Orientadores: Fernando Cendes, Priscila Camile Barioni Salgado / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-20T06:49:09Z (GMT). No. of bitstreams: 1 Nogueira_MateusHenrique_M.pdf: 10853006 bytes, checksum: 6d8dedc970380ba9d12e0c0d44f6b32e (MD5) Previous issue date: 2012 / Resumo: Os transtornos de humor e de ansiedade são transtornos psiquiátricos frequentemente encontrados na epilepsia de lobo temporal mesial (ELTM). Essas comorbidades aparentam ser o produto de uma complexa interação entre os efeitos das drogas antiepilépticas (DAEs), das alterações neurobiológicas associadas às crises epilépticas, das experiências subjetivas e da vulnerabilidade social causada pelo impacto psicossocial da epilepsia. A proposta deste estudo foi avaliar os transtornos de humor e de ansiedade em pacientes com ELTM e correlacionar os dados clínicos e demográficos dos pacientes com as comorbidades psiquiátricas. Os participantes, avaliados no período de 02/2010 a 07/2011, eram pacientes com ELTM acompanhados no ambulatório de epilepsia do Departamento de Neurologia da FCM/UNICAMP. As entrevistas ocorreram de forma individual, com duração média de uma hora e meia. Foram utilizados como instrumentos de avaliação a Entrevista Clínica Estruturada para o DSM-IV-Transtornos do Eixo I (SCID I), o Inventário de Ansiedade Estado e Traço - IDATE I e II e o Inventário de Depressão de Beck (BDI). Os pacientes que concordaram em participar do estudo assinaram o Termo de Consentimento Livre e Esclarecido, aprovado pelo Comitê de Ética da UNICAMP. Utilizamos para a análise estatística o programa SYSTAT 9®, com os testes Qui-quadrado ou Teste Exato de Fisher, Mann-Whitney e as correlações de Spearman. Foram avaliados 104 pacientes, considerando os critérios de inclusão e exclusão. Os transtornos psiquiátricos foram encontrados em 34 (32,70%) pacientes, sendo que destes, 47,06% não possuíam um diagnóstico prévio de transtornos psiquiátricos. O grupo de pacientes com comorbidades psiquiátricas se mostrou mais refratário ao tratamento clínico (p=0,03), apresentando maior frequência mensal de crises epilépticas (p=0,004) e maiores escores no BDI, IDATE-Traço e IDATE-Estado (p<0,0001). Houve associação significativa entre a refratariedade ao tratamento clínico da epilepsia e maiores escores no BDI (p<0,0001), IDATE-Traço e IDATE-Estado (p=0,002). Os resultados apontaram uma tendência em relação à maior ocorrência de transtornos psiquiátricos no gênero feminino (p=0,052), apresentando também escores maiores no BDI em comparação ao gênero masculino (p=0,029). Subdividimos o grupo de pacientes com transtornos psiquiátricos em dois grupos: um apenas com indicativos de transtornos de humor (GTH) e o outro com transtornos de humor e de ansiedade (GTHA). Os pacientes do GTHA apresentaram maior frequência mensal de crises (p=0,029) em relação aos pacientes do GTH. A presença de ideação suicida foi significativa (p<0,0001) no grupo de pacientes com transtornos psiquiátricos e correlacionada a maiores escores no BDI, IDATE-Traço e IDATE-Estado (p<0,0001). Maiores escores no BDI foram relacionados aos pacientes que não possuíam uma atividade profissional (p=0,046). A associação entre as comorbidades psiquiátricas e a epilepsia ainda é subdiagnosticada e pode influenciar negativamente o tratamento clínico da epilepsia / Abstract: Mood and anxiety disorders are psychiatric disorders often found in mesial temporal lobe epilepsy (MTLE). These comorbidities appear to be the product of a complex interaction between the effects of antiepileptic drugs (AEDs), neurophysiological changes resulting from epileptic seizures, subjective experiences and social vulnerability caused by psychosocial impact of epilepsy. The purpose of this study was to assess mood disorders and anxiety in patients with MTLE and correlate the clinical and demographic data of patients with psychiatric comorbidities. Patients were evaluated between February 2010 and July 2011. Patients were followed at the epilepsy outpatient clinic from the department of neurology at FCM / UNICAMP. The interviews took place individually, with an average time of one and a half hour. We used the following instruments: Structured Clinical Interview for DSM-IV Axis I Disorders (SCID I), State