External forcings, oceanographic processes and particle flux dynamics in Cap de Creus submarine canyon, NW Mediterranean Sea

Particle fluxes (including major components and grain size), and oceanographic parameters (near-bottom water temperature, current speed and suspended sediment concentration) were measured along the Cap de Creus submarine canyon in the Gulf of Lions (GoL; NW Mediterranean Sea) during two consecutive winter-spring periods (2009 2010 and 2010 2011). The comparison of data obtained with the measurements of meteorological and hydrological parameters (wind speed, turbulent heat flux, river discharge) have shown the important role of atmospheric forcings in transporting particulate matter through the submarine canyon and towards the deep sea. Indeed, atmospheric forcing during 2009 2010 and 2010 2011 winter months showed differences in both intensity and persistence that led to distinct oceanographic responses. Persistent dry northern winds caused strong heat losses (14.2 × 103 W m−2) in winter 2009 2010 that triggered a pronounced sea surface cooling compared to winter 2010 2011 (1.6 × 103 W m−2 lower). As a consequence, a large volume of dense shelf water formed in winter 2009 2010, which cascaded at high speed (up to ∼1 m s−1) down Cap de Creus Canyon as measured by a current-meter in the head of the canyon. The lower heat losses recorded in winter 2010 2011, together with an increased river discharge, resulted in lowered density waters over the shelf, thus preventing the formation and downslope transport of dense shelf water. High total mass fluxes (up to 84.9 g m−2 d−1) recorded in winter-spring 2009 2010 indicate that dense shelf water cascading resuspended and transported sediments at least down to the middle canyon. Sediment fluxes were lower (28.9 g m−2 d−1) under the quieter conditions of winter 2010 2011. The dominance of the lithogenic fraction in mass fluxes during the two winter-spring periods points to a resuspension origin for most of the particles transported down canyon. The variability in organic matter and opal contents relates to seasonally controlled inputs associated with the plankton spring bloom during March and April of both years.

