Orientador: Fernando Cendes / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciencias Médicas / Made available in DSpace on 2018-08-06T21:33:07Z (GMT). No. of bitstreams: 1 Coan_AnaCarolina_M.pdf: 12048096 bytes, checksum: d15d096a9718e3a561f6bbab24d2755e (MD5) Previous issue date: 2006 / Resumo: Introdução: Inúmeros fatores influenciam a recidiva de crises epilépticas em pacientes em remissão submetidos à retirada da droga anti-epiléptica (DAE). O tipo de síndrome epiléptica e a pesença de anormalidades estruturais são consideradas variáveis importantes. Porém, nenhum estudo demonstrou a influência de alterações em exames de ressonância magnética (RM) no prognóstico de reciva de crises destes pacientes. Objetivo: Determinar a presença e o grau de atrofia hipocampal (AH) e a presença de hipersinal hipocampal em pacientes há pelo menos dois anos livres de crises, que foram submetidos à retirada da DAE, e correlacionar a AH e a hiperintensidade do sinal hipocampal com a recorrência de crises. Métodos: De um grupo de 99 pacientes com epilepsia parcial acompanhados em um protocolo para retirada da DAE após o controle adequado das crises, foi realizado o estudo volumétrico da estrutura hipocampal nos 84 pacientes que puderam ser submetidos a exame de Ressonância Magnética (RM). Todos os pacientes foram acompanhados por um longo período (5,7 a 11 anos). Os volumes hipocampais (VHc) foram determinados em imagens coronais T1-IR de 3 mm através do software NIH, com correção pelo volume intra-cranial total de cada paciente. A intensidade do sinal hipocampal foi determinada através da relaxometria de T2 duplo-eco (realizada em 57 pacientes) através do software NIH. A AH foi caracterizada tanto pelo VHc como pelo índice de assimetria hipocampal (IAHc, razão do menor lado/maior lado) abaixo de dois desvios-padrão da média do grupo controle. A hiperintensidade de sinal foi determinada pelo prolongamento do tempo de T2 pelo menos dois desvios-padrão acima da média do grupo controle. Resultados: Um total de 50/84 pacientes (59,5%) apresentaram recorrência de crises após a retirada da DAE. A AH esteve presente em 39/84 (46%). A presença de AH se relacionou significativamente a uma maior freqüência de recorrência de crises (29/39; 74%) após a retirada da DAE em comparação com os pacientes sem AH (21/45; 47%) (x2, p=0,01) A análise de sobrevivência (Kaplan-Meier) demonstrou uma diferença significativa de recorrência de crises entre pacientes com ou sem AH (Mantel, p=0,006). A probabilidade estimada de permanecer livre de crises 5 anos após a retirada da DAE foi 62% para aqueles sem AH e aproximadamente 28% para aqueles com AH. Relaxometria de T2 com sinal anormal foi mais freqüente em pacientes com recorrência de crises (10/31; 32%) do que naqueles que permaneceram livres de crise (3/26, 11%), apesar desta diferença não ser significativa, (x2, p=0,1) devido ao tamanho da amostra. No entanto, a análise de sobrevivência (Kaplan-Meier) demonstrou uma diferença significativa de recorrência de crises entre pacientes com ou sem sinal anormal à relaxometria de T2 (Tarone-Ware, p= 0,013). A probabilidade estimada de permanecer livre de crises 5 anos após a retirada da DAE foi 62% para aqueles sem sinal hipocampal anormal e aproximadamente 23% para aqueles com sinal hipocampal anormal à relaxometria de T2. Conclusões: A recorrência de crises após a retirada da DAE foi mais frequente entre os pacientes com AH ou sinal hipocampal anormal. A avaliação de imagens de RM deve ser levada em consideração antes da decisão da retirada da DAE em pacientes com epilepsias parciais / Abstract: Background: Various factors influence the proportion of patients with epilepsy who were previously seizure-free under medication, and will present seizure recurrence after antiepileptic drug (AED) withdrawal. Epilepsy syndrome and presence of structural abnormalities are considered important variables. Objective: To determine the presence and degree of hippocampal atrophy (HA) and hyperintense hippocampal signal in patients who were seizure-free for at least two years and underwent AED withdrawal, and to correlate HA and hyperintense hippocampal signal with seizure recurrence. Methods: From a group of 99 patients with partial epilepsy followed in a protocol for AED withdrawal after seizure control, we performed hippocampal volumetric study in 84 patients with available high resolution MRI All patients were followed up for a long period of time. Hippocampal volumes (HcV) were determined using 3 mm Tl-IR coronal images using NIH software, with correction by the variation of total intracranial volumes. Signal intensity of hippocampal structure was determined by double-echo T2 relaxometry (obtained for 57 patients) using NTH software HA was determined for either HcV or hippocampal asymmetry index (HcAI, smaller/larger ratio) below two standard deviations from the