipples and that of infant bottles, of which one of the most widely used was the « Robert Bottle. »Labeÿ Robert. Christophe Colomb, le caoutchouc et les tétines. In: Revue d'histoire de la pharmacie, 82ᵉ année, n°300, 1994. pp. 55-63
Pathophysiology of information processing in neocortical dendrites in Fragile X Syndrome
No full textLe Syndrome de l’X Fragile (SXF) est la forme héréditaire de retard mental la plus fréquente et la cause la mieux caractérisée de troubles du spectre autistique (TSA). Elle est causée par une mutation causant l’inactivation du gène Fmr1 (codant pour la protéine FMRP). La sensibilité accrue aux stimuli sensoriels est une caractéristique importante du SXF et des TSA, mais les mécanismes sous-jacents sont encore mal compris. Nous avons constaté que la suppression du gène Fmr1 entrainait une hyperexcitabilité sensorielle dans le modèle murin du SXF. Les souris Fmr1KO nécessitaient significativement moins d'informations tactiles pour l'exploration haptique, et les représentations évoquées par les informations tactiles provenant des vibrisses dans le cortex somatosensoriel primaire (S1) se propageaient à une vitesse plus élevée chez les souris Fmr1KO par rapport aux souris témoins sauvages.Au niveau cellulaire, il a été montré que les ARNm de plusieurs sous-unités de canaux ioniques (par exemple HCN1, KCNMA1) jouant un rôle clé dans le traitement de l'information dendritique / neuronale étaient des cibles de la protéine FMRP (Liao et al, 2008; Darnell et al, 2011). Sur la base de ces observations, nous avons étudié les canalopathies comme une caractéristique importante du SXF. Nous avons testé de possibles dysfonctionnement des canaux ioniques, et leurs conséquences sur le traitement de l'information dendritique dans les neurones pyramidaux du néocortex de la couche 5 chez les souris Fmr1KO, en utilisant une combinaison d’approches électrophysiologiques et d’imagerie calcique bi-photonique. Nos résultats ont montré que les dendrites des neurones pyramidaux du S1 étaient hyperexcitables, facilitant ainsi le couplage des entrées d’information synaptique à la génération de potentiel d'action en sortie dans les neurones. Cette altération était, au moins en partie, attribuable à un dysfonctionnement des canaux Ih et BKCa et a été partiellement restaurée par l'activation pharmacologique des canaux BKCa. Ces résultats plaident en faveur d'un rôle nouveau et crucial des canalopathies dans l'expression de l'hyperexcitabilité sensorielle dans le SXF.Fragile X Syndrome (FXS) is the most common form of inherited mental retardation syndrome and most well characterized cause of Autism Spectrum Disorders (ASD), and it is caused by a silencing mutation of the gene Fmr1 (encoding the protein FMRP). Increased sensitivity to sensory stimuli is a prominent feature of FXS and ASD, but its underlying mechanisms are poorly understood. We found that deletion of the Fmr1 gene results in somatosensory hyper-excitability in a mouse model for FXS. Fmr1 knockout (Fmr1KO) mice required significantly less tactile information for haptic exploration, and touch-evoked whisker representations in the primary somatosensory cortex (S1) spread with increased velocity in Fmr1KO mice compared to wild-type control. At the cellular level, it has been shown that the mRNAs of several ion channel subunits (e.g. HCN1, KCNMA1) playing key roles in dendritic/neuronal information processing are regulated by FMRP (Liao et al., 2008; Darnell et al., 2011). Based on these observations, we investigated channelopathies as a prominent feature of FXS. We probed ion channel dysfunction, and its consequence for dendritic information processing in neocortical pyramidal neurons of layer 5 in Fmr1KO mice, using a combination of electrophysiological and 2-photon calcium imaging approaches. Our results showed that dendrites of S1 pyramidal neurons were hyper-excitable, facilitating the coupling of synaptic input to the generation of action potential output in these neurons. This defect was, at least in part, attributable to a dysfunction of Ih channels and BKCa channels and was partially rescued by pharmacological activation of BKCa channels. These findings argue for a novel and critical role for channelopathies in the expression of sensory hyper-excitability in FXS
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