Behavioural queueing : cellular automata and a laboratory experiment

Dans cette thèse, nous étudions les aspects comportementaux d'agents qui interagissent dans des systèmes de files d'attente à l'aide de modèles de simulation et de méthodologies expérimentales. Chaque période les clients doivent choisir un prestataire de servivce. L'objectif est d'analyser l'impact des décisions des clients et des prestataires sur la formation des files d'attente. Dans un premier cas nous considérons des clients ayant un certain degré d'aversion au risque. Sur la base de leur perception de l'attente moyenne et de la variabilité de cette attente, ils forment une estimation de la limite supérieure de l'attente chez chacun des prestataires. Chaque période, ils choisissent le prestataire pour lequel cette estimation est la plus basse. Nos résultats indiquent qu'il n'y a pas de relation monotone entre le degré d'aversion au risque et la performance globale. En effet, une population de clients ayant un degré d'aversion au risque intermédiaire encoure généralement une attente moyenne plus élevée qu'une population d'agents indifférents au risque ou très averses au risque. Ensuite, nous incorporons les décisions des prestataires en leur permettant d'ajuster leur capacité de service sur la base de leur perception de la fréquence moyenne d'arrivées. Les résultats montrent que le comportement des clients et les décisions des prestataires présentent une forte "dépendance au sentier". En outre, nous montrons que les décisions des prestataires font converger l'attente moyenne pondérée vers l'attente de référence du marché. Finalement, une expérience de laboratoire dans laquelle des sujets jouent le rôle de prestataire de service nous a permis de conclure que les délais d'installation et de démantèlement de capacité affectent de manière significative la performance et les décisions des sujets. En particulier, les décisions du prestataire, sont influencées par ses commandes en carnet, sa capacité de service actuellement disponible et les décisions d'ajustement de capacité qu'il a prises, mais pas encore implémentées. - Queuing is a fact of life that we witness daily. We all have had the experience of waiting in line for some reason and we also know that it is an annoying situation. As the adage says "time is money"; this is perhaps the best way of stating what queuing problems mean for customers. Human beings are not very tolerant, but they are even less so when having to wait in line for service. Banks, roads, post offices and restaurants are just some examples where people must wait for service. Studies of queuing phenomena have typically addressed the optimisation of performance measures (e.g. average waiting time, queue length and server utilisation rates) and the analysis of equilibrium solutions. The individual behaviour of the agents involved in queueing systems and their decision making process have received little attention. Although this work has been useful to improve the efficiency of many queueing systems, or to design new processes in social and physical systems, it has only provided us with a limited ability to explain the behaviour observed in many real queues. In this dissertation we differ from this traditional research by analysing how the agents involved in the system make decisions instead of focusing on optimising performance measures or analysing an equilibrium solution. This dissertation builds on and extends the framework proposed by van Ackere and Larsen (2004) and van Ackere et al. (2010). We focus on studying behavioural aspects in queueing systems and incorporate this still underdeveloped framework into the operations management field. In the first chapter of this thesis we provide a general introduction to the area, as well as an overview of the results. In Chapters 2 and 3, we use Cellular Automata (CA) to model service systems where captive interacting customers must decide each period which facility to join for service. They base this decision on their expectations of sojourn times. Each period, customers use new information (their most recent experience and that of their best performing neighbour) to form expectations of sojourn time at the different facilities. Customers update their expectations using an adaptive expectations process to combine their memory and their new information. We label "conservative" those customers who give more weight to their memory than to the xiv Summary new information. In contrast, when they give more weight to new information, we call them "reactive". In Chapter 2, we consider customers with different degree of risk-aversion who take into account uncertainty. They choose which facility to join based on an estimated upper-bound of the sojourn time which they compute using their perceptions of the average sojourn time and the level of uncertainty. We assume the same exogenous service capacity for all facilities, which remains constant throughout. We first analyse the collective behaviour generated by the customers' decisions. We show that the system achieves low weighted average sojourn times when the collective behaviour results in neighbourhoods of customers loyal to a facility and the customers are approximately equally split among all facilities. The lowest weighted average sojourn time is achieved when exactly the same number of customers patronises each facility, implying that they do not wish to switch facility. In this case, the system has achieved the Nash equilibrium. We show that there is a non-monotonic relationship between the degree of risk-aversion and system performance. Customers with an intermediate degree of riskaversion typically achieve higher sojourn times; in particular they rarely achieve the Nash equilibrium. Risk-neutral customers have the highest probability of achieving the Nash Equilibrium. Chapter 3 considers a service system similar to the previous one but with risk-neutral customers, and relaxes the assumption of exogenous service rates. In this sense, we model a queueing system with endogenous service rates by enabling managers to adjust the service capacity of the facilities. We assume that managers do so based on their perceptions of the arrival rates and use the same principle of adaptive expectations to model these perceptions. We consider service systems in which the managers' decisions take time to be implemented. Managers are characterised by a profile which is determined by the speed at which they update their perceptions, the speed at which they take decisions, and how coherent they are when accounting for their previous decisions still to be implemented when taking their next decision. We find that the managers' decisions exhibit a strong path-dependence: owing to the initial conditions of the model, the facilities of managers with identical profiles can evolve completely differently. In some cases the system becomes "locked-in" into a monopoly or duopoly situation. The competition between managers causes the weighted average sojourn time of the system to converge to the exogenous benchmark value which they use to estimate their desired capacity. Concerning the managers' profile, we found that the more conservative Summary xv a manager is regarding new information, the larger the market share his facility achieves. Additionally, the faster he takes decisions, the higher the probability that he achieves a monopoly position. In Chapter 4 we consider a one-server queueing system with non-captive customers. We carry out an experiment aimed at analysing the way human subjects, taking on the role of the manager, take decisions in a laboratory regarding the capacity of a service facility. We adapt the model proposed by van Ackere et al (2010). This model relaxes the assumption of a captive market and allows current customers to decide whether or not to use the facility. Additionally the facility also has potential customers who currently do not patronise it, but might consider doing so in the future. We identify three groups of subjects whose decisions cause similar behavioural patterns. These groups are labelled: gradual investors, lumpy investors, and random investor. Using an autocorrelation analysis of the subjects' decisions, we illustrate that these decisions are positively correlated to the decisions taken one period early. Subsequently we formulate a heuristic to model the decision rule considered by subjects in the laboratory. We found that this decision rule fits very well for those subjects who gradually adjust capacity, but it does not capture the behaviour of the subjects of the other two groups. In Chapter 5 we summarise the results and provide suggestions for further work. Our main contribution is the use of simulation and experimental methodologies to explain the collective behaviour generated by customers' and managers' decisions in queueing systems as well as the analysis of the individual behaviour of these agents. In this way, we differ from the typical literature related to queueing systems which focuses on optimising performance measures and the analysis of equilibrium solutions. Our work can be seen as a first step towards understanding the interaction between customer behaviour and the capacity adjustment process in queueing systems. This framework is still in its early stages and accordingly there is a large potential for further work that spans several research topics. Interesting extensions to this work include incorporating other characteristics of queueing systems which affect the customers' experience (e.g. balking, reneging and jockeying); providing customers and managers with additional information to take their decisions (e.g. service price, quality, customers' profile); analysing different decision rules and studying other characteristics which determine the profile of customers and managers.

Geological structure, recharge processes and underground drainage of a glacierised karst aquifer system, Tsanfleuron-Sanetsch, Swiss Alps

The relationships between stratigraphic and tectonic setting, recharge processes and underground drainage of the glacierised karst aquifer system `Tsanfleuron-Sanetsch' in the Swiss Alps have been studied by means of various methods, particularly tracer tests (19 injections). The area belongs to the Helvetic nappes and consists of Jurassic to Palaeogene sedimentary rocks. Strata are folded and form a regional anticlinorium. Cretaceous Urgonian limestone constitutes the main karst aquifer, overlain by a retreating glacier in its upper part. Polished limestone surfaces are exposed between the glacier front and the end moraine of 1855/1860 (Little Ice Age); typical alpine karrenfields can be observed further below. Results show that (1) large parts of the area are drained by the Glarey spring, which is used as a drinking water source, while marginal parts belong to the catchments of other springs; (2) groundwater flow towards the Glarey spring occurs in the main aquifer, parallel to stratification, while flow towards another spring crosses the entire stratigraphic sequence, consisting of about 800 m of marl and limestone, along deep faults that were probably enlarged by mass movements; (3) the variability of glacial meltwater production influences the shape of the tracer breakthrough curves and, consequently, flow and transport in the aquifer.

Influence of triamcinolone intravitreal injection on retinochoroidal healing processes.