Anxiety Inventory-Trait - STAI I and II, and Beck Depression Inventory (BDI). Patients who agreed to participate signed a consent form approved by the Ethics Committee of UNICAMP. We used the program SYSTAT ® 9 for statistical analysis. We performed chi-square or Fisher's exact test, Mann- Whitney and Spearman correlations. We evaluated 104 patients considering the criteria for inclusion and exclusion. Psychiatric disorders were found in 34 (32,70%) patients, of whom 47,06% did not have a prior diagnosis of psychiatric disorders. The group of patients with psychiatric comorbidity was more refractory to medical therapy (p = 0,03), with higher monthly frequency of seizures (p = 0,004) and higher BDI, STAI-Trait and STAI-State (p <0,0001) scores. We found a significant association between the clinical treatment of refractory epilepsy and higher scores on BDI (p <0,0001) and STAI-Trait and STAI-State (p = 0,002). The results showed a trend toward higher incidence of psychiatric disorders in females (p = 0,52), who showed higher BDI scores compared to males (p = 0,029). We subdivided the group of patients with psychiatric disorders in two groups: one with only indicative of mood disorders (GTH) and the other with mood disorders and anxiety (GTHA). Patients in the GTHA had higher monthly frequency of seizures (p = 0,029) compared to patients of GTH. The presence of suicidal ideation was significant (p <0,0001) in patients with psychiatric disorders and correlated with higher scores on BDI, STAI-Trait and STAI-state (p <0,0001). Higher BDI scores were correlated with patients who did not have a professional activity (p = 0,046). The association between psychiatric comorbidities and epilepsy still is underdiagnosed and can negatively influence the clinical treatment of epilepsy / Mestrado / Fisiopatologia Médica / Mestre em Ciências

Abstract Epilepsy is associated with reproductive disorders and decreased fertility. The role of antiepileptic medication and type of epilepsy in development of these disorders has been widely debated. The effects of oxcarbazepine on reproductive function in women and the effects of antiepileptic medication on male fertility have not been previously studied, and only a few studies have evaluated fertility in subjects with epilepsy in a population based setting. This study aimed to analyze predictors of reproductive disorders and the effects of oxcarbazepine on reproductive function in women. Moreover, the effects of antiepileptic medication on male reproductive health were also evaluated, and finally, the reproductive health of patients with epilepsy and the normal population was compared in a population based setting. The study was conducted in the Departments of Neurology, Gynecology and Obstetrics and Public Health Science and General Practice in the University of Oulu. Studies I–III were cross-sectional studies consisting of 249 subjects with epilepsy and 247 control subjects. Study IV was a retrospective study; the data was based on Northern Finland Birth Cohort 1966(NFBC1966), consisting of 12,058 subjects, of which 222 had epilepsy. In studies I–III all subjects were interviewed, clinical examinations were done, blood samples were analyzed and ovarian ultrasound examination or testicular ultrasound examination and sperm samples were studied. In study IV all subjects with epilepsy were identified from NFBC1966 and patient files were reviewed. Fertility analyses were based on information obtained from the Finnish Population Center and Finnish Birth Register. Reproductive disorders were more common in women with idiopathic generalized epilepsy and in women taking valproate. Also young age increased the risk of these disorders. Oxcarbazepine was associated with reproductive disorders in women with epilepsy. In men all antiepileptic drugs studied were associated with sperm abnormalities, and sperm abnormalities in men taking valproate were associated with decreased testicular volume. In a population based setting active epilepsy and antiepileptic medication during adulthood decreased fertility. The reproductive endocrine effects of AEDs should be taken into consideration when prescribed to fertile aged men and women, especially, if the anticipated duration of treatment is long.

