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

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.

M.A. (Clinical Psychology) / Attention deficit hyperactivity disorder (ADHD) is a form of psychopathology characterised by difficulties with hyperactivity, attention and behavioural inhibition. Although ADHD has been historically considered a disorder specific to children, the contemporary consensus among researchers is that, in some cases, ADHD may persist into adulthood. Neurologically, ADHD is associated with deficits in the executive functions, located in the frontal lobe. Cortical arousal, which refers to the level of neuronal activity in the cerebral cortex and is measurable using electroencephalograph machinery (EEGs), is usually lower in the frontal lobes in those with ADHD when compared to individuals without the disorder. With regards to anxiety disorders, which are dysfunctional variations in the normal fear response, the aetiology of the pathologies in this category are multifaceted and complex. At the neurological level, however, there is a clear link between anxiety disorders and hypervigilance, which is characterised by high cortical arousal in the frontal lobes. Despite the fact that, at face value, it seems impossible for an increase and decrease in cortical arousal to occur simultaneously, ADHD is often diagnosed alongside anxiety disorders, and it is this anomaly which is the focus of this study. Electroencephalograph machinery (EEGs) are able to measure levels of cortical arousal using electrodes placed on the scalp. This research employs this equipment in order to elucidate on how cortical arousal manifests during a task that demands significant involvement from the frontal areas of the brain. A quasi-xperimental research design using non-parametric statistics (Mann-Whitney U Test) was used in order to compare the levels of cortical arousal between 4 groups of 5 research participants with either ADHD, an anxiety disorder, comorbid ADHD and anxiety and no discernible psychopathology. The significant results found in this study point to the fact that, in cases where ADHD and anxiety disorders occur comorbidly, there is a possibility that the anxiety component enables an individual to achieve more pronounced levels of attention, concentration and focus than normal participants and those with ADHD alone...

Orientador: Mary Massumi Itoyama / Resumo: Dentro da subordem Heteroptera algumas espécies pertencentes a família Pentatomidae apresentam espermátides polimórficas em seus lobos testiculares que são oriundas de dois fenômenos: a polimegalia e o lobo harlequin. Para a investigação dessas espermátides polimórficas e de seu desenvolvimento comparadas às espermátides não polimórficas, foram coletadas em São José do Rio Preto as espécies Antiteuchus tripterus, Edessa meditabunda, Loxa deducta e Proxys albopunctulatus (Pentatomidae, Heteroptera). Os machos tiveram seus testículos extraídos, fixados e incluídos em parafina ou resina. O material incluído em parafina foi cortado e corado com hematoxilina/eosina (HE) e analisados em microscopia de luz, outros testículos, destas quatro espécies, foram incluídos em resina e analisados em microscópio eletrônico de transmissão (MET). Os testículos A. tripterus são alongados e divididos em 6 lobos, sendo 1 a 3 sem alterações, 4 e 6 com polimegalia e o 5 o lobo harlequin. Foi possível observar, também, que os lobos 4 e 6 possuem espermátides grandes enquanto o 5 apresenta espermátides pequenas. Em MET foi possível observar em todos os lobos a formação da cauda da espermátide onde ocorre a formação dos derivados mitocondriais e a posterior formação do axonema entre os derivados. Além disso, foi detectada a presença de três tipos de espermatozoides no ducto eferente, sendo estes oriundos dos diferentes lobos. Há o espermatozoide maior com morfologia alongada com pequenas protuberâncias... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Within the suborder Heteroptera, some species belonging to the Pentatomidae family show alterations in some of their testicular lobes, where we can show the polymegaly and the harlequin lobe. For the investigation of these polymorphic cells and their development compared to non-polymorphic cells, the species Antiteuchus tripterus, Edessa meditabunda, Loxa deducta and Proxys albopunctulatus (Pentatomidae, Heteroptera) were collected in São José do Rio Preto. The males had their testicles extracted, fixed and included in paraffin or resin. The material included in paraffin was cut and stained with hematoxylin / eosin (HE) and analyzed under light microscopy, other testicles, of these four species, were included in resin and analyzed using a transmission electron microscope (TEM). The testicles A. tripterus are elongated and divided into 6 lobes, 1 to 3 without changes, 4 and 6 with polymegaly and the 5 the harlequin lobe. It was also possible to observe that lobes 4 and 6 have large cells while 5 has small cells. In TEM, it was possible to observe the formation of the sperm tail in all lobes where the formation of the mitochondrial complex occurs, which divides into two mitochondrial derivatives and the subsequent formation of the axoneme between the derivatives. In addition, the presence of three types of sperm in the efferent duct was detected, these coming from different lobes. There is the larger sperm with elongated morphology with small protuberances at its ends; the inte... (Complete abstract click electronic access below) / Doutor