mean of the control group. Signal hyperintensity was determined for prolonged T2 relaxation time above two standard deviations from the mean of control group. Results: A total of 50/84 patients (59.5%) had seizure recurrence after AED withdrawal HA was present in 39/84 (46%). The presence of HA was associated with a significant higher frequency (29/39, 74%) of seizure recurrence after AED withdrawal compared to those without HA (21/45; 47%) (x2, p=0.01). Survival analysis (Kaplan-Meier) demonstrated a significant difference of seizure recurrence between patients with or without HA (Mantel, p= 0 006). The estimated probability of remaining seizure free 5 years after AED withdrawal was 62% for those without HA and approximately 28% for those with HA. Abnormal T2 relaxometry was more frequent in patients with seizure recurrence (10/31; 32%) than in those who remained seizure free (3/26; 11%) although this difference was not significant (x2, p=0.1) due to sample size. Survival analysis (Kaplan-Meier) demonstrated a significant difference of seizure recurrence between patients with or without abnormal T2 relaxometry (Tarone-Ware, p= 0.013) The estimated probability of remaining seizure free at 5 years after AED withdrawal was 62% for those without abnormal hippocampal signal and approximately 23% for those with abnormal hippocampal signal. Conclusions: Seizure recurrence after AED withdrawal was more frequent among patients with HA and abnormal hippocampal T2 signal MRI evaluation should be considered before decision for AED discontinuation in seizure-free patients with partial epilepsies / Mestrado / Neurociencias / Mestre em Fisiopatologia Médica

A administração sistêmica da peçonha bruta da cascavel Sul-Americana (Crotalus durissus terrificus, Cdt) induz prejuízos de memória espacial em ratos, sugerindo a ocorrência de dano hipocampal. No presente estudo, foram utilizadas técnicas In vivo e In vitro, a fim de testar os potenciais efeitos centrais do veneno de Cdt e da crotoxina (CTX), principal neurotoxina componente da peçonha bruta da serpente. Foram investigados (1) efeitos comportamentais da administração sistêmica e intrahipocampal do veneno de Cdt e da CTX envolvendo diferentes versões do labirinto aquático de Morris; (2) a morte neuronal, o curso temporal e a ação protetora do soro anticrotálico à exposição de culturas organotípicas de hipocampo a doses crescentes (0,05-1 μg/mL) do veneno bruto de Cdt e de CTX, por meio da coloração fluorescente por Iodeto de propídeo (PI) (que reflete a morte neuronal); e (3) alguns dos possíveis mecanismos fisiopatológicos do envenenamento experimental em culturas de hipocampo utilizando o antagonista de receptores AMPA de glutamato, NBQX, à administração de 0,05 μg/mL do veneno bruto de Cdt, para avaliar a possível participação do glutamato e seus receptores na morte neuronal. A administração sistêmica e intrahipocampal do veneno bruto de cascavel em ratos promoveu marcados prejuízos de memória espacial de referência e operacional. Diferentemente, a CTX promoveu prejuízos comportamentais apenas quando a administração foi intrahipocampal. As culturas expostas ao veneno bruto apresentaram dano sobretudo nas células piramidais de CA1, enquanto as culturas incubadas com CTX não apresentaram danos. Essa seletividade em relação ao CA1 foi mais proeminente nas doses baixas. O soro anticrotálico preveniu a morte neuronal quando administrado até uma hora após a exposição do veneno. O NBQX preveniu parcialmente o dano em CA1 indicando a participação de receptores AMPA de glutamato na cascata de eventos que subjazem os danos a estas células causados pela peçonha bruta de Cdt. Estes achados podem contribuir para a tanto para elucidação dos mecanismos quanto para a conduta utilizada durante o envenenamento crotálico / Systemic administration of the South American rattlesnake (Crotalus durissus terrificus, Cdt) venom in rats permanently disrupts performance in hippocampus-dependent spatial memory tasks, thus indicating the occurrence of damage to this brain area. In vivo and In vitro approaches were employed in the present study to investigate the effects of the venom and its main compound, crotoxin (CTX). We investigated (1) behavioral effects after both systemic and intrahipocampal administration of either venom or CTX in rats on performance in different versions of the Morris\' water maze task; (2) neuronal death, evaluated by the intensity of propidium iodide (PI) fluorescence (neurotoxicity of the venom and of CTX, and the protection by the antivenom, on organotypic hippocampal slice cultures, were evaluated by comparing slices exposed to venom and (1) slices treated only with vehicle - negative control - and (2) slices incubated with 1 mM glutamate - positive control for maximal neuronal death), and the time course of neurotoxic effects of 0.05-1 μg/mL of crude Cdt venom and CTX, and