To analyze the effects of triamcinolone intravitreal injection on the wound healing processes after argon laser retinal photocoagulation, wild type C57BL/6J mice, 8-12 weeks old underwent a standard argon laser photocoagulation protocol. After pentobarbital anesthesia and pupil dilatation, argon laser lesions were induced (50microm, 400mW, 0.05s). Two photocoagulation impacts created two disc diameters from the optic nerve in both eyes. The photocoagulated mice were divided into four groups: Group I (n=12), photocoagulation controls, did not receive any intravitreous injection. Group II (n=12), received an intravitreous injection of 1microl of balanced salt solution (BSS). Group III (n=12), received an intravitreous injection of 1microl containing 15microg of triamcinolone acetonide (TAAC) in BSS. Two mice from each of these three groups were sacrificed at 1, 3, 7, 14 days and 2 and 4 months after photocoagulation. Group IV (n=10) received 1.5, 3, 7.5, 15, or 30microg of TAAC and were all sacrificed on day 14. The enucleated eyes were subjected to systematic analysis of the cellular remodeling processes taking place within the laser lesion and its vicinity. To this purpose, specific antibodies against GFAP, von Willebrand factor, F4/80 and KI67 were used for the detection of astrocytes, activated Müller cells, vascular endothelial cells, infiltrating inflammatory cells and actively proliferating cells. TUNEL reaction was also carried out along with nuclear DAPI staining. Temporal and spatial observations of the created photocoagulation lesions demonstrate that 24h following the argon laser beam, a localized and well-delineated affection of the RPE cells and choroid is observed in mice in Groups I and II. The inner retinal layers in these mice eyes are preserved while TUNEL positive (apoptotic) cells are observed at the retinal outer nuclear layer level. At this stage, intense staining with GFAP is associated with activated retinal astrocytes and Müller cells throughout the laser path. From day 3 after photocoagulation, dilated new choroidal capillaries are detected on the edges of the laser lesion. These processes are accompanied by infiltration of inflammatory cells and the presence of proliferating cells within the lesion site. Mice in Group III treated with 15microg/mul of triamcinolone showed a decreased number of infiltrating inflammatory cells and proliferating cells, which was not statistically significant compared to uninjected laser treated controls. The development of new choroidal capillaries on the edges of the laser lesion was also inhibited during the first 2 months after photocoagulation. However, on month 4 the growth of new vessels was observed in these mice treated with TAAC. Mice of Group IV did not show any development of new capillaries even with small doses. After argon laser photocoagulation of the mouse eye, intravitreal injection of triamcinolone markedly influenced the retina and choroid remodeling and healing processes. Triamcinolone is a powerful inhibitor of the formation of neovessels in this model. However, this inhibition is transient. These observations should provide a practical insight for the mode of TAAC use in patients with wet AMD.

Standardizing admission and discharge processes to improve patient flow: A cross sectional study

Background: The aim of this study was to evaluate how hospital capacity was managed focusing on standardizing the admission and discharge processes. Methods: This study was set in a 900-bed university affiliated hospital of the National Health Service, near Barcelona (Spain). This is a cross-sectional study of a set of interventions which were gradually implemented between April and December 2008. Mainly, they were focused on standardizing the admission and discharge processes to improve patient flow. Primary administrative data was obtained from the 2007 and 2009 Hospital Database. Main outcome measures were median length of stay, percentage of planned discharges, number of surgery cancellations and median number of delayed emergency admissions at 8:00 am. For statistical bivariate analysis, we used a Chi-squared for linear trend for qualitative variables and a Wilcoxon signed ranks test and a Mann–Whitney test for non-normal continuous variables. Results: The median patients’ global length of stay was 8.56 days in 2007 and 7.93 days in 2009 (p<0.051). The percentage of patients admitted the same day as surgery increased from 64.87% in 2007 to 86.01% in 2009 (p<0.05). The number of cancelled interventions due to lack of beds was 216 patients in 2007 and 42 patients in 2009. The median number of planned discharges went from 43.05% in 2007 to 86.01% in 2009 (p<0.01). The median number of emergency patients waiting for an in-hospital bed at 8:00 am was 5 patients in 2007 and 3 patients in 2009 (p<0.01). Conclusions: In conclusion, standardization of admission and discharge processes are largely in our control. There is a significant opportunity to create important benefits for increasing bed capacity and hospital throughput.

The "handwriting brain": a meta-analysis of neuroimaging studies of motor versus orthographic processes.

INTRODUCTION: Handwriting is a modality of language production whose cerebral substrates remain poorly known although the existence of specific regions is postulated. The description of brain damaged patients with agraphia and, more recently, several neuroimaging studies suggest the involvement of different brain regions. However, results vary with the methodological choices made and may not always discriminate between "writing-specific" and motor or linguistic processes shared with other abilities. METHODS: We used the "Activation Likelihood Estimate" (ALE) meta-analytical method to identify the cerebral network of areas commonly activated during handwriting in 18 neuroimaging studies published in the literature. Included contrasts were also classified according to the control tasks used, whether non-specific motor/output-control or linguistic/input-control. These data were included in two secondary meta-analyses in order to reveal the functional role of the different areas of this network. RESULTS: An extensive, mainly left-hemisphere network of 12 cortical and sub-cortical areas was obtained; three of which were considered as primarily writing-specific (left superior frontal sulcus/middle frontal gyrus area, left intraparietal sulcus/superior parietal area, right cerebellum) while others related rather to non-specific motor (primary motor and sensorimotor cortex, supplementary motor area, thalamus and putamen) or linguistic processes (ventral premotor cortex, posterior/inferior temporal cortex). CONCLUSIONS: This meta-analysis provides a description of the cerebral network of handwriting as revealed by various types of neuroimaging experiments and confirms the crucial involvement of the left frontal and superior parietal regions. These findings provide new insights into cognitive processes involved in handwriting and their cerebral substrates.

