Thesis (Ph. D.)--University of Washington, 2003. / Vita. Includes bibliographical references (leaves 87-99).

Thesis (Ph. D.)--University of Washington, 2003. / Vita. Includes bibliographical references (leaves 67-80).

Diss. (sammanfattning) Umeå : Umeå universitet, 2009. / Härtill 4 uppsatser. Även tryckt utgåva.

We are interested in understanding the molecular mechanisms that govern synaptic growth and plasticity. Recent evidence from several laboratories suggests that small GTPases play an important role in the promotion of neurite outgrowth; however, their role in the control of synaptic growth and functional plasticity is not well understood. The goal of this thesis is to investigate the role of small GTPases (including Rac, Rho and Cdc42) in the regulation of synaptic growth in vivo, using the Drosophila larval neuromuscular junction (NMJ) synapses as a model system. Our results show that presynaptic overexpression of Rac, but not of Rho or Cdc42, positively regulates both synaptic structure and function. Genetic loss of Rac leads to embryonic lethality, making it impossible to assess the full loss-of-function phenotype using conventional mutants. To circumvent this, we use the MARCM (Mosaic Analysis with a Repressible Cell Marker) technique to generate single motor neuron clones devoid of all genetic copies of Rac. Our data suggest that Rac activity is crucial for normal synaptic development. In support of this conclusion, we demonstrate that genetic removal of trio, a guanine nucleotide exchange factor (GEF) that is known to activate Rac, leads to a drastic reduction in the number of synaptic boutons. In addition, genetic removal of one copy of trio is sufficient to suppress the gain-of-function phenotype of Rac. Moreover, we demonstrate that partial removal of the fragile X mental retardation gene (dfmr1), a known suppressor of Rac, enhances the gain-of-function phenotype of Rac. Taken together, our findings support a model in which Rac signaling positively regulates synaptic growth and function at the Drosophila larval NMJ.

Morphologic and metabolic change is a known property of the adult brain. A number of behavioural tasks alter local cerebral blood flow and glucose utilisation. The expression of the glucose transporter 1 (GLUT1), which allows the entry of glucose to the brain, also has been shown to change in response to long-lasting neuronal activation. However, little is known about the effect of acute neuronal activation on GLUT1 expression. Using immunohistochemistry and Western blot, we investigated cerebral GLUT1 expression and vasculature density in mice undergoing a 48-hour voluntary wheel running period. The results showed that the striatum was the main region where GLUT1 protein was up-regulated: There was a trend for GLUT1 expression and blood vessels density to be associated with the distance run during the experiment. These results indicate that short-term increased neuronal activation is associated with rapid changes in glucose transport and possibly vascular remodelling.

Thesis (Ph.D. in Neuroscience) -- University of Colorado at Denver and Health Sciences Center, 2006. / Typescript. Includes bibliographical references (leaves 86-96). Free to UCDHSC affiliates. Online version available via ProQuest Digital Dissertations;