the Brazilian anticrotalid serum on organotypic hippocampal slice cultures; and (3) provide an initial screening of the mechanisms underlying the venom action on cultured hippocampal slices, using the AMPA receptor antagonist NBQX and the crude venom at 0.5 μg/mL dose, to test whether the neuronal damage is mediated by glutamate following the experimental envenomation. Both systemic and intrahippocampal administration of Cdt venom induces permanent disruption of spatial reference memory and spatial working memory in rats. In contrast, CTX only promoted behavioral effects when administered intrahippocampally. Slices exposed to Cdt venom, but not to CTX, exhibited substantial neuronal damage. This effect was particularly prominent in the CA1 sub-field at lower concentrations of the venom. The antivenom prevented damage when applied until 1 hour after the exposure of the venom. The NBQX partially prevented damage to CA1, thus indicating that AMPA receptors play a role in the damage caused by the Cdt venom. These findings may be helpful in the elucidation and management of rattlesnake envenomation

O presente estudo avaliou (1) a memória espacial de ratas Wistar expostas a diferentes formas de experiência maternal [primiparidade (PRIM), sensibilização maternal pela exposição a filhotes adotivos (SENS) e primiparidade com privação de contato com os filhotes por 6 horas diárias (PRIV), em comparação com a nuliparidade (NUL)] em tarefas espaciais de memórias de referência e operacional no labirinto aquático de Morris. Os resultados revelaram ausência de diferenças quando os animais são avaliados após o desmame dos filhotes, enquanto que o período de contato com os mesmos resultou em comportamentos relacionados à ansiedade em ratas SENS. Adicionalmente, ratas PRIV exibiram melhor desempenho no teste de memória operacional, possivelmente devido a um efeito de enriquecimento ambiental pela exposição intermitente e contínua aos filhotes durante o período de lactação. Ainda, comparou- se (2) o desempenho de ratas PRIM e NUL das linhagens Wistar (W) e Sprague-Dawley (SDW) nas mesmas tarefas comportamentais no labirinto aquático. As ratas PRIM-SDW exibiram melhores desempenhos em relação às NUL-SDW e também em relação às PRIM-W no teste de memória de referência, enquanto que no teste de memória operacional, as ratas PRIM-SWD exibiram um desempenho superior em relação às PRIM-W. Adicionalmente, ratas SWD, independentemente da experiência maternal, exibiram estratégias de busca mais eficientes para cumprir a tarefa. Por fim, (3) avaliou-se o desempenho de ratas PRIM e NUL da linhagem SWD expostas a um ambiente enriquecido (AE) em comparação com ratas PRIM e NUL mantidas em gaiolas comuns de laboratório (STD) em tarefas de localização e reconhecimento de objetos, bem como as taxas de neurogênese hipocampal desses animais. Os resultados mostraram que as ratas PRIM-AE foram mais sensíveis à alteração na disposição espacial de um objeto familiar em comparação com as NUL-AE. Adicionalmente, as ratas NUL e STD exibiram mais comportamentos relacionados com ansiedade e estresse. Os resultados anatômicos mostraram que a experiência maternal e a exposição ao AE por 45 dias não geraram alterações na neurogênese hipocampal em ratas SWD. Estes resultados mostram que diferentes formas de experiência maternal exercem alterações distintas sobre o comportamento de ratas, de forma dependente do momento em que os animais são avaliados e da linhagem de ratas utilizadas. Adicionalmente, mostram que o AE gera alterações de memória espacial e produz efeitos ansiolíticos, particularmente em ratas PRIM. / Maternal experience in rats induces changes in brain and behavior. This study compared (1) spatial memory performances of primiparous (PRIM), pup-induced maternal behaviors (SENS), 6h/daily pup-deprived primiparous (DEP) and nuliparous (NUL) Wistar rats in the reference and working memory versions of the Morris water maze task. No differences were found when the animals were tested after pup\'s weaning/exposure. On the other hand, lactation/pup exposure period induced anxiety-like behaviors in SENS rats when tested during this period. In addition, DEP rats showed better performances in the working memory task, which may be an \"environmental enrichment effect\" due the intermittent exposure to the offspring during lactation. We also compared (2) spatial performances of PRIM and NUL rats from 2 different strains, e, Wistar (W) and Sprague-Dawley (SDW) in the same tasks described in the first experiment. The results showed better performances of PRIM-SDW groups relative to both NUL-SWD and PRIM-W in the reference memory task, while PRIM-SWD outperformed PRIM-W rats in the working memory task. Additionally, SWD rats, regardless their reproductive status, showed better performance in relation to their search strategies to find the hidden platform. Lastly, we (3) evaluated performances of PRIM and NUL Sprague-Dawley rats exposed to an enriched environment (EE) compared to NUL and PRIM rats housed in standard laboratory cages (STD) in an object placement and object recognition task. All groups were injected with BrdU in order to assess hippocampal cell proliferation, differentiation/migration, and cell survival in these animals. In the place object task, PRIM-AE rats exhibited better performances compared to NUL-AE rats. In addition, NUL and STD rats showed more anxiety and stress-related behaviors. The anatomical data showed no differences among all groups, indicating that enriched environment in a regimen of 45 days exposure or maternal experience had no influence on the hippocampal neurogenesis in Sprague-Dawley rats. Taken together, these data show that different forms of maternal experience in rats induce different effects on behavior, in a time and strain-dependent manner. Also, the results showed that exposure to an enriched environment induced spatial memory alterations and anxyolitic effects, mainly in PRIM rats.

Orientador: Elenice Aparecida de Moraes Ferrari / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-24T19:42:43Z (GMT). No. of bitstreams: 1 Canova_Fernando_D.pdf: 2567655 bytes, checksum: 90b1604e178613eecc7fb38539152db1 (MD5) Previous issue date: 2014 / Resumo: A memória espacial tem importante valor para a sobrevivência do organismo. Está relacionada com sua habilidade em utilizar informações que possibilitam a formação de mapas espaciais de acordo com a relevância das relações entre as pistas ambientais. A memória espacial envolve alterações plásticas no hipocampo de roedores e de aves, dentre essas a indução da transcrição de genes de expressão imediata, tais como o zenk, que regula a expressão de inúmeros outros genes e proteínas. O gene zenk atua na regulação da expressão da proteína Sinapsina. A expressão de sinapsinas também parece estar relacionada com mecanismos sinápticos mediados por receptores GABAB. O presente trabalho teve como objetivo avaliar, em pombos, a aprendizagem, a consolidação e a persistência da memória espacial em situação de escolha alimentar. A plasticidade neural foi investigada pela análise da imunorreatividade para as proteínas Zenk e Sinapsina I no hipocampo. Foram realizados dois experimentos com pombos (Columba livia): No Experimento I foi avaliada a aprendizagem e memória espacial em animais submetidos ao treino de escolha alimentar em 2 ou 7 sessões de treino. Cada sessão teve 6 tentativas experimentais, numa arena circular com 4 comedouros onde apenas um tinha alimento. A resposta de escolha foi definida como orientar-se, aproximar-se e bicar um comedouro. Foi aplicado também o teste de estratégia espacial, que consistiu em 1 sessão, com 3 tentativas experimentais onde na terceira tentativa foram retirados os comedouros e os animais foram colocados na arena circular para análise do tempo de permanência no quadrante onde antes existia o comedouro com alimento. No Experimento II foram avaliados os efeitos do antagonista do receptor GABAB, faclofeno administrado (i.p) imediatamente após as sessões de treino. O teste de persistência da memória espacial foi realizado 7 dias após o treino. A análise imunoistoquímica foi utilizada para avaliação da expressão das proteínas Zenk e Sinapsina I no hipocampo. O número de escolhas corretas aumentou em função do treino, variando entre 50 e 88% na primeira e sétima sessões respectivamente (ANOVA, p < 0,001). O número de núcleos Zenk - positivos e células Sinapsina I - positivas aumentou no hipocampo de pombos treinados durante 7 sessões (ANOVA, p < 0,001). O tratamento com o inibidor do receptor GABAB teve um efeito facilitador nas sessões de treino em comparação com os grupos controles (ANOVA, p < 0,001). Os dados indicam que a experiência em escolha espacial desencadeia a ativação de mecanismos sinápticos que resultam na expressão de Zenk e Sinapsina I no hipocampo de pombos, os quais estão envolvidos na consolidação da memória espacial de escolha alimentar. Os efeitos do tratamento com faclofeno sugerem que o receptor GABAB participa destes mecanismos. Em conjunto os presentes dados demonstraram que o pombo é um modelo experimental importante para a análise de mecanismos de plasticidade neural no hipocampo que são conservados entre espécies de mamíferos e aves / Abstract: The spatial memory has important value for the organism's survival, because it is related with ability to acquire and use space information, enabling the formation of cognitive and spatial maps. The experience with the environment triggers behavioral, cellular and molecular changes in central nervous system that an essential to the formation and consolidation of spatial memory. Therefore, acquisition, consolidation and stability of spatial memory involve plastic changes in the hippocampus of rodents