I am interested in discovering the role of field potential oscillations in producing synchrony within the song system of the male zebra finch brain. An important function attributed to neural synchrony is sensorimotor integration. In the production of birdsong, sensorimotor integration is crucial, as auditory feedback is necessary for the maintenance of the song. A cortical-thalamic-cortical feedback loop is thought to play a role in the integration of auditory and motor information for the purpose of producing song. Synchronous activity has been observed between at least two nuclei in this feedback loop, MMAN and HVC. Since low frequency field potential oscillations have been shown to play a role in the synchronization of nuclei within the brain of other model animals, I hypothesized that this may be the case in the zebra finch song system. In order to investigate whether oscillatory activity is a mechanism behind the synchronous activity observed between HVC and MMAN, I performed dual extracellular recordings of neural activity within the zebra finch song system. Results suggest that oscillations are likely not involved in the synchrony observed in these nuclei. Future study may reveal that the structure of the feedback loop is necessary, and possibly even sufficient, for the synchronous activity in the zebra finch song system.

Sequences of stereotyped actions are central to the everyday lives of humans and animals, from the kingfisher's dive to the performance of a piano concerto. Lashley asked how neural circuits managed this feat nearly 6 decades ago, and to this day it remains a fundamental question in neuroscience. Toward answering this question, vocal performance in the songbird was used as a model to study the performance of learned, stereotyped motor sequences. The first component of this work considers the song motor cortical zone HVC in the zebra finch, an area that sends precise timing signals to both the descending motor pathway, responsible for stereotyped vocal performance in the adult, and the basal ganglia, which is responsible for both motor variability and song learning. Despite intense interest in HVC, previous research has exclusively focused on describing the activity of small numbers of neurons recorded serially as the bird sings. To better understand HVC network dynamics, both single units and local field potentials were sampled across multiple electrodes simultaneously in awake behaving zebra finches. The local field potential and spiking data reveal a stereotyped spatio-temporal pattern of inhibition operating on a 30 ms time-scale that coordinates the neural sequences in principal cells underlying song. The second component addresses the resilience of the song circuit through cutting the motor cortical zone HVC in half along one axis. Despite this large-scale perturbation, the finch quickly recovers and sings a near-perfect song within a single day. These first two studies suggest that HVC is functionally organized to robustly generate neural dynamics that enable vocal performance. The final component concerns a statistical study of the complex, flexible songs of the domesticated canary. This study revealed that canary song is characterized by specific long-range correlations up to 7 seconds long-a time-scale more typical of human music than animal vocalizations. Thus, the neural sequences underlying birdsong must be capable of generating more structure and complexity than previously thought.

Parmi les procédés de mise en forme de pièces industrielles, la métallurgie des poudres autorise une haute cadence de production avec une faible perte de matière première. L'élaboration de composants multi-matériaux par compression et frittage permet de minimiser le nombre d'étapes de conception afin de combiner des propriétés complémentaires. L'objet de cette thèse est l'étude d'un procédé de compression innovant qui peut augmenter la cadence de production et diminuer les contraintes sur l'outillage : la CGV (compression grande vitesse), et de l'appliquer à la mise en forme de pièces multimatériaux. Une presse à grande vitesse a été développée au laboratoire afin d'étudier l'influence de la CGV sur deux couples de matériaux : un couple base Fe / base WC, associant dureté et ténacité, un couple Acier 1.4313 / Stellite 6 associant résistance mécanique et résistance à la corrosion. Des modélisations numériques des procédés de compression conventionnelle et CGV ont été réalisées dans le but de mieux analyser les phénomènes observés et de prévoir le comportement en compression de plusieurs poudres simultanément. Ce travail aboutit à des recommandations pour la mise en oeuvre de la compression de composants multi-matériaux et met au jour quelques caractéristiques de la CGV. / Among industrial shape processes, powder metallurgy has a high production rate associated to low waste of materials. Manufacturing of multi-material components by die pressing and sintering is useful to reduce the production cycle in order to combine complementary properties. The aim of this thesis is to study an innovative compaction process which can increase production rate and reduce stresses applied on tools: the HVC (High Velocity Compaction), and use it to shape multi-material components. A HVC device has been developed in our laboratory to investigate the influence of HVC on two couples of powders : A couple Fe / WC which combines toughness and hardness properties, a couple 1.4313 steel/Stellite 6 with mechanical resistance associated to corrosion resistance. Numerical simulations of conventional and HVC compaction process have been achieved to analyze phenomenon and predict the behavior of multi-materials components during compaction. This work ends in recommendations for the implementation of the compression of multimaterials components and brings to light some characteristics of the CGV.