Angelman syndrome (AS) is a rare genetic and neurological disorder presenting with severe developmental delay, ataxia, epilepsy, and lack of speech. AS is associated with a neuron-specific loss of function of the maternal UBE3A allele, a gene encoding an E3 ubiquitin ligase. Currently, no cure exists for this disorder; however, recent research using an AS mouse model suggests that pharmacological intervention is plausible, and can alleviate some of the detrimental phenotypes reported in AS patients. Although there is no curative treatment for AS, seizure medication and behavioral therapies are most commonly prescribed in order to minimize symptoms. However, these options only moderately improve quality of life and can cause adverse side effects, such as alterations in mood and cognition following seizure treatment. Unfortunately, epilepsy is a common cause of death in AS and affects greater than 80% of AS patients, with 77% of those patients remaining refractory. The severity of seizures and lack of consistently effective anti-epileptic medications for AS patients demonstrates a considerable need for other therapeutic options. The goal of this work was to evaluate the effects of seizure therapies that have proven beneficial for treating refractory epilepsy in seizure-related disorders. These studies focused specifically on advances in both a pharmacological and dietary therapy evaluated in the AS mouse model. Previous work in our lab has demonstrated the importance of interneurons and GABAergic tone in hippocampal network regulation and cognition. GABA is an important modulator of synaptic plasticity, and learning increases both inhibitory synaptogenesis and GABA release from hippocampal inhibitory neurons. A neuronal excitatory/inhibitory imbalance, coupled with decreased GABAergic tone, altered synaptic plasticity, and impaired cognition have been reported in the AS mouse model. Therefore, we proposed to examine two therapeutic strategies used in seizure treatment – a ketone ester (KE) supplement, which is thought to increase the [GABA]/[glutamate] ratio via alterations in brain metabolism, and ganaxolone, a positive allosteric modulator of GABAA receptors. We evaluated the effects of each therapeutic on learning and cognitive enhancement, alterations in synaptic function, and anticonvulsant activity. We hypothesized that both the KE and ganaxolone would demonstrate anticonvulsant efficacy in both behavioral and chemiconvulsant seizure models. Additionally, as chronic epilepsy has been linked to progressive cognitive and memory impairment which may be related to GABA deficiencies, we hypothesized that both therapeutics would improve cognition and modulate synaptic plasticity (i.e., synaptic function). KE administration produced sustained ketosis and improved motor coordination, learning and memory, and synaptic plasticity in AS mice. The KE was also anticonvulsant and altered brain amino acid metabolism in AS treated animals. Ganaxolone was anxiolytic, anticonvulsant, and improved motor deficits in AS mice. Four weeks of treatment also led to recovery of spatial working memory and hippocampal synaptic plasticity deficits. This study demonstrates that the KE and ganaxolone ameliorate many of the behavioral abnormalities in the adult AS mouse, possibly through modulations of GABAergic tone. These results support clinical investigation of both the KE and ganaxolone in AS, which may lead to the development of a novel treatment for AS patients.

No description available.

Background: Despite the so-called "generalised" nature of many epileptic seizures, patient specific spatio-temporal properties have been shown using imaging data at the macroscopic level of the cortex. Previous computational models have failed to account for spatial heterogeneities at the scale of the entire cortex. Furthermore, one of they key benefits of developing a model is the ability to easily test stimulation protocols. Previous studies of generalised spike-wave (the hallmark of absence epilepsy) have abstracted away from this.METHODSIn this work we develop a set of models of epileptic activity, one of which is at the scale of the entire cortex and incorporates anatomically relevant connectivity from human subjects. A similar model incorporating physiologically relevant thalamocortical circuitry is developed in order to test hypotheses regarding stimulation protocols.RESULTSWe show that the model can account for large-scale spatio-temporal dynamics similar to those seen in epileptic patients. We demonstrate, using the model of thalamocortical interaction, that such a modelling approach can be used for the evaluation of stimulation protocols which are shown to successfully abort the seizure prematurely.CONCLUSIONThis work highlights the importance of computational modelling to support existing data and to make specific predictions regarding testable hypotheses. For example, a stimulus given at the correct time with the correct amplitude will stop the seizure.