and birds, among these the induction of the expression of early gene transcription such as zenk gene which regulates the expression of numerous other genes and the proteins they control. The gene zenk acts in the formation of new memories and has a role in the regulation of the synapsin protein expression. The expression of synapsins also seems to be related with synaptic mechanisms mediated by the GABAB receptor. This study aimed to evaluate, in pigeons, consolidation and persistence of spatial memory in a situation of food location. The underlying neural plasticity was investigated through the analysis of immunoreactivity for Zenk and Synapsin I proteins in neurons of the hippocampus. Adult, male pigeons (Columba livia) were used in two experiments. Experiment I investigated learning and spatial memory during short (2 sessions) or long duration training (7 sessions). Sessions had 6 experimental trials and were conducted in a arena with 4 feeders. In each trial the bird was released at a different point of the arena, and the time between the release of the bird and the first feeder pecking response (latency of the choice response) was recorded. Testing for spatial strategy was conducted in the arena without any feeder and the time spent in each quadrant was analyzed. Experiment II analyzed the effects of the post-training administration of phaclofen on the consolidation and persistence of spatial memory. Testing of memory persistency occurred 7 days after the 7th training session. Immunohistochemistry of hippocampal tissue was used for the analysis of expression of Zenk and Synapsin I proteins. Number of correct choice increased across training, with values varying between 50 and 88% in the first and 7th session respectively (Anova, p < 0.001). Treatment with GABABantagonist had a facilitatory effect on choice performance which was indicated by lower latency values and higher accuracy values (Anova < 0.001). The values of Zenk positive and Synapsin I positive cells counting were higher in the hippocampus of pigeons that were trained during 7 days and in pigeons treated with phaclofen as compared to their respective controls (Anova p < 0.001). These data indicated that experience with spatial learning of food choice induced expression of Zenk and Synapsin I proteins in the hippocampus of pigeons during the consolidation of spatial memory. Post-training administration of phaclofen suggested a role of GABAB receptor in these experience-dependent synaptic mechanisms. The present results point to the pigeon as interesting animal model for the analysis of neuroplastic mechanisms involved with spatial memory, which are conservative across mammalian and birds species / Doutorado / Fisiologia / Doutor em Biologia Funcional e Molecular

Orientadoesr: Patrícia Aline Boer, José Antônio Rocha Contijo / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-21T03:17:34Z (GMT). No. of bitstreams: 1 Oliveira_AgnesdaSilvaLopes_M.pdf: 2796027 bytes, checksum: 870f793968de3a84abd8d4b40ecd4034 (MD5) Previous issue date: 2012 / Resumo: Estudos têm demonstrado que a deficiência de nutrientes durante a gravidez ou nos primeiros anos de vida pós-natal resultam em anormalidades estruturais no hipocampo da prole, bem como comprometimento cognitivo em animais com 16 semanas de vida. Na tentativa de analisar se a restrição protéica gestacional pode induzir a déficits de aprendizagem e perda de memória associada a alterações estruturais no hipocampo, realizou-se teste de water maze (MWM) e uma análise detalhada morfométrica da citoarquitetura dendrítica do hipocampo de ratos machos adultos. Além disso, analisamos no hipocampo dorsal e ventral a expressão e localização de receptores de mineralocorticóides (MR) e glicocorticóides (GR), angiotensina II receptor tipo 1 (AT1) e receptores específicos de serotonina (5HT1A e 5HT2A). No MWM não foram encontradas diferenças significativas entre os grupos LP e NP, em qualquer um dos parâmetros analisados no teste de memória espacial, sugerindo que tais funções de hipocampo não foram alteradas com a restrição proteica gestacional. No entanto, através da aplicação da técnica de Gogi-Cox realizamos a reconstrução dendrítica nos neurônios do hipocampo dorsal. Nossos resultados demonstram que a restrição proteica gestacional leva a uma diminuição do comprimento dos dendritos basais e no número de intersecções dos dendritos apicais de CA3. A citoarquitetura de CA1 e do giro denteado não foi alterada. O presente estudo revelou uma clara dissociação entre a resposta do teste comportamental e alterações de neurônios do hipocampo, como conseqüência da programação fetal. Encontramos diferentes padrões de expressão dos receptores analisados no hipocampo dorsal e ventral o que sugere que redução na expressão de GR e 5HT1A paralelamente a maior expressão de 5HT2A estão envolvidos no comportamento ansioso e que a