Standardizing admission and discharge processes to improve patient flow: A cross sectional study

Background: The aim of this study was to evaluate how hospital capacity was managed focusing on standardizing the admission and discharge processes. Methods: This study was set in a 900-bed university affiliated hospital of the National Health Service, near Barcelona (Spain). This is a cross-sectional study of a set of interventions which were gradually implemented between April and December 2008. Mainly, they were focused on standardizing the admission and discharge processes to improve patient flow. Primary administrative data was obtained from the 2007 and 2009 Hospital Database. Main outcome measures were median length of stay, percentage of planned discharges, number of surgery cancellations and median number of delayed emergency admissions at 8:00¿am. For statistical bivariate analysis, we used a Chi-squared for linear trend for qualitative variables and a Wilcoxon signed ranks test and a Mann¿Whitney test for non-normal continuous variables. Results:The median patients' global length of stay was 8.56 days in 2007 and 7.93 days in 2009 (p<0.051). The percentage of patients admitted the same day as surgery increased from 64.87% in 2007 to 86.01% in 2009 (p<0.05). The number of cancelled interventions due to lack of beds was 216 patients in 2007 and 42 patients in 2009. The median number of planned discharges went from 43.05% in 2007 to 86.01% in 2009 (p<0.01). The median number of emergency patients waiting for an in-hospital bed at 8:00¿am was 5 patients in 2007 and 3 patients in 2009 (p<0.01). Conclusions: In conclusion, standardization of admission and discharge processes are largely in our control. There is a significant opportunity to create important benefits for increasing bed capacity and hospital throughput.

Quantification of magmatic and hydrothermal processes in a peralkaline syenite-alkali granite complex based on textures, phase equilibria, and stable and radiogenic isotopes

The Puklen complex of the Mid-Proterozoic Gardar Province, South Greenland, consists of various silica-saturated to quartz-bearing syenites, which are intruded by a peralkaline granite. The primary mafic minerals in the syenites are augite +/- olivine + Fe-Ti oxide + amphibole. Ternary feldspar thermometry and phase equilibria among mafic silicates yield T = 950-750degreesC, a(SiO2) = 0.7-1 and an f(O2) of 1-3 log units below the fayalite-magnetite-quartz (FMQ) buffer at 1 kbar. In the granites, the primary mafic minerals are ilmenite and Li-bearing arfvedsonite, which crystallized at temperatures below 750degreesC and at f(O2) values around the FMQ buffer. In both rock types, a secondary post-magmatic assemblage overprints the primary magmatic phases. In syenites, primary Ca-bearing minerals are replaced by Na-rich minerals such as aegirine-augite and albite, resulting in the release of Ca. Accordingly, secondary minerals include ferro-actinolite, (calcite-siderite)(ss), titanite and andradite in equilibrium with the Na-rich minerals. Phase equilibria indicate that formation of these minerals took place over a long temperature interval from near-magmatic temperatures down to similar to300degreesC. In the course of this cooling, oxygen fugacity rose in most samples. For example, late-stage aegirine in granites formed at the expense of arfvedsonite at temperatures below 300degreesC and at an oxygen fugacity above the haematite-magnetite (HM) buffer. The calculated delta(18)O(melt) value for the syenites (+5.9 to +6.3parts per thousand) implies a mantle origin, whereas the inferred delta(18)O(melt) value of <+5.1parts per thousand for the granitic melts is significantly lower. Thus, the granites require an additional low-delta(18)O contaminant, which was not involved in the genesis of the syenites. Rb/Sr data for minerals of both rock types indicate open-system behaviour for Rb and Sr during post-magmatic metasomatism. Neodymium isotope compositions (epsilonNd(1170 Ma) = -3.8 to -6.4) of primary minerals in syenites are highly variable, and suggest that assimilation of crustal rocks occurred to variable extents. Homogeneous epsilon(Nd) values of -5.9 and -6.0 for magmatic amphibole in the granites lie within the range of the syenites. Because of the very similar neodymium isotopic compositions of magmatic and late- to post-magmatic minerals from the same syenite samples a principally closed-system behaviour during cooling is implied. In contrast, for the granites an externally derived fluid phase is required to explain the extremely low epsilon(Nd) values of about -10 and low delta(18)O between +2.0 and +0.5parts per thousand for late-stage aegirine, indicating an open system in the late-stage history. In this study we show that the combination of phase equilibria constraints with stable and radiogenic isotope data on mineral separates can provide much better constraints on magma evolution during emplacement and crystallization than conventional whole-rock studies.