The developmental stress hypothesis predicts that an aversive condition, such as decreased food intake, predation, and social isolation, in the early developmental stage could have long term effects on behaviors and brain development of an animal. In nature, bird nestlings are susceptible to various factors, such as malnutrition, infections, and parasites. Effects of early life stress on adulthood have been extensively studied with some stressors including malnutrition. However, immune challenges as an early life stressor and their long-term programming effects on adult behaviors are yet to be studied in detail. The goal of the current study was to investigate changes in growth rate, personality, mate selection behaviors and brain development in zebra finch nestlings after injection with a viral infection mimicking agent, Polyinosinic: polycytidylic acid (Poly I:C). By using Poly I:C, it was possible to isolate long-term effects to the immune response of the bird. After Poly I:C injection on post-hatch day (PD) 14, morphological measures were conducted to detect changes in body growth rate. When birds became sexually mature (> ~PD 200), behaviors of birds were observed in different conditions to detect changes associated with the personality traits of animals. In mate choice trials, both attractiveness of males and mate selection behaviors of males and females were investigated. Finally, the development and neuronal activity of specific brain nuclei involved in courtship (i.e., HVC and RA) and social/sexual behaviors (nucleus taeniae of the amygdala, TnA) were investigated. The results showed that nestlings’ growth rate was not affected. However, Poly I:C injection had some effects on certain, but not all, personality traits observed in the study. Such effects were found only in female zebra finches, suggesting that there was a limited sex-specific influence of an early life immune challenge on personalities of adults. The results also showed that Control females tended to choose untreated males over Poly I:C injected males in mate choice trials. Finally, Poly I:C injection negatively affected the overall development of targeted brain nuclei. In addition, neuronal activity in TnA was higher in Poly I:C injected birds. Results of the present study suggest that one time injection with Poly I:C early in the life causes long term effects on adulthood. These findings are further discussed regarding their relevance to the developmental stress hypothesis.

Made available in DSpace on 2015-02-04T21:42:06Z (GMT). No. of bitstreams: 1 Maria Heloiza.pdf: 504743 bytes, checksum: 4fde6c28fc11e48070d8c696d00b78e9 (MD5) Previous issue date: 2009-10-23 / Este estudo epidemiológico foi realizado com o objetivo de definir a soroprevalência das hepatites virais B e C em escolares que freqüentavam regularmente as escolas municipais de Ensino Fundamental no município de Santos, no período de novembro de 2008 a março de 2009,. Foram analisadas 98 amostras de sangue de escolares com idade entre 4 e 14 anos, através da técnica de ELISA, para detecção da presença de anticorpos anti-HVC, anti-HBc anti-HBs e HBsAg, Setenta e duas amostras foram positivas para a detecção do anti-HBs, mostrando uma eficácia na vacinação contra hepatite B de 75%. Em duas crianças foi detectada a presença do HVC, demonstrando uma prevalência de 2,8%. A pesquisa do anti-HBc foi positiva em duas crianças (prevalência de 2,8%) e o HBs-Ag foi detectado em uma criança (prevalência de 1,3%).

This thesis presents an exploration of stars and gas in the halo of our Galaxy. A sample of 8321 field horizontal branch (FHB) stars was selected from the Hamburg/ESO Survey. The stars make excellent tracers of the Milky Way halo, and we studied the kinematics of a subset of the HES FHB stars, comparing their velocity dispersions to those predicted by several models. Since these stars are intrinsically luminous, hot and numerous they make ideal probes of the distances to high-velocity clouds (HVCs) - clouds of neutral hydrogen gas whose distances are largely unknown and which do not fit simple models of Galaxy rotation. A catalogue of stars which align with the HVCs was developed. High resolution spectroscopy of 16 such HVC probes with the Magellan telescope has yielded a remarkably tight distance constraint to complex WB. This is one of only a handful of such distance limits so far established. Lower distance limits were set for several other clouds. Finally, we have suggested that some of the HVCs may be associated with the accretion onto the MilkyWay of the Sagittarius dwarf galaxy.