Ce travail consiste à étudier l’expression, le rôle et l’échange des microARNs (miARNs), dans le système nerveux (SN). Les microARNs (miARNs) sont des petits ARNs endogènes non-codants qui exercent une régulation négative sur l’expression desgènes, en s’hybridant sur la région 3’ non-codante des ARNm cibles.Dans un premier, nous avons dévoilé le rôle spécifique du miR-92a, dans lecontrôle de l’expression de la sous-unité GluA1 des récepteurs AMPA, dans un paradigme de plasticité synaptique homéostatique, dans lequel la plasticité synaptique est inhibée.Nous avons ensuite montré, par RNA-Seq que les miARNs sont différentiellement exprimés entre les structures du SN. Cette technologie nous a aussi permis de révéler la présence de nouvelles espèces de miARNs. De plus les résultats suggèrent que l’expression des miARNs (connus et nouveaux) participe à la signature transcriptomique singulière de chacune des structures.Dans un troisième temps, notre travail montre que les miARNs peuvent être échangés entre les cellules du système nerveux via les vésicules extracellulaires (EVs). Le contenu des EVs en miARNs varie en fonction de l’activité neuronale, et ces derniers ont pour cibles prédictives des protéines impliquées dans la régulation de la plasticité neuronale. Nos résultats suggèrent donc que l’échange de miARNs via les EVs est un nouveau mécanisme de modulation de la plasticité neuronale.Enfin, nous proposons un nouvel outil de purification des EVs, qui à l’avenir permettrait de purifier les EVs du SNC selon leur origine cellulaire.Pour conclure, ce travail apporte une meilleure compréhension du rôle des miARNs dans la régulation de la physiologie du SNC. / This work consists in stuying the expression, the role and the transport of microRNAs (miRNAs) in the central nervous system (CNS). microRNAs (miRNAs) are small endogenous non coding RNAs, exerting a negative regulation on gene expression.They inhibit protein translation by hybridization on the 3’ untranslated region of mRNA.First, we have revealed the specific role of miR-92a in the control of the expressionof GluA1, in an homeostatic plasticity paradigm in which the synaptic plasticity is inhibited.Second, by using RNA-Seq technology, we showed that miRNAs are differentially expressed in the different structures of the CNS. Moreover, we have discovered new species of miRNAs. Finally, our results suggest that the miRNA expression (of known and new miRNAs) participate in the singular transcriptomique signature of each structure.Third, we have shown that miRNAs are transported into EVs, and can be exchanged between the cells of the CNS. The miRNA content of EVs varies depending on neuronal activity. Target prediction of these miRNAs includes genes involved in the regulation of neuronal plasticity. Together, our results suggest that the exchange of miRNAs through EVs is a new mechanism involved in the modulation of neuronal plasticity. Finally, we propose a new tool for purifiying EVs depending on their cellular origin.To conclude, this study allows a better understanding of the role of miRNAs in the regulation of the physiology of the CNS.