Characterization of polyphosphate synthesis by isolated yeast vacuoles

Polyphosphate (iPOP) is a linear polymer of orthophosphate units linked together by high energy phosphoanhydride bonds. It is found in all organisms, localized in organelles called acidocalcisomes and ranges from a few to few hundred monomers in length. iPOP has been found to play a vast array of roles in all organisms, including phosphate and energy metabolism, regulation of enzymes, virulence, pathogenicity, bone remodelling and blood clotting, among many others. Recently it was found that iPOP levels were increased in myeloma cells. The growing interest in iPOP in human cell lines makes it an interesting molecule to study. However, not much is known about its metabolism in eukaryotes. Acidocalcisomes are electron dense, acidic organelles that belong to the group of Lysosome Related Organelles (LROs). The conservation of acidocalcisomes among all kingdoms of life is suggestive of their important roles for the organisms. However, they are difficult to analyse because of limited biochemical tools for investigation. Yeast vacuoles present remarkable similarities to acidocalcisomes in terms of their physiological and structural features, including synthesis and storage of iPOP, which make them an ideal candidate to study biological processes which are shared between vacuoles and acidocalcisomes. The availability of tools for genetic manipulation and isolation of vacuoles makes yeast a candidate of choice for the characterization of iPOP synthesis in eukaryotes. Our group has identified the Vacuolar Transporter Chaperone (VTC) complex as iPOP polymerase and identified the catalytic subunit (Vtc4). The goal of my study was to characterize the process of iPOP synthesis by isolated vacuoles and to reconstitute iPOP synthesis in liposomes. The first step was to develop a method for monitoring iPOP by isolated vacuoles over time and comparing it with previously known methods. Next, a detailed characterization was performed to determine the modulators of the process, both for intact as well as solubilized vacuoles. Finally, attempts were made to purify the VTC complex and reconstitute it in liposomes. A parallel line of study was the translocation and storage of synthesized iPOP in the lumen of the vacuoles. As a result of this study, it is possible to determine distinct pools of iPOP- inside and outside the vacuolar lumen. Additionally, I establish that the vacuolar lysate withstands harsh steps during reconstitution on liposomes and retains iPOP synthesizing activity. The next steps will be purification of the intact VTC complex and its structure determination by cryo-electron microscopy. - Les organismes vivants sont composés d'une ou plusieurs cellules responsables des processus biologiques élémentaires tels que la digestion, la respiration, la synthèse et la reproduction. Leur environnement interne est en équilibre et ils réalisent un très grand nombre de réactions chimiques et biochimiques pour maintenir cet équilibre. A différents compartiments cellulaires, ou organelles, sont attribuées des tâches spécifiques pour maintenir les cellules en vie. L'étude de ces fonctions permet une meilleure compréhension de la vie et des organismes vivants. De nombreux processus sont bien connus et caractérisés mais d'autres nécessitent encore des investigations détaillées. L'un de ces processus est le métabolisme des polyphosphates. Ces molécules sont des polymères linéaires de phosphate inorganique dont la taille peut varier de quelques dizaines à quelques centaines d'unités élémentaires. Ils sont présents dans tous les organismes, des bactéries à l'homme. Ils sont localisés principalement dans des compartiments cellulaires appelés acidocalcisomes, des organelles acides observés en microscopie électronique comme des structures denses aux électrons. Les polyphosphates jouent un rôle important dans le stockage et le métabolisme de l'énergie, la réponse au stress, la virulence, la pathogénicité et la résistance aux drogues. Chez l'homme, ils sont impliqués dans la coagulation du sang et le remodelage osseux. De nouvelles fonctions biologiques des polyphosphates sont encore découvertes, ce qui accroît l'intérêt des chercheurs pour ces molécules. Bien que des progrès considérables ont été réalisés afin de comprendre la fonction des polyphosphates chez les bactéries, ce qui concerne la synthèse, le stockage et la dégradation des polyphosphates chez les eucaryotes est mal connu. Les vacuoles de la levure Saccharomyces cerevisiae sont similaires aux acidocalcisomes des organismes supérieurs en termes de structure et de fonction. Les acidocalcisomes sont difficiles à étudier car il n'existe que peu d'outils génétiques et biochimiques qui permettent leur caractérisation. En revanche, les vacuoles peuvent être aisément isolées des cellules vivantes et