Orientador: Donizeti Cesar Honorato / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-07T11:34:40Z (GMT). No. of bitstreams: 1 Veronez_DjaniraAparecidadaLuz_D.pdf: 1713916 bytes, checksum: 0bb9082c7d06dda2f3716d4715c53cd9 (MD5) Previous issue date: 2006 / Resumo: A epilepsia do lobo temporal (ELT) apresenta-se em um percentual elevado de pacientes que são refTatários ao tratamento clínico. Fato este que faz da ELT a epilepsia que melhor responde ao tratamento cirúrgico baseado na lobectomia do lobo temporal e na amígdalo-hipocampectomia. Esta última técnica cirúrgica consiste na ressecção parcial do corpo amigdalóide, do hipocampo e do giro para-hipocampal; estruturas consideradas epileptogênicas. Destas estruturas, o corpo amigdalóide é o único ainda com volume morfológico e morfométrico indeterminado. Com base nestes fatos, este trabalho apresenta como objetivos, determinar em cérebros de cadáveres humanos, parâmetros quantitativos e tridimensionais do corpo amigdalóide a partir da análise de cortes bidimensionais; definir os limites topográficos macroscópicos do corpo amigdalóide; estabelecer as distâncias entre o como temporal do ventrículo lateral e as superficies utilizadas nos acessos cirúrgicos; expressar os resultados em escalas métricas, da análise morfométrica e estereológica e sugerir o uso destas nas amígdalo-hipocampectomias. Para isto realizamos análise de cortes seriados, coronais e parassagitais do corpo amigdalóide, que foram avaliados através de um sistema de processamento e análise de imagem. O método de Cavalieri foi utilizado para estabelecer o volume absoluto do corpo amigdalóide. Por último, foram feitas análises estatísticas e apresentação gráfica dos resultados. Os hemisférios cerebrais, direitos e esquerdos foram analisados separadamente, para verificar a morfometria do corpo amigdalóide. Foram observadas diferenças inter-hemisféricas na volumetria do corpo amigdalóide. Nos cortes coronais a média do volume absoluto do corpo amigdalóide direito foi de 1.870mm3 e do corpo amigdalóide esquerdo foi de 1.807mm3, com índice de assimetria de 3,4%. Nos cortes parassagitais a média do volume absoluto do corpo amigdalóide direito foi de 1.927mm3 e do corpo amigdalóide esquerdo foi de 1.878mm3, com índice de assimetria de 2,6%. Não foi identificada variabilidade anatômica significativa na topografia dos ventrículos laterais e do corpo amigdalóide. Nossos resultados demonstram que a análise morfométrica e estereológica aplicada ao corpo amigdalóide e as medidas obtidas na topografia do lobo temporal constitui instrumento confiável, que pode auxiliar nas cirurgias de amígdalo-hipocampectomias / Abstract: The temporallobe epilepsy (TLE) has a high percentage of patients who are reffactory to a clinical treatment. This fact makes TLE the epilepsy that best responds to a surgi cal treatment. It is based on lobectomy of temporallobe and on amygdalohippocampectomy. This surgi cal technique consists of a partial resection of the amygdaloid complex, hippocampus, and gyrus parahippocampal. These structures are considered as epileptogenics. Among these structures, the amygdaloid complex is the only with undetermined morphological and morphometric volume. Based on these facts, this study will have the following objectives: to determine in brains of human corpses the quantitative and three-dimensional parameters of the amygdaloid complex obtained ITom analyses of two-dimensional slices; to define the topographic macroscopic limits of the amygdaloid complex; to establish the distances between the temporal hom of the lateral ventricles and the surfaces used for the surgi cal approaches; to express the results in metrical scales of morphometric and estereological analysis of the amygdaloid complex, and to suggest the use of them in an amygdalohippocampectomy. In order to do this, we have accomplished analysis of serial, coronary and parasagital slices of the amygdaloid complex, which were evaluated through a processing system and an image analysis. The Cavalieri Method was used to establish the absolute volume of the amygdaloid complexo Finally, estatistics analyses and a graphical presentation of the results were done. Right and left brain hemispheres were analyzed separately in order to verify the morphometry of the amygdaloid complexo The inter-hemispherical differences in the volumetry of the amygdaloid complex were observed. In the coronal slices the absolute medium volume of the right amygdaloid complex was 1.894mm3, and the left was 1.837mm3 with an index of asymmetry of 3,4%. In the parassagital si ices the absolute medium volume of the right amygdaloid complex was of 1.927mm3 and the left amygdaloid complex was 1.878mm3 with an index of asymmetry of 2,6%. We have verified that there was no significant anatomical variation in the topography of lateral ventricles and the amygdaloid complexo Our results demonstrate that a morphometric and estereological analysis applied to the amygdaloid complex and measurements obtained in the topography of the temporallobe is a reliable instrument, which can help in surgeries ofthe amygdalohippocampectomy / Doutorado / Ciencias Biomedicas / Doutor em Ciências Médicas