Despite the trillions of dollars spent over the past decades on foreign aid 80% of humanity still lives on less than $10 dollars a day. There is an alarming need to deliver quality healthcare services and products to developing countries. The healthcare industry for developing countries is estimated to be $202 million and growing exponentially. However, intrinsic obstacles have prevented companies from fully deploying solutions in these countries. With the emergence of Citizen-Sector Organizations companies now have an alley to create High Value Chains enabling companies to deliver solutions to developing countries. Thereby, increasing shareholder value and increasing the living conditions of global citizens. As citizens of developing countries have better health care they are better equipped to succeed economically and consume other services and products the company has to offer. This paper discusses how an engineering manager can increase shareholder value by aligning corporate responsibility with the company’s strategic goals by leveraging High Value Chains. Specifically this paper discusses how open source methodologies can be utilized to improve healthcare in developing countries while increasing shareholder value. / text

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HCV genome is a positive sense single-stranded RNA containing a single open reading frame (ORF) flanked by untranslated regions (UTRs), 5’UTR and 3’UTR.Initiation of HCV RNA translation is mediated by internal ribosome entry site (IRES) present in 5’ UTR and this process is independent of cap-structure and requires only a small subset of canonical initiation factors. Hence, HCV IRES-mediated translation initiation mechanism is quite different from canonical cellular mRNA translation initiation. The IRES is organized into highly structured domains, namely domain II, III and IV. High affinity interactions between structured RNA elements present in the IRES and 40S ribosomal proteins mediate 40S recruitment to HCV IRES. However, details of the RNA elements and region of ribosomal proteins involved in these interactions are poorly understood. In recent days, RNA-based molecules like siRNAs, antisense RNAs and RNA decoys have become promising candidates for antiviral molecules. So designing short RNA molecules that target unique HCV translation initiation mechanism might help in developing novel anti-HCV molecules. HCV 3’UTR and antisense-5’ UTRs serve as sites for replication initiation to synthesize negative and positive strand and this process is catalyzed by NS5B protein (RNA-dependent RNA polymerase). Hence, host proteins binding to both 3’UTR and antisense-5’UTR might play important role in HCV replication. This puts the study of HCV RNA–host protein interactions and its role in viral translation and replication in perspective. Studying the HCV IRES-ribosomal protein S5 interactions and its role in HCV IRES function Previous studies from our laboratory have demonstrated that binding of La protein to GCAC close to initiator AUG enhances ribosomal protein S5 (RPS5) binding with HCV IRES and stimulates HCV translation. However in-detail study on HCV IRES–RPS5 interactions and its implication on HCV translation initiation were lacking. In present study computational modelling suggested that domain II and IV interact majorly with the beta hairpin structure and C-terminal helix of RPS5. Filter-binding and UV cross-linking studies with peptides derived from predicated RNA-binding region of RPS5 and mutational studies with RPS5 demonstrated that beta hairpin structure present in RPS5 is critical for IRES–RPS5 interaction. In parallel, we have studied RNA elements involved in the IRES–RPS5 interactions using deletions and substitution mutations, which we had generated on the basis of the computational model. Direct and competition UV cross-linking experiments performed with these IRES mutants and 40S subunits as a source of RPS5 suggested that structure and sequence of both domain II and IV play crucial role in IRES–RPS5 interactions. We further investigated the effect of these mutations on IRES activity by in vitro translation assay and found that all the mutants that were compromised in binding to RPS5 showed reduced IRES activity. Moreover, ribosome assembly experiments on HCV IRES demonstrated that mutations affecting IRES–RPS5 interactions result in reduction of 80S peak and slight increase of 48S peak. Since the 40S subunit had been previously reported to bind with HCV 3’UTR, we explored the possible interaction of RPS5 with HCV 3’UTR. From direct and competition UV cross-linking assays, we found that RPS5 does not bind to 3’UTR and the interaction is unique to IRES (5’UTR). Interestingly, partial silencing of RPS5 preferentially inhibited HCV translation with marginal effect on cap-dependent translation. Recently, reduction in 40S subunit abundance was reported to preferentially inhibit HCV translation. So, we investigated the abundance of free 40S subunit