significativa diminuição na expressão de AT1 pode ter um efeito protetor. Essas alterações neuroquímicas podem ter consequências importantes para comportamento de ansiedade e depressão. Nosso estudo não é capaz de responder se as alterações encontradas estão relacionadas com subdesenvolvimento no útero ou resulta de uma adaptação pós-natal para a fisiologia programada na vida adulta. Outros estudos devem ser feitos para responder essas questões / Abstract: Studies have demonstrated that maternal nutritional restriction during pregnancy or in early postnatal life results in hippocampus cognitive impairment and structural abnormalities in the 16-wk-old offspring. In an attempt to analyze whether gestational protein restriction might induce learning and memory impairment associated with structural changes in the hippocampus we carried out MWM test and a detailed morphometric analysis of dendritic cytoarchitecture of the hippocampus from male adult rats. In addition, we analyzed the dorsal and ventral hippocampal expression and localization of mineralo- (MR) and glucocorticoid (GR), type 1 angiotensina II receptor (AT1) and serotonin specific receptors (5HT1A and 5HT2A). By MWM we did not found significant differences between LP and NP groups, in any of the parameters analyzed, suggesting that such functions of hippocampus were not altered by gestational protein restriction. However, by applying 3-dimensional analysis of dendrites from the dorsal hippocampus, this study demonstrates that gestational protein restriction leads to decreases in total basal dendritic length and inapical intersections of CA3 pyramidal neurons. The dendritic architecture of CA1 and dentate gyros was unchanged. The current study revealed a clear dissociation between behavioral test response and hippocampal neuron changes as consequence of fetal programming. We found different patterns of dorsal and ventral expression of analyzed receptors and we suggests that reduced GR and 5HT1A and enhanced 5HT2A expression are involved in anxious behavior and that AT1 down regulation may has a protective effect. These neurochemical alterations may have important consequences for anxiety- and depressive-like behavior. Our study is not able to answer the question whether these alterations are related to in uteri underdevelopment or results from a postnatal adaptation to programmed physiology in adult life. Further time-course studies should be done to answer this question / Mestrado / Clinica Medica / Mestre em Clinica Medica

Orientadores: Li Li Min, Fernando Cendes / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-04T03:09:51Z (GMT). No. of bitstreams: 1 Bonilha_LeonardoFatorGouvea_D.pdf: 8420238 bytes, checksum: 016f0b30e07800517df57c5fc2672c09 (MD5) Previous issue date: 2004 / Resumo: A esclerose hipocampal (EH) é a alteração histológica mais comum em pacientes com epilepsia do lobo temporal medial (ELTM). A Ressonância Magnética (RM) de crânio possibilita a detecção in vivo de sinais associados à EH, permitindo que pacientes com EL TM reftatária à medicação sejam submetidos à ressecção cirúrgica do hipocampo para tratamento de crises epilépticas. As causas de reftatariedade à medicação e ao tratamento cirúrgico ainda são desconhecidas, porém supõe-se que um dos motivos seja a presença de lesão neuronal acometendo outras áreas cerebrais além do hipocampo. O uso da morfometria por RM permite avaliação do dano neuronal tanto no hipocampo como em outras estruturas cerebrais através da avaliação e quantificação da atrofia presente nestas estruturas. Para avaliação pormenorizada das estruturas cerebrais foi realizada a implementação e validação de um protocolo anatômico para mensuração da região mesial do lobo temporal, com uso de RM tridimensional de alta definição. Foi também definido um protocolo para volumetria automatizada baseada em voxel de todo o cérebro. Foi observado que o dano neuronal em pacientes com EL TM se estende além do hipocampo e acomete regiões que se conectam funcionalmente e anatomicamente ao hipocampo. Tál achado sugere que exista lesão abrangendo uma rede neuronal, o que pode ser responsável em conjunto pelas manifestações clínicas observadas nesses pacientes / Abstract: Hippocampal sclerosis (HS) is the most common histological finding in patients with media! temporal lobe epilepsy (MTLE). Magnetic resonance imaging (MRl) permits in vivo detection of signs that are associated to HS, permitting the surgical treatment for these patients. The causes of medical and surgical reftactoriness observed in patients with MTLE are still unknown. One possible explanation is the fact that the neuronalloss encountered in these patients spans over other brain areas beyond the hippocampus. The use of morphometric quantification of brain structures through MRI is a powerful tool to investigate the neuronalloss in the hippocampus and in other areas