Orientador: Marilisa Mantovani Guerreiro / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-22T07:18:22Z (GMT). No. of bitstreams: 1 Capelatto_LiviaLucenadeMedeiros_D.pdf: 6633660 bytes, checksum: 2c203cf41487511bbd4f62288b5f851f (MD5) Previous issue date: 2013 / Resumo: A Epilepsia focal benigna da infância com paroxismos centro-temporais (EBICT), ou epilepsia rolândica (ER), é a forma mais frequente de epilepsia idiopática da infância. Os objetivos deste estudo foram aprofundar o exame neurológico através da detecção de sinais menores e avaliar criteriosamente as anormalidades neurofisiológicas de pacientes com ER. Para atingirmos o primeiro objetivo aplicamos o QNST II e WISC III em um grupo de pacientes com ER que foram comparados a um grupo-controle composto por crianças normais. Para atingirmos o segundo objetivo, analisamos os EEGs de um grupo de pacientes com ER que foram comparados a um grupo de pacientes com epilepsia sintomática e descargas na área rolândica. A primeira parte do estudo incluiu 40 sujeitos durante o período de Março de 2007 a Dezembro de 2009. As crianças eram de ambos os gêneros e faixa etária de 9 a 15 anos (média de idade de 12 anos) e foram subdivididos em dois grupos: G1 (Grupo 1) - 20 pacientes com ER (8 meninas), 85% controladas; G2 (Grupo 2) - 20 sujeitos do grupo controle (10 meninas), sem queixas neurológicas e pareados por idade e sexo. Os resultados demostraram que entre os grupos quanto ao QI de execução e total houve com melhor desempenho das crianças com epilepsia rolândica (p = 0,001 e p = 0,004, respectivamente). Houve proporcionalidade entre dados obtidos do QNST II e o WISC III pela análise de correlação de Spearmann, tanto diante do QI total (p=0,015), QI execução (p=0.045) como do QI verbal (p=0.031). Concluímos que crianças com epilepsia rolândica em fase de controle de crises podem não apresentar sinais neurológicos menores, sendo o QNST II uma instrumento confiável para este tipo de avaliação. Na segunda parte do estudo, foram selecionados e analisados 45 EEGs de pacientes, sendo subdivididos em dois grupos: G1 (Grupo 1) - 21 pacientes com ER (13 meninas), com idade média de 9,2 anos ( + 2,6); G2 (Grupo 2) - 24 pacientes com epilepsia sintomática (16 meninas), com idade média de 10,7 ( +3,5). Os traçados foram realizados entre Janeiro de 2001 e Março de 2009. Em relação à atividade epileptiforme, as variáveis estudadas e comparadas foram dipolo horizontal, fenômeno de dupla ponta, extensão das descargas além da área rolândica e atividade de base. Os resultados não mostraram diferença estatisticamente significativa entre os dois grupos, a não ser em relação à atividade de base (p=0,008). Concluímos que as características eletrencefalográficas avaliadas neste estudo refletem, na realidade, um distúrbio elétrico da área rolândica / Abstract: The benign epilepsy of childhood with centrotemporal spikes (BECTS), or rolandic epilepsy (RE), is the most frequent form of idiopathic epilepsy of childhood. The aims of this study were to deep the neurological examination by searching for soft signs and to evaluate carefully the neurophysiological abnormalities of patients with RE. To achieve the first aim we applied the QNST II and WISC III in a group of patients with RE that were compared to a control group composed of normal children. To achieve the second aim we analyzed the EEGs of a group of patients with RE that were compared to a group of patients with symptomatic epilepsy and discharges in rolandic area. The first part of the study included 40 subjects during the period of March 2007 to December 2009. Children of both genders and aging from 9-15 years were included and divided into two groups: G1 (Group 1) - 20 patients with RE, 85% were controlled; G2 (Group 2) - 20 subjects in the control group without any neurological complaint, and matched with G1 by age and sex. Results showed that a significant difference between the groups against the run and total IQ with better performance in children with rolandic epilepsy. There was correlation between the data obtained QNST II with WISC III (total IQ, verbal and executive), both compared to the total IQ (p = 0.015), executive IQ (p = 0.045) and verbal IQ (p = 0.031). We conclude that children with rolandic epilepsy under control seizures may not have neurological soft signs, QNST II being a reliable tool for this type of evaluation. In the second part of the