upon silencing RPS5 and results showed reduction in free 40S subunit level. So, we hypothesize that silencing of RPS5 reduces free 40S abundance to inhibit HCV translation. Taken together, results identified specific RNA elements present in HCV IRES that are critical for IRES–RPS5 interactions and demonstrated the role of these interactions in HCV translation initiation. Targeting ribosome assembly on HCV IRES using short RNAs Stem-loops (SL) IIIe and IIIf of HCV IRES are known to play an important role in stable IRES–40S complex formation. However interaction of these stem-loops with 40S subunit in isolation, independent of other regions of HCV IRES, was not studied. In this study, using electrophoretic mobility shift assay (EMSA) and sucrose gradient centrifugation experiments, we demonstrate that short RNA containing both SLIIIe and SLIIIf together (SLRef RNA) binds to 40S subunit, while short RNAs containing either of the stem-loops (SLRe RNA and SLRf RNA) lose their ability to interact with 40S subunit. Further, SLRef RNA inhibited ribosome assembly on the IRES, whereas SLRe and SLRf RNA failed to inhibit the same. Since SLRef RNA is derived from IRES, we investigated the interaction SLRef RNA with IRES–trans-acting factors (ITAFs). UV cross-linking of radio-labelled HCV IRES with cytoplasmic extract (S10) in presence of unlabelled short RNAs suggested possible interactions of La and RPS5 proteins with SLRef RNA. Studies with recombinant La protein and RPS5 further confirmed their interaction with SLRef RNA. Ex vivo experiments with HCV bicistronic RNA suggested that SLRef RNA specifically inhibits HCV translation. In addition to that SLRef RNA inhibited the HCV RNA synthesis in JFH1 HCV cell culture system. Moreover, specific delivery of pSUPER construct expressing SLRef RNA (pSUPERSLRef) to mice liver along with HCV bicistronic construct using Sendai virosomes demonstrated specific inhibition of HCV IRES activity by SLRef RNA in mice hepotocytes. In summary, short RNA derived from HCV IRES was shown to bind with La protein and RPS5 to inhibit ribosome assembly on HCV IRES. Further, targeted delivery of SLRef RNA into mice liver using Sendai virosome resulted in inhibition of HCV RNA translation in mice hepatocytes. Characterizing the interaction of host proteins with antisense-5’UTR and 3’UTR and its significance in HCV replication Antisense-5’UTR and 3’UTR of HCV RNA are the sites of replication initiation. Hence, host proteins binding to both of these RNA sequences are potential candidates for regulation of HCV replication. In this study, we have investigated host proteins binding with antisense-5’UTR and 3’UTRof HCV RNA by performing UV cross-linking experiments with cytoplasmic extract of Huh7 cells, and found that a protein of ~42kDa protein interacts with both antisense-5’UTR and 3’UTR. Based on earlier report, we predicted that the ~42kDa protein could be hnRNPC1/C2. Results of UV cross-linking followed by immuno pull-down (UV-IP assay) and UV cross-linking experiments with recombinant hnRNPC1 protein confirmed that hnRNPC1 indeed binds to antisense-5’UTR and 3’UTR. Further, filter-binding experiments demonstrated that hnRNPC1 protein binds to 3’UTR with higher affinity compared to antisense-5’UTR. Subsequently, we investigated the regions within 3’UTR and antisense-5’UTR that interact with hnRNPC1protein. Results demonstrated that poly-(U/UC) region of 3’UTR and region containing stem-loops SL-IIIa’, SL-IIIb’, SL-IIIcdef’ and SL-IV’ in antisense-5’UTR were mostly involved in the interaction. Interestingly, studies with confocal microscopy suggested that hnRNPC1/C2 re-localizes from nucleus to cytoplasm upon JFH1 infection, which might in turn influence HCV replication. To investigate the role of hnRNPC1/C2 in HCV replication, partial silencing of hnRNPC1/C2 was performed in HCV cell culture system (JFH1) and results demonstrated that hnRNPC1/C2 is critical for HCV RNA synthesis. However experiments with HCV bicistronic RNA suggested that hnRNPC1/C2 does not play significant role in HCV translation. Taken together, results suggested that hnRNPC1/C2 re-localizes from nucleus to cytoplasm upon JFH1 infection and binds to HCV 3’UTR and antisense- 5’UTR to regulate HCV replication. In summary, this thesis provides novel insights into the interaction of host proteins with HCV RNA and its significance in HCV translation and replication. Inhibition of the ribosome assembly and consequent reduction in HCV translation with mutations interfering with IRES–RPS5 interaction, reported in the present study, unfolds the novel role of this interaction in HCV translation. Further, results obtained in the present study with a small RNA SLRef, derived from HCV IRES, provide proof of concept for using short RNAs to specifically inhibit HCV translation. In addition, studies of interaction of hnRNPC1/C2 with HCV RNA and its re-localization upon HCV infection sheds light on the significance of host–virus interaction in viral RNA replication.