manipulées génétiquement. Les vacuoles comme les acidocalcisomes synthétisent et stockent les polyphosphates. Ainsi, les découvertes faites grâce aux vacuoles de levures peuvent être extrapolées aux acidocalcisomes des organismes supérieurs. Le but de mon projet était de caractériser la synthèse des polyphosphates par des vacuoles isolées. Au cours de mon travail de thèse, j'ai mis au point une méthode de mesure de la synthèse des polyphosphates par des organelles purifés. Ensuite, j'ai identifié des composés qui modulent la réaction enzymatique lorsque celle-ci a lieu dans la vacuole ou après solubilisation de l'organelle. J'ai ainsi pu mettre en évidence deux groupes distincts de polyphosphates dans le système : ceux au-dehors de la vacuole et ceux en-dedans de l'organelle. Cette observation suggère donc très fortement que les vacuoles non seulement synthétisent les polyphosphates mais aussi transfère les molécules synthétisées de l'extérieur vers l'intérieur de l'organelle. Il est très vraisemblable que les vacuoles régulent le renouvellement des polyphosphates qu'elles conservent, en réponse à des signaux cellulaires. Des essais de purification de l'enzyme synthétisant les polyphosphates ainsi que sa reconstitution dans des liposomes ont également été entrepris. Ainsi, mon travail présente de nouveaux aspects de la synthèse des polyphosphates chez les eucaryotes et les résultats devraient encourager l'élucidation de mécanismes similaires chez les organismes supérieurs. - Les polyphosphates (iPOP) sont des polymères linéaires de phosphates inorganiques liés par des liaisons phosphoanhydres de haute énergie. Ces molécules sont présentes dans tous les organismes et localisées dans des compartiments cellulaires appelés acidocalcisomes. Elles varient en taille de quelques dizaines à quelques centaines d'unités phosphate. Des fonctions nombreuses et variées ont été attribuées aux iPOP dont un rôle dans les métabolismes de l'énergie et du phosphate, dans la régulation d'activités enzymatiques, la virulence, la pathogénicité, le remodelage osseux et la coagulation sanguine. Il a récemment été montré que les cellules de myélome contiennent une grande quantité de iPOP. Il y donc un intérêt croissant pour les iPOP dans les lignées cellulaires humaines. Cependant, très peu d'informations sur le métabolisme des iPOP chez les eucaryotes sont disponibles. Les acidocalcisomes sont des compartiments acides et denses aux électrons. Ils font partie du groupe des organelles similaires aux lysosomes (LROs pour Lysosome Related Organelles). Le fait que les acidocalcisomes soient conservés dans tous les règnes du vivant montrent l'importance de ces compartiments pour les organismes. Cependant, l'analyse de ces organelles est rendue difficile par l'existence d'un nombre limité d'outils biochimiques permettant leur caractérisation. Les vacuoles de levures possèdent des aspects structuraux et physiologiques très similaires à ceux des acidocalcisomes. Par exemple, ils synthétisent et gardent en réserve les iPOP. Ceci fait des vacuoles de levure un modèle idéal pour l'étude de processus biologiques conservés chez les vacuoles et les acidocalcisomes. De plus, la levure est un organisme de choix pour l'étude de la synthèse des iPOP compte-tenu de l'existence de nombreux outils génétiques et la possibilité d'isoler des vacuoles fonctionnelles. Notre groupe a identifié le complexe VTC (Vacuole transporter Chaperone) comme étant responsable de la synthèse des iPOP et la sous-unité Vtc4p comme celle possédant l'activité catalytique. L'objectif de cette étude était de caractériser le processus de synthèse des iPOP en utilisant des vacuoles isolées et de reconstituer la synthèse des iPOP dans des liposomes. La première étape a consisté en la mise au point d'un dosage permettant la mesure de la quantité de iPOP synthétisés par les organelles isolés en fonction du temps. Cette nouvelle méthode a été comparée aux méthodes décrites précédemment dans la littérature. Ensuite, la caractérisation détaillée du processus a permis d'identifier des composés modulateurs de la réaction à la fois pour des vacuoles intactes et des vacuoles solubilisées. Enfin, des essais de purification du complexe VTC et sa reconstitution dans des liposomes ont été entrepris. De façon parallèle, une étude sur la translocation et le stockage des iPOP dans le lumen des vacuoles a été menée. Il a ainsi été possible de mettre en évidence différents groupes de iPOP : les iPOP localisés à l'intérieur et ceux localisés à l'extérieur des vacuoles isolées. De plus, nous avons observé que le lysat vacuolaire n'est pas détérioré par les étapes de reconstitution dans les liposomes et conserve l'activité de synthèse des iPOP. Les prochaines étapes consisteront en la purification du complexe intact et de la détermination de sa structure par cryo-microscopie électronique.