study we selected and analyzed EEGs of 45 patients, divided into two groups: G1 (Group 1) - 21 patients with RE, G2 (Group 2) - 24 patients with symptomatic epilepsy and rolandic discharges. The tracings were performed between January 2001 and March 2009. The studied variables in epileptiform activity were horizontal dipole, double spike phenomenon, extent of the discharges beyond the rolandic area and background activity. The results showed no statistically significant difference between the two groups, except in relation to background activity (p = 0.008). We concluded that the neurophysiological features assigned to the RE reflect in fact an electrical disturbance of rolandic area and they can be found in any form of epilepsy that involves this region / Doutorado / Neurologia / Doutora em Ciências Médicas

Orientadores: Iscia Teresinha Lopes Cendes, André Schwambach Vieira / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-28T00:14:14Z (GMT). No. of bitstreams: 1 MoratodoCanto_Amanda_M.pdf: 2477839 bytes, checksum: f2e9db3a96d271cc686d6d66697b0e59 (MD5) Previous issue date: 2015 / Resumo: O uso de modelos animais para estudo de doenças humanas é importante para o entendimento dos mecanismos fisiopatológicos destas doenças. Particularmente, os modelos que reproduzem a Epilepsia do Lobo Temporal (ELT) em roedores, apresentam uma epileptogenicidade similar à encontrada em tecidos "epilépticos" humanos quando estudados ex vivo. A ELT afeta cerca de 40% dos pacientes adultos, e é caracterizada clinicamente por um desenvolvimento progressivo de crises epilépticas com foco no lobo temporal. Pacientes que apresentam ELT normalmente não respondem aos tratamentos. Destas, a Epilepsia do Lobo Temporal Mesial (ELTM) e a mais comum e se caracteriza pelo acometimento das estruturas mesiais do lobo temporal, como o caso da Esclerose Hipocampal. A proteômica dispõe de ferramentas poderosas que nos permitem elucidar mecanismos biológicos complexos, encontrar proteínas alteradas em todo o organismo e descrever padrões de expressões proteicas em diferentes condições fisiológicas e patológicas. Portanto, é relevante analisar esse padrão de expressão no hipocampo de modelos animais de ELTM usando técnicas de proteômica, a fim de gerar informações que nos auxiliem no entendimento dos mecanismos envolvidos na epileptogênese desses modelos. No presente estudo, as proteínas identificadas podem nos indicar novas vias envolvidas com a epileptogênese. Além disso, nossos dados demonstram que uma complexidade molecular adicional pode ser observada quando analisamos as diferentes sub-regiões do hipocampo separadamente. Portanto, acreditamos que a integração dos dados de proteômica com dados obtidos por outras "ômicas" podem gerar dados ainda mais informativos sobre esses processos neuronais / Abstract: Studies about human diseases using animal models are really important to our understanding about the physiopathology mechanisms from those diseases. Particularly, the models that reproduce the Temporal Lobe Epilepsy (TLE) in rodents, presents epileptogenicity similar to that found in ex vivo human tissues. The TLE affects around 40% of the adult patients and it is clinic characterized by a progressive development of seizures with temporal lobe focus, caused by an unbalance between the excitatory and inhibitory neurotransmission. Patients who present that type of epilepsy normally don¿t respond well to the treatments. Of this type of epilepsy, the Mesial Temporal Lobe Epilepsy (MTLE) is the most common one and it is characterized by the commitment of the mesial temporal lobe structures, such as in the Hipocampal Sclerosis. To realize these studies the proteomics has many powerful tools that allow us to elucidate complex biological mechanisms, to find altered proteins in the whole organism and describe protein expression patterns in different physiological and pathological conditions. Therefore, it¿s relevant to study this protein expression pattern in the hippocampus of animal models of MTLE using proteomics techniques, searching for informative data that lead us to the understanding of the involved mechanisms in the epileptogenicity. In this study we identified proteins that can indicate new pathways involved in the epileptogenesis processes. Furthermore, our data demonstrate that additional molecular complexity could be observed as hippocampal subfields were analyzed separately. We believe that the further integration of the proteomic data with other "omics" approaches could generate even more informative data about those neuronal processes / Mestrado / Fisiopatologia Médica / Mestra em Ciências