Made available in DSpace on 2016-06-02T20:19:22Z (GMT). No. of bitstreams: 1 5748.pdf: 1975827 bytes, checksum: e6ad26dbbd3bf2dd7360047fe5b4b42b (MD5) Previous issue date: 2014-02-19 / Financiadora de Estudos e Projetos / Background: The stroke is the leading cause of adult disability in the world. One of the main complaints of individuals post-stroke refers to the loss of function of the upper limb, as evidenced during the performance of activities of daily living. This difficulty may be related to an important component of sensorimotor control, joint position sense, a submodality of proprioception. The relationship of sense of joint and the difficulties of upper limb movements are not clear in the literature. Objectives: To investigate whether the joint position sense of both shoulders of chronic hemiparetic is altered during the abduction and flexion, and whether there is any correlation between joint position sense and sensorimotor performance in these subjects. Methods: The study included 13 subjects with chronic hemiparesis due to ischemic stroke and 13 healthy subjects matched for gender and age. The proprioception was assessed using an isokinetic dynamometer and by joint sense position. The variable calculated was absolute error for shoulder abduction and flexion at the 30° and 60°. The motor performance was assessed by the upper extremity of the Fugl-Meyer Motor Assessment Scale. For the comparison between paretic, nonparetic and control limbs was utilized Kruskal Wallis test following post-hoc of Mann-Whitney with Bonferroni adjustment. For the correlation were used the Spearman. Results: No difference was found between the paretic and non-paretic limbs in both movements at 30o and 60o (p>0.05). But higher values of absolute error for both paretic and nonparetic limbs compared to control were observed during abduction at 30o (p <0.01) and at 60o (p=0.03 and p=0.04, respectively). During flexion, higher values of absolute error was observed in paretic limbs compared to control at 30o (p=0.01) and also at 60o (p=0.02). Non paretic limbs presented higher values of absolute error only at 30o, when compared to control (p=0.01). Total absolute error was inversely correlated to sensibility and proprioception Fugl-Meyer Motor Assessment Subscales (EFM). Absolute error also was directly correlated right hemisphere injury. Furthermore, absolute error was to shoulder´s subluxation during abduction. Conclusion: The joint position sense is affected in both shoulders of chronic post-stroke ischemic hemiparetic subjects during flexion and abduction. These proprioceptive deficits of the paretic limb are correlated to subscale score of proprioception and sensibility EFM, shoulder´s subluxation and damaged hemisphere. / O Acidente Vascular Cerebral (AVC) é a principal causa de incapacidade em adultos no mundo. Uma das principais queixas de indivíduos pós- AVC refere-se à perda da função do membro superior, evidenciado durante a realização de atividades da vida diária. Essa pode estar relacionada a um componente importante do controle sensório-motor, o senso de posição articular, uma submodalidade da propriocepção. A relação do senso de articular e as dificuldades de movimentos do membro superior ainda não estão claras na literatura. Objetivos: Investigar se o senso de posição articular de ambos os ombros de hemiparéticos crônicos está alterada durante a abdução e flexão, e se há correlação entre o senso de posição articular, o desempenho sensório-motor e o lado de lesão. Métodos: O estudo incluiu 13 indivíduos com hemiparesia crônica decorrente de AVC isquêmico e 13 indivíduos saudáveis pareados por sexo e idade. A propriocepção foi avaliada através do senso de posição articular em um dinamômetro. A variável calculada foi erro absoluto para abdução e flexão do ombro a 30° e 60°. O desempenho motor foi avaliado Escala de Avaliação de Fugl-Meyer (EFM) para membros superiores. Para a comparação entre o membro parético, não parético e controle foi utilizado o teste de Kruskal Wallis seguinte post-hoc de Mann-Whitney com ajuste de Bonferroni. Para a correlação foi utilizado a Correlação de Spearman. Resultados: Não foi encontrada diferença entre os membros parético e não parético em ambos os movimentos, a 30o e 60o (p>0,05). Mas, foram observados altos valores de erro absoluto para ambos os membros, parético e não parético, comparado ao controle durante a abdução a 30o (p<0,01) e a 60o (p=0,03 e p=0,04, respectivamente). Durante a flexão, foram constatados altos valores absolutos no membro parético comparado ao controle a 30° (p=0,01) e também a 60° (p=0,02). O membro não parético apresentou altos valores de erro absoluto somente a 30°, comparado ao controle (p=0,01). O erro absoluto do membro parético foi inversamente correlacionado com a pontuação da subescala de sensibilidade e propriocepção da EFM. O erro absoluto foi diretamente correlacionado com a lesão do hemisfério direito. Além disso, o erro absoluto do membro parético durante a abdução foi correlacionado com a subluxação do ombro. Conclusão: O senso de posição articular está alterado em ambos os ombros de indivíduos hemiparéticos crônicos pós-AVC isquêmico durante a flexão e abdução. Esses déficits proprioceptivos do membro parético estão correlacionados com a pontuação da subescala de propriocepção e sensibilidade da EFM, lesão do hemisfério e subluxação do ombro.

O exercício físico traz inúmeros benefícios para o sistema nervoso, dentre eles a melhora da memória e cognição, além de um efeito protetor em relação ao declínio mental decorrente do envelhecimento e de lesões do sistema nervoso. Este estudo teve como objetivo observar os efeitos plásticos do exercício moderado de curta duração no hipocampo e em regiões motoras do encéfalo de ratos, frequentemente afetadas por lesões ou doenças neurodegenerativas. As metodologias empregadas nestas análises foram a imuno-histoquímica, o Western blotting,o PCR em tempo real, avaliação dos níveis de neurogênese pela injeção de BrdU e imageamento de Ca2+ de astrócitos corticais. Os resultados encontrados mostram que o exercício moderado de curta duração promoveu alterações plásticas específicas em todas as regiões estudadas, variando na dependência do marcador utilizado e do decurso temporal do exercício. Acreditamos que este é suficiente para promover plasticidade difusa no sistema nervoso, que pode ser parte do substrato do efeito benéfico do exercício no sistema nervoso. / Evidence shows that physical exercise is neuroprotective and enhances brain function by improving cognition, learning and memory. It has also been associated with structural changes such as angiogenesis, synaptogenesis and neurogenesis. The aim of this study was to observe the effects of a moderate, short-term exercise protocol on the hippocampus and brain regions related to motor function, commonly affected by neurodegenerative diseases. The methods used for these analyses were immunohistochemistry, Western blotting, real-time PCR, evaluation of the levels of hippocampal neurogenesis with injections of BrdU and Ca2+ imaging of cortical astrocytes. Our results show that short-term moderate physical exercise induced specific plastic changes in all regions studied, which varied depending on the marker and time course of exercise and is enough to modulate synaptic and structural elements of neurons as well as astrocytes, playing an important role in the diffuse exercise-dependent plasticity which may underlie the beneficial effects of exercise in the brain.