of the brain. In order to assess the neuronal damage in brain structures of patients with MTLE, we developed a protocol for manual MRI morphometry of the media! temporallobe structures. We also developed an automatic protocol to assess the concentration of gray matter in the whole brain of these patients through the use of Voxel Based Morphometry. We observed that patients with MTLE exhibit neuronal loss that is not restricted to the hippocampus, but affects di:fferent areas throughout the brain that are functionally and anatomica1ly connected to the hippocampus. These findings suggest that a lesion of a network of neural structures may be responsible for the clinical symptomatology exhibited by patients with MTLE / Doutorado / Neurologia / Doutor em Ciências Médicas

Ischemic stroke causes irreversible damage to the brain. The hippocampus is a vulnerable region and plays an important role in cognition and locomotor activity. Puerarin is a phytoestrogen that has beneficial effects in treating neurological disorders. How puerarin protects against hippocampal injury and its molecular mechanisms remain to be elucidated. Transient global brain ischemia was induced by 4-vessel occlusion in adult male Sprague-Dawley rats. The rats were pretreated with puerarin alone or together with LY294002 (an PI3K inhibitor) before ischemia/ reperfusion (I/R). The open-and closed-field tasks and Morris water maze (MWM) test were used to assess the effects of puerarin on anxiety-like behavioral and cognitive impairment following I/R. Hematoxylin-eosin staining(HE) was used to examine the survival of hippocampal CA1 pyramidal neurons, and immunoblotting was performed to examine the expression of the related proteins. By using the rat model for transient I/R, we demonstrated that puerarin pretreatment significantly increased the travelling distance and number of crossings in the open-and closedfield tests, reduced latency and increased the proportion of distance and time in zone IV in the MWM. The number of live cells in the hippocampus is sharply increased by puerarin pretreatment. We further observed that the levels of phosphorylated Akt1, GSK-3 beta and MCL-1were elevated and those of cleaved-caspase-3 were reduced in the puerarin-treatment group. Notably, the PI3K inhibitor LY294002 counteracted all of the effects of puerarin. Our findings suggest that puerarin protects the hippocampus from I/R damage by activating the PI3K/Akt1/GSK-3 beta/MCL-1 signaling pathway.

The main aim of this thesis was to assess if early deprivation (ED) and glucocorticoid (GC) treatment exert long-term effects on the volume of the brain regions implicated in responses to stress, and if it associates with alterations in the distribution and structure of astroglia, which are known to support brain plasticity. This study also investigated the effects of prenatal dexamethasone (Dex) treatment on selected brain receptors, namely the oxytocin and 5-HT1A receptors, as they are implicated in the regulation of responses to stress. In addition, in vitro effects of Dex on neural stem cells were studied, in order to explore the drug effects on cell proliferation and differentiation, and on glial cell markers. Unbiased stereological estimation was employed to determine the regional brain volume, astroglial morphology and total cell count. Peripheral quantitative computed tomography (pQCT) technique was used to quantify total brain volume. Autoradiography technique was employed to visualise and analyse oxytocin and 5HT-1A serotonin receptor binding using selective radioactive ligands. The results of the present study demonstrate that both ED and prenatal Dex exposure leads to long-term effects on hippocampal remodelling with volume losses and impoverished astroglial morphology in the form of reduced primary process length. The observed deterioration in astroglial morphology adds further evidence that astrocytic changes contribute to hippocampal volume losses, a phenomenon that deserves more research in the context of effects of corticosteroid overload and stress-related pathologies. The present results also demonstrate that prenatal Dex induces long-term effects at the level of central neuroregulatory processes. Thus significant region- and sex-dependent reductions or increases in the oxytocin and 5-HT1A receptor binding were observed. The in vitro study has shown that Dex affects both proliferation and differentiation of GFAP positive NSCs with no toxic effects as such. Overall, both early postnatal or prenatal manipulations that increase levels of stress and/or glucocorticoids as the chemical mediators of stress, lead to a long-term maladaptive brain remodelling with losses in the hippocampal volume, impoverishment of hippocampal astroglial morphology and changes in the properties of central regulatory receptors in the brain areas involved in the reaction to stress.