The synapse is essential for normal neuronal communication and synaptic abnormalities could underlie many neuronal pathologies leading to such diseases as epilepsy and autism. Recent reports suggest that the Wnt signaling pathway is essential for normal synaptic development and function. However the role of specific Wnt ligands and their downstream signaling molecules play in synapse formation and function remain unclear. PRICKLE1 (PK1) and PRICKLE2 (PK2) are downstream Wnt signaling molecules which are suggested to play essential roles in neuronal function. PK1 was recently shown to be mutated in three large families with epilepsy, and Pk2 interacts with post-synaptic density 95 and subunits of the NMDA receptor. Although it seems clear that PK1 and PK2 are critical for normal neuronal function, their role in synaptic function and animal behavior remain to be investigated. In Aim1, we show that mutations in prickle (pk) genes are associated with seizures in humans, mice, and flies. We identified human epilepsy patients with heterozygous mutations in either PK1 or PK2. In overexpression assays in zebrafish, pk mutations resulted in aberrant pk function. A seizure phenotype was present in the Pk1-null mutant mouse, two Pk1 point mutant (missense and nonsense) mice, and a Pk2-null mutant mouse. Drosophila with pk mutations displayed seizures that were responsive to anti-epileptic medication, and homozygous mutant embryos showed neuronal defects. In aim 2, we describe two families with ASD-specific mutations in the non-canonical Wnt gene PK2. These mutations reduced the co-localization of the human PK2 protein with PSD-95, another protein implicated in ASDs. Studying Pk2 function in mice, we found that disrupting Pk2 in mouse hippocampal neurons reduced dendrite branching, synapse number, and post-synaptic density size. Consistent with these findings, disrupting Pk2 decreased the frequency and size of spontaneous miniature synaptic currents. Interestingly, these phenotypes were rescued by wild-type human PK2, but not the ASD-associated PK2 mutants suggesting that these mutations cause a critical loss of PK2 function. Behavioral studies in Pk2-/- mice suggest that loss of Pk2 function lead to ASD-like behaviors. These studies provide new insight into the biological roles of PK2, its behavioral importance, and firmly link non-canonical Wnt signaling abnormalities and ASDs. Together, Aim1 and 2 show that the Pk proteins are critical regulators of normal neuronal function and suggest that Pk2 could be a link between epilepsy and autism.

Epilepsy and Autism Spectrum disorders (ASD) are both complex neurodevelopmental disorders which share approximately 30% comorbidity. Epilepsy is characterized by unprovoked recurrent seizures and affects ~1% of the population while ASDs are characterized by deficits in language, social, and behavior and found 1 in 68 people. Variants in synaptic genes suggest disruptions in synaptic regulation underlie both conditions. PRICKLE1 and PRICKLE2 are known core WNT/ PCP genes implicated in Progressive myoclonic epilepsy in families and in the general population. Humans, mice, zebrafish, and Drosophila with disrupted Prickle exhibit epileptic behavior and other neurological deficits. Prickle is implicated in several aspects of neuronal development and mutated proteins display aberrant activity in vivo and in vitro. Recently, variations in PRICKLE1 were associated with ASDs in humans. The mechanisms by which PRICKLE could contribute to ASDs are unknown. Results presented here show Prickle1+/- mice exhibit ASD-like behavior. Prickle1 associates with Synapsin I; a phosphoprotein important for synaptogenesis, axonogenesis, and neurotransmitter release. Mutant R104QPRICKLE1 protein causes a reduction in sizes of dense-core vesicles in neuronal-like PC12 cells. Results indicate PRICKLE1 may be associated with ASDs and possibly involved in synaptic homeostasis. Prickle deregulation has also been associated with neural tube defects and cancers. The mechanism(s) by which Prickle is regulated is incompletely understood. To further elucidate the role of PRICKLE in disease, immunoprecipitates from PRICKLE-expressing stable tetracycline-regulated neuronal-like PC12 cells were identified by mass spectrometry. The deubiquitinating enzyme USP9X was identified as a novel interacting PRICKLE protein. USP9X is a substrate-specific deubiquitinating enzyme implicated in several aspects of neuronal development, associated with X-linked intellectual disability and a candidate gene for epilepsy. Results show that USP9X robustly deubiquitinates and protects PRICKLE from proteasomal degradation. USP9X variants found in the ARRA ASD cohort directly associates USP9X with ASDs. The identified USP9X mutations delete the PRICKLE-interacting domain and provide a possible mechanism for PRICKLE deregulation. Already a target for treating cancer, USP9X can serve as a therapeutic target to regulate PRICKLE levels.