Regulation of dendritic development by neurotrophic factors

AbstractEstablishment of a functional nervous system occurs through an orchestrated multistep process during embryogenesis. As dendrites are the primary sites of synaptic connections, development of dendritic arborization is essential for the formation of functional neural circuits. Maturation of dendritic arbor occurs through dynamic processes that are regulated by intrinsic genetic factors and external signals, such as environmental stimuli, neuronal activity and growth factors. Among the latter, the neurotrophic factor BDNF is a key regulator of dendritic growth. However, the mechanisms by which BDNF controls dendritic development remain elusive.In this study, we first showed that activation of the MAPK signaling pathway and phosphorylation of the transcription factor CREB are required to mediate the effects of BDNF on dendritic development of cortical neurons. However, phosphorylation of CREB alone is not sufficient to induce dendritic growth in response to BDNF. Thus, by using a mutant form of CREB unable to bind its coactivator CRTC1, we demonstrated that BDNF-induced dendritic elaboration requires the functional interaction between CREB and CRTC1. Consistent with these observations, inhibition of CRTC1 expression by shRNA-mediated knockdown was found to suppress the effects of BDNF on dendritic length and branching of cortical neurons.The nuclear translocation of CRTC1, a step necessary for the interaction between CREB and CRTC1, was shown to result from the activation of NMD A receptors by glutamate, leading to the dephosphorylation of CRTC1 by the protein phosphatase calcineurin. In line with these findings, prevention of CRTC1 nuclear translocation in the absence of glutamate, or by inhibiting NMDA receptors or calcineurin suppressed the promotion of dendritic growth by BDNF.Increasing evidence supports a role for the growth factor HGF in the regulation of dendritic morphology during brain development. Despite these observations, little is known about the cellular mechanisms underlying the effects of HGF on dendritic elaboration of cortical neurons. The second part of this study was aimed at elucidating the cellular processes that mediate the effects of HGF on dendritic differentiation. We found that HGF increases cortical dendritic growth through mechanisms that involve MAPK-dependent phosphorylation of CREB, and interaction of CREB with its coactivator CRTC1. These data indicate that the mechanisms underlying the promotion of dendritic growth by HGF are similar to those that mediate the effects of BDNF, suggesting that the role of CREB and CRTC1 in the regulation of dendritic development may not be limited to HGF and BDNF, but may extend to other neurotrophic factors that control dendritic differentiation.Together, these results identify a previously unrecognized mechanism by which CREB and its coactivator CRTC1 mediate the effects of BDNF and HGF on dendritic growth of cortical neurons. Moreover, these data highlight the important role of the cooperation between BDNF/HGF and glutamate that converges on CREB to stimulate the expression of genes that contribute to the development of dendritic arborization.RésuméL'établissement d'un système nerveux fonctionnel s'accomplit grâce à des mécanismes précis, orchestrés en plusieurs étapes au cours de l'embryogenèse. Les dendrites étant les principaux sites de connexions synaptiques, le développement de l'arborisation dendritique est essentiel à la formation de circuits neuronaux fonctionnels. La maturation de l'arbre dendritique s'effectue grâce à des processus dynamiques qui sont régulés par des facteurs génétiques intrinsèques ainsi que par des facteurs externes tels que les stimuli environnementaux, l'activité neuronale ou les facteurs de croissance. Parmi ces derniers, le facteur neurotrophique BDNF est - connu pour être un régulateur clé de la croissance dendritique. Cependant, les mécanismes par lesquels BDNF contrôle le développement dendritique demeurent mal connus.Au cours de cette étude, nous avons montré dans un premier temps que l'activation de la voie de signalisation de la MAPK et la phosphorylation du facteur de transcription CREB sont nécessaires aux effets du BDNF sur le développement dendritique des neurones corticaux. Toutefois, la phosphorylation de CREB en tant que telle n'est pas sûffisante pour permettre la pousse des dendrites en réponse au BDNF. Ainsi, en utilisant une forme mutée de CREB incapable de se lier à son coactivateur CRTC1, nous avons démontré que l'élaboration des dendrites induite par le BDNF nécessite également une interaction fonctionnelle entre CREB et CRTC1. Ces résultats ont été confirmés par d'autres expériences qui ont montré que l'inhibition de l'expression de CRTC1 par l'intermédiaire de shRNA supprime les effets du BDNF sur la longueur et le branchement dendritique des neurones corticaux.Les résultats obtenus au cours de ce travail montrent également que la translocation nucléaire de CRTC1, qui est une étape nécessaire à l'interaction entre CREB et CRTC1, résulte de l'activation des récepteurs NMDA par le glutamate, entraînant la déphosphorylation de CRTC1 par la protéine phosphatase calcineurine. De plus, le blocage de la translocation nucléaire de CRTC1 en absence de glutamate, ou suite à l'inhibition des récepteurs NMDA ou de la calcineurine, supprime complètement la pousse des dendrites induite par le BDNF.De nombreuses d'évidences indiquent que le facteur de croissance HGF joue également un rôle important dans la régulation de la morphologie dendritique au cours du développement cérébral. Malgré ces observations, peu d'éléments sont connus quant aux mécanismes cellulaires qui sous-tendent les effets du HGF sur la croissance dendritique des neurones corticaux. Le but de la seconde partie de cette étude a eu pour but d'élucider les processus cellulaires responsables des effets du HGF sur la différenciation dendritique des neurones corticaux. Au cours de ces expériences, nous avons pu mettre en évidence que le HGF induit la pousse dendritique par des mécanismes qui impliquent la phosphorylation de CREB par la MAPK, et l'interaction de CREB avec son coactivateur CRTC1. Ces données indiquent que les mécanismes impliqués dans la stimulation de la croissance dendritique par le HGF sont similaires à ceux régulant les effets du BDNF, ce qui suggère que le rôle de CREB et de CRTC1 dans la régulation du développement dendritique n'est vraisemblablement pas limité aux effets du HGF ou du BDNF, mais pourrait s'étendre à d'autres facteurs neurotrophiques qui contrôlent la différenciation dendritique.En conclusion, ces résultats ont permis l'identification d'un nouveau mécanisme par lequel CREB et son coactivateur CRTC1 transmettent les effets du BDNF et du HGF sur la croissance dendritique de neurones corticaux. Ces observations mettent également en évidence le rôle important joué par la coopération entre BDNF/HGF et le glutamate, dans l'activation de CREB ainsi que dans l'expression de gènes qui participent au développement de l'arborisation dendritique des neurones corticaux.