Alzheimer´s disease (AD) is an age-related, irreversible and progressive neurodegenerative pathology that deteriorates cognitive function including learning and memory. AD is characterised neuropathologically by the presence of neuritic plaques (Aβ), neurofibrillary tangles (NFTs), synaptic loss and neuronal death. AD affects specific brain regions involved in mnestic function such as the neocortex and the hippocampus. The dorsal (DR) and the median raphe (MR) nuclei give rise to serotonergic (5-HT) projections that innervate multiple brain regions including the cortex and the hippocampus, playing an important role in learning and memory processes. For a long time the degeneration of cholinergic (ACh) system was considered as the main neurochemical changes in AD brains, however, more recent studies highlight the involvement of other neurotransmitter systems including 5-HT. This thesis entitled “Altered serotonergic neurotransmission as a main player in the pathophysiology of Alzheimer’s disease: structural and ultrastructural studies in a triple transgenic mouse model of the disease” demonstrates that there exist specific alterations in the serotonergic projections of the hippocampus during the progression of AD using the triple transgenic (3xTg-AD) mouse model of the disease, which closely resemble human AD. Mr. Harun N. Noristani is submitting this thesis to the University of Manchester for the degree of PhD in the Faculty of Life Science. The results obtained in this thesis show for the first time a biphasic increase in serotonergic fibre sprouting in the 3xTg-AD mouse model of AD that occurs in parallel with evident intraneuronal/extracellular Aβ deposition in the hippocampus (Chapter 3). In addition, serotonergic fibre sprouting correlated with reduced perforated synapses in the hippocampus, suggesting a structural remodeling process to maintain hippocampal connectivity (Chapter 4). Increased 5-HT neurotransmission (via high dietary intake of tryptophan, 5-HT precursor) reduced intraneuronal Aβ accumulation in the hippocampus, suggesting a direct role of 5-HT neurotransmission in modifying AD neuropathology (Chapter 5). Given the protective role of increased 5-HT neurotransmission, treatment with 5-HT enhancing drugs may be beneficial in reducing the underlying pathology as well as improving the behavioural and cognitive abnormalities associated with AD. Nevertheless, the role of specific 5-HT receptors responsible for such neuro-protective effect of 5-HT in AD awaits further research.

The perirhinal cortex (PER), which is critical for associative memory and stimulus discrimination, has been described as a wall of inhibition between the neocortex and hippocampus. With advanced age, rats show deficits on PER-dependent behavioral tasks and fewer PER principal neurons are activated by stimuli, but the role of PER interneurons in these altered circuit properties in old age has not been characterized. In the present study, PER neurons were recorded while rats traversed a circular track bidirectionally in which the track was either empty or contained eight novel objects evenly spaced around the track. Putative interneurons were discriminated from principal cells based on the autocorrelogram, waveform parameters, and firing rate. While object modulation of interneuron firing was observed in both young and aged rats, PER interneurons recorded from old animals had lower firing rates compared with those from young animals. This difference could not be accounted for by differences in running speed, as the firing rates of PER interneurons did not show significant velocity modulation. Finally, in the aged rats, relative to young rats, there was a significant reduction in detected excitatory and inhibitory monosynaptic connections. Together these data suggest that with advanced age there may be reduced afferent drive from excitatory cells onto interneurons that may compromise the wall of inhibition between the hippocampus and cortex. This circuit dysfunction could erode the function of temporal lobe networks and ultimately contribute to cognitive aging.