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The research of my thesis focused on the use of transcriptomic open discovery approaches to interrogate the molecular basis of two distinct yet related neurological disorders that are both associated with cognitive decline, Temporal Lobe Epilepsy and Alzheimer’s Disease. Interestingly, a potential role for compromised synaptogenesis early in disease was common to both, as was the direct role that neurons may play in brain inflammatory processes involving glia. Temporal lobe epilepsy (TLE) is a progressive disorder mediated by pathological changes in molecular cascades and hippocampal neural circuit remodeling that results in spontaneous seizures and cognitive dysfunction. Targeting these cascades may provide disease-modifying treatments for TLE patients. Janus Kinase/Signal Transducer and Activator of Transcription (JAK/STAT) inhibitors have emerged as potential disease-modifying therapies; however, a more detailed understanding of the contribution of JAK/STAT signaling to epileptogenesis is required to increase the potential therapeutic efficacy and reduce adverse effects associated with un-targeted JAK/STAT inhibition. With our collaborators, my lab developed a mouse line in which tamoxifen treatment conditionally abolishes STAT3 signaling from forebrain excitatory neurons (nSTAT3KO). Seizure frequency (continuous in vivo electroencephalography) and memory (contextual fear conditioning and motor learning) were analyzed in wildtype (Wt) and nSTAT3KO mice after intrahippocampal kainate (IHKA) injection as a model of TLE. Selective STAT3 KO in excitatory neurons reduced seizure progression and hippocampal memory deficits without reducing the extent of cell death or mossy fiber sprouting induced by IHKA injection. In my thesis, RNA was extracted from harvested hippocampi 24 h after IHKA and libraries were prepared for bulk RNA-sequencing (70–80 million reads/sample) using the NextSeq 500 Illumina system. 3190 genes were differentially expressed in Wt mice injected with KA vs saline (fold change |1.5|, FDR=<0.05). Ingenuity Pathway Analysis (IPA) revealed significant enrichment in 2 overarching sets of pathways: 1) those related to synaptic signaling and 2) those related to inflammation. As expected, many of the IHKA-induced genes were previously associated with epilepsy or seizure disorders (260 for Seizure Disorder, 267 for Epilepsy or Neurodevelopmental Disorder), and Seizure Disorder had the highest activation score in Neurological Disease based on gene expression patterns. Interestingly, a closer analysis of the IHKA-induced gene set revealed an enrichment of STAT3-associated genes (216), most of which were upregulated by IHKA. Compared to the 3190 Differentially Expressed Genes (DEGs) between IHKA and saline-injected Wt mice 24 hours after SE, more than half of these DEGs (1609) were rescued when comparing IHKA-injected nSTAT3KO mice and saline-injected Wt mice, indicating a significant rescue of gene expression when nSTAT3 is absent in excitatory neurons. While nSTAT3 KO influences the expression of genes in many different pathways, including the reversal of genes whose expression was inhibited in pathways of learning and memory by IHKA, the greatest surprise came from the predicted regulatory control over microglial function. nSTAT3KO mice displayed the greatest number of rescued DEGs compared to IHKA-injected WT mice in pathways that regulate inflammation and ion transport, and while inflammation was an expected response to IHKA, we were surprised to find evidence for its rescue in nSTAT3 KO mice. We also interrogated the expression of the Alzheimer’s disease genome as modeled using a rat model (TgF344-AD ) of familial AD that allows for behavioral and molecular characterization of AD, and expresses an endogenous pathogenic form of tau in addition to Abeta oligomers and plaques. AD is a neuropsychiatric disorder characterized initially by short term memory loss and disorientation, followed by declining cognitive functioning, and eventually, death. Widespread failure of 99% of AD drugs that make it to clinical trials has led to renewed interest in early signatures of disease in hopes of altering disease trajectory through early intervention. Key to such efforts is capturing a molecular window into AD at its earliest stages. The TgF344-AD rat shows overt pathology (including Aβ plaques, frank neuronal loss, and endogenous tau pathology) at 16 and 26 mo, but only to a very limited extent at 6 mo (Towne, 2013). Thus, in my thesis research, we set out to uncover any cell-type specific transcriptomic alterations that may be present in advance of major behavioral deficits or appearance of pathology, given that a strong body of literature suggests a long pre-symptomatic stage of illness in which subtle abnormalities may be present. 10x Genomics’ v3 gene expression assays were used to perform snRNA-seq on freshly dissected hippocampi from 6 mos, 9 mos and 19 mos littermate pairs of Tg and Wt rats (n=16 for 6 months and 9 months, with 8 for 19 months). ~2000 cells/subject were collected, and cDNA libraries were sequenced to a depth of ~120k reads/nuclei. Interestingly, data analysis revealed wide-scale gene changes in dentate granule cells (DGCs) and non-DGC excitatory neurons (Excit Ns) at 6 mos, suggestive of a significant decrease in synaptogenesis in Tg vs their Wt littermates, as well as small increases in cholesterol biosynthesis in the Tg rats in these cell types. By 9 months, some differentially expressed genes were observed across genotype in classes of glial cells, but the strongest impact on gene expression could still be seen in Excit Ns and DGCs, which continued to display evidence of decreased synaptogenesis, though to a lesser extent than at 6 mos. Interestingly, 9 mos Tg rats displayed an even stronger upregulation in genes related to cholesterol biosynthesis than 6 mos for both DGCs and Excit Ns. At 19 months, cholesterol and steroid biosynthesis were amongst the top biological pathways enriched for in Excit Ns and Inhibitory neurons of the Tg, to an even greater extent than changes in synaptogenesis. Altogether, our results suggest the transcriptional basis for a profound suppression of synapse formation or maintenance during early stages of illness in the TgF344-AD rat model, as well as abnormalities in neuronal cholesterol biosynthesis. Given that cholesterol is a key component of plasma membranes and lipid rafts, structures needed for the generation of new synapses and the stability of their receptor populations, it may be that deficiencies in the available cholesterol of Tg neuronal cells is leading to the impaired synaptogenesis in these cell types. Future work will focus on identifying whether these transcriptional alterations can be detected at even earlier time points, whether they are prescient for changes at the membrane in vivo that are correlated with memory impairment, and whether they are related to the alterations in the genome seen in our acquired epilepsy models, suggesting a common theme for the brain’s genomic response to injury of the hippocampus. / 2025-02-12T00:00:00Z

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