Multiple processes regulate long-term population dynamics of sea urchins on Mediterranean rocky reefs

We annually monitored the abundance and size structure of herbivorous sea urchin populations (Paracentrotus lividus and Arbacia lixula) inside and outside a marine reserve in the Northwestern Mediterranean on two distinct habitats (boulders and vertical walls) over a period of 20 years, with the aim of analyzing changes at different temporal scales in relation to biotic and abiotic drivers. P. lividus exhibited significant variability in density over time on boulder bottoms but not on vertical walls, and temporal trends were not significantly different between the protection levels. Differences in densities were caused primarily by variance in recruitment, which was less pronounced inside the MPA and was correlated with adult density, indicating density-dependent recruitment under high predation pressure, as well as some positive feedback mechanisms that may facilitate higher urchin abundances despite higher predator abundance. Populations within the reserve were less variable in abundance and did not exhibit the hyper-abundances observed outside the reserve, suggesting that predation effects maybe more subtle than simply lowering the numbers of urchins in reserves. A. lixula densities were an order of magnitude lower than P. lividus densities and varied within sites and over time on boulder bottoms but did not differ between protection levels. In December 2008, an exceptionally violent storm reduced sea urchin densities drastically (by 50% to 80%) on boulder substrates, resulting in the lowest values observed over the entire study period, which remained at that level for at least two years (up to the present). Our results also showed great variability in the biological and physical processes acting at different temporal scales. This study highlights the need for appropriate temporal scales for studies to fully understand ecosystem functioning, the concepts of which are fundamental to successful conservation and management.

Limited agreement between body mass index, waist and body fat in diagnosing obesity in the Swiss population

Objective: to assess the agreement between different anthropometric markers in defining obesity and the effect on the prevalence of obese subjects. Methods: population-based cross-sectional study including 3213 women and 2912 men aged 35-75 years. Body fat percentage (%BF) was assessed using electric bioimpedance. Obesity was defined using established cut-points for body mass index (BMI) and waist, and three population-defined cut-points for %BF. Between-criteria agreement was assessed by the kappa statistic. Results: in men, agreement between the %BF cut-points was significantly higher (kappa values in the range 0.78 - 0.86) than with BMI or waist (0.47 - 0.62), whereas no such differences were found in women (0.41 - 0.69). In both genders, prevalence of obesity varied considerably according to the criteria used: 17% and 24% according to BMI and waist in men, and 14% and 31%, respectively, in women. For %BF, the prevalence varied between 14% and 17% in men and between 19% and 36% in women according to the cut-point used. In the older age groups, a fourfold difference in the prevalence of obesity was found when different criteria were used. Among subjects with at least one criteria for obesity (increased BMI, waist or %BF), only one third fulfilled all three criteria and one quarter two criteria. Less than half of women and 64% of men were jointly classified as obese by the three population-defined cut-points for %BF. Conclusions: the different anthropometric criteria to define obesity show a relatively poor agreement between them, leading to considerable differences in the prevalence of obesity in the general population.

Post-switching beta synchronization reveals concomitant sensory reafferences and active inhibition processes

It is known that post-movement beta synchronization (PMBS) is involved both in active inhibition and in sensory reafferences processes. The aim of this study was examine the temporal and spatial dynamics of the PMBS involved during multi-limb coordination task. We investigated post-switching beta synchronization (assigned PMBS) using time-frequency and source estimations analyzes. Participants (n = 17) initiated an auditory-paced bimanual tapping. After a 1500 ms preparatory period, an imperative stimulus required to either selectively stop the left while maintaining the right unimanual tapping (Switch condition: SWIT) or to continue the bimanual tapping (Continue condition: CONT). PMBS significantly increased in SWIT compared to CONT with maximal difference within right central region in broad-band 14âeuro"30 Hz and within left central region in restricted-band 22âeuro"26 Hz. Source estimations localized these effects within right pre-frontal cortex and left parietal cortex, respectively. A negative correlation showed that participants with a low percentage of errors in SWIT had a large PMBS amplitude within right parietal and frontal cortices. This study shows for the first time simultaneous PMBS with distinct functions in different brain regions and frequency ranges. The left parietal PMBS restricted to 22âeuro"26 Hz could reflect the sensory reafferences of the right hand tapping disrupted by the switching. In contrast, the right pre-frontal PMBS in a broad-band 14âeuro"30 Hz is likely reflecting the active inhibition of the left hand stopped. Finally, correlations between behavioral performance and the magnitude of the PMBS suggest that beta oscillations can be viewed as a marker of successful active inhibition.

Limited telomere shortening in hematopoietic stem cells after transplantation.

The number of cell divisions in hematopoietic stem cells (HSCs) following transplantation of bone marrow or mobilized peripheral blood into myelo-ablated recipients is unknown. This number is expected to depend primarily on the number of transplanted stem cells, assuming that stem cells do not differ in engraftment potential and other functional properties. In a previous study, we found that the telomere length in circulating granulocytes in normal individuals shows a biphasic decline with age, most likely reflecting age-related changes in the turnover of HSCs. In order to study HSCs' proliferation kinetics following stem cells transplantation, we analyzed the telomere length in donor-derived nucleated blood cells in four HLA-matched bone marrow transplant recipients relative to comparable cells from the sibling donors. In each case, the telomeres in granulocytes were shorter in the recipient than in the donor. This difference was established in the first year post transplantation and did not change after that. The telomere length in naïve and memory T cells showed marked differences after transplantation, complicating the interpretation of telomere length data using unseparated nucleated blood cells. Interestingly, the telomere length in naïve T cells that were first observed six months post transplantation was very similar in donor and recipient pairs. Our observations are compatible with a limited number of additional cell divisions in stem cell populations after bone marrow transplantations and support the idea that different populations of stem cells contribute to short-term myeloid and long-term lympho myeloid hematopoiesis.