Introdução: A perda auditiva na infância, mesmo perdas auditivas mínimas, pode dificultar ou atrasar a aquisição de linguagem da criança. Quanto mais tardio for o diagnóstico e o início da intervenção, maiores serão os efeitos da privação sensorial na via auditiva. Em crianças usuárias de aparelho de amplificação sonora individual (AASI), a utilização dos Potenciais Evocados Auditivos de Longa Latência (PEALL) torna-se uma ferramenta de verificação capaz de mensurar a maturação do sistema nervoso auditivo central (SNAC) ao longo do tempo de uso da amplificação. Objetivo: Caracterizar os potenciais evocados auditivos de longa latência (PEALL) em crianças com perda auditiva neurossensorial pré e pós-adaptação do aparelho de amplificação sonora individual (AASI). Metodologia: Estudo longitudinal constituído por 32 sujeitos e dividido em dois grupos: grupo estudo e grupo controle. O grupo estudo foi composto por 18 crianças, sendo cinco do gênero feminino e 13 do gênero masculino, com idade entre sete e 12 anos (média de idade: 9 anos e 2 meses), com perda auditiva neurossensorial bilateral simétrica de grau leve a moderado, sem experiência prévia com qualquer tipo de amplificação. Todas as crianças foram adaptadas com AASI bilateral após a primeira avaliação eletrofisiológica para a captação dos PEALL (componentes P1, N1, P2, N2 e P300) com estímulo de fala e estímulo tone burst, sendo que esta avaliação se repetiu 3 meses e 9 meses após o uso do AASI. O grupo controle foi composto por 14 crianças, sendo seis do gênero feminino e 8 do gênero masculino, com idade entre sete e 12 anos (média de idade: 9 anos e 8 meses). Todas as crianças deste grupo apresentaram audição normal e a avaliação para obtenção dos PEALL foi realizada respeitando o mesmo intervalo de tempo do grupo estudo: M0 - avaliação inicial; M3 e M9 - avaliação após três e nove meses da avaliação inicial. Resultados: Na comparação dos valores de latência e amplitude com o estímulo tone-burst no GE (condição sem AASI), observou-se diferença estatisticamente significante entre os três momentos de avaliação (M0xM3xM9) para a latência do P1 na orelha esquerda (OE) e latência do P300 na orelha direita (OD), com diminuição da latência ao longo do tempo. Na comparação dos momentos de avaliação dois a dois, para o componente P1 a diferença significante deu-se entre os momentos M0xM9 (p-valor=0,022), e para o componente P300 entre M0xM3 (p-valor=0,013). Com relação ao estímulo de fala pode-se observar uma diminuição estatisticamente significante nas latências dos componentes P2 (p-valor=0,010) e N2 (p-valor=0,007) (OE) entre os momentos M3 e M9. Quanto à presença e ausência dos componentes dos PEALL com estímulo de fala, verificou-se ausência dos componentes P1 e N1 no momento M3 assim como para N1, P2 e P300 no momento M9. Com estímulo tone burst observou-se ausência de respostas para os componentes N1 e P2, nos diferentes momentos de avaliação. No que diz respeito à correlação entre o tempo de privação sensorial e os componentes do PEALL obtidos com estímulo tone-burst na OD (condição com AASI) observou-se correlação estatisticamente significante entre tempo de privação sensorial e amplitude do P300 (p-valor=0,006), sendo que quanto maior o tempo de privação sensorial, menor a amplitude do P300 (r=-0,655). Com relação à correlação entre a frequência de uso do AASI e os componentes dos PEALL observou-se correlação estatisticamente significante entre frequência de uso do AASI e amplitude P2-N2 (p-valor=0,033) para o estímulo de fala na condição com AASI na OE. Conclusão: O PEALL demonstrou ser uma ferramenta clínica viável na avaliação de crianças usuárias de AASI, permitindo monitorar e mensurar a plasticidade neuronal do Sistema Nervoso Auditivo Central após um período de estimulação auditiva / Introduction: Hearing loss in childhood, even when very small, can hinder or even delay the process of language acquisition. The effects of sensory deprivation in the auditory pathway worsen, as diagnosis and the beginning of intervention are delayed. The use of Long Latency Auditory Evoked Potentials (LLAEP) in children that use individual hearing aids becomes a scanning tool capable of measuring the central auditory nervous system\'s (CANS) maturation throughout the period of use of the hearing aid. Purpose: To characterize the long latency auditory evoked potentials (LLAEP) in children with sensorineural hearing loss before and after fitting of hearing aids. Methodology: Longitudinal study composed of 32 subjects and divided into 2 groups: study group and control group. The study group was composed of 18 children, of these 5 were female and 13 were male, with ages between 7 and 12 years (age average: 9 years and 2 months), with mild to moderate bilateral symmetrical sensorineural hearing loss, with no previous experience using any kind of hearing aid. All of the children were fitted with bilateral hearing aids after the first electrophysiological assessment in order to record the LLAEP (components P1, N1, P2, N2 and P300), with speech and tone-burst stimuli; this assessment was repeated 3 and 9 months after the fitting of the hearing aids. The control group was composed of 14 children, of these 6 were female and 8 were male, with ages between 7 and 12 years (age average 9 years and 8 months). All of the children in this group showed normal hearing and the assessments to record the LLAEP were carried out in the same intervals of time as the study group: A0 - initial assessment; A3 and A9 - assessments 3 and 9 months after the initial assessment. Results: When comparing the latency and amplitude results with the tone-burst stimulus in the study group (SG) (without hearing aids), a statistically significant difference between the 3 assessments (A0xA3xA9) was observed for the P1 latency in the left ear (LE) and the P300 latency in the right ear (RE), with a decrease in latency with the passing of time. When comparing the assessments two by two, for the P1 component, the significant difference was recorded between A0xA9 (p-value=0,022), and for the P300 component, the significant difference was recorded between A0xA3 (p-value=0,013). Regarding the speech stimulus, a statistically significant decrease can be observed in the latency of the components P2 (p-value=0,010) and N2 (p-value=0,007) (LE) between A3 and A9. As for the presence and absence of the LLAEP components with the speech stimulus, an absence of the components P1 and N1 at A3 was observed, as well as for N1, P2 and P300 at A9. An absence of response was observed with the tone-burst stimulus for the N1 and P2 components, at the different times of assessment. As for the correlation between the period of sensory deprivation and the LLAEP components obtained from the tone-burst stimulus in the RE (with hearing aids), a statistically significant correlation was observed between the period of sensory deprivation and the amplitude of the P300 (p-value=0,006) - where there was a higher sensory deprivation time, the amplitude of the P300 was lower (r=-0,655). As for the correlation between the frequency of the use of hearing aids and the components of the LLAEP, a statistically significant relationship was observed between the frequency of use of hearing aids and the P2-N2 amplitude (p-value=0,033) for the speech stimulus when using hearing aids in the LE. Conclusion: The LLAEP proved to be a viable clinical tool in the assessment of children using hearing aids, allowing for the monitoring and measuring of the neural plasticity of the Central Auditory Nervous System after a period of hearing stimulation