This thesis focuses on the regulation of the ubiquitous second messenger Ca2+ in neuronal physiology and disease. Ca2+ signalling in neurons is regulated by ion channels located in the plasma membrane, as well as in the endoplasmic reticulum (ER) and mitochondrial membranes. Ca2+ signalling is essential for numerous cellular processes, including neuronal excitability, neurotransmitter release, synaptic plasticity, and induction of cell death. Age-related disruptions in Ca2+ signalling may contribute to decline of cognitive function and motor control associated with aging. Furthermore, disruption in neuronal Ca2+ signalling is implicated in several neurodegenerative disorders including Alzheimer’s disease (AD), Parkinson’s disease (PD), Huntington’s disease (HD), and Amyotrophic Lateral Sclerosis (ALS). In this thesis, I studied neuronal Ca2+ signalling and how it is affected in neurodegenerative disease. First, I examined the role of the ER Ca2+ binding protein Calreticulin (CRT) in AD. CRT is involved in regulation of ER Ca2+ signalling and modulation of susceptibility to cell death. I found that there was an increase in the expression of CRT in in vitro and in vivo models of AD. However, increased levels of CRT did not alter susceptibility of neuronal cells to death induced by AD-related stressors. Second, I examined the role of X-Linked Inhibitor of Apoptosis Protein (XIAP) in the modulation of neuronal Ca2+ signalling. I found that overexpression of XIAP in neuronal cells modified Ca2+ signalling by decreasing Ca2+ flux through multiple plasma membrane and ER channels. These effects appear to be independent of caspase inhibition, which is one of the ways that XIAP can inhibit apoptosis. Third, I examined a compound found in green tea, L-theanine, a glutamate receptor antagonist that is protective in models of excitotoxic neuronal injury. I found that 24 hour L-theanine treatment reduces the amount of Ca2+ released from neuronal intracellular stores in response to both glutamate stimulation and passive leak through ER channels. An acute 30 minute L-theanine treatment had similar effects. In conclusion, these observations further the understanding of the regulation of Ca2+ signalling in neurons and may lead to novel therapeutic strategies in neurodegenerative disease. / October 2008

This thesis focuses on the regulation of the ubiquitous second messenger Ca2+ in neuronal physiology and disease. Ca2+ signalling in neurons is regulated by ion channels located in the plasma membrane, as well as in the endoplasmic reticulum (ER) and mitochondrial membranes. Ca2+ signalling is essential for numerous cellular processes, including neuronal excitability, neurotransmitter release, synaptic plasticity, and induction of cell death. Age-related disruptions in Ca2+ signalling may contribute to decline of cognitive function and motor control associated with aging. Furthermore, disruption in neuronal Ca2+ signalling is implicated in several neurodegenerative disorders including Alzheimer’s disease (AD), Parkinson’s disease (PD), Huntington’s disease (HD), and Amyotrophic Lateral Sclerosis (ALS). In this thesis, I studied neuronal Ca2+ signalling and how it is affected in neurodegenerative disease. First, I examined the role of the ER Ca2+ binding protein Calreticulin (CRT) in AD. CRT is involved in regulation of ER Ca2+ signalling and modulation of susceptibility to cell death. I found that there was an increase in the expression of CRT in in vitro and in vivo models of AD. However, increased levels of CRT did not alter susceptibility of neuronal cells to death induced by AD-related stressors. Second, I examined the role of X-Linked Inhibitor of Apoptosis Protein (XIAP) in the modulation of neuronal Ca2+ signalling. I found that overexpression of XIAP in neuronal cells modified Ca2+ signalling by decreasing Ca2+ flux through multiple plasma membrane and ER channels. These effects appear to be independent of caspase inhibition, which is one of the ways that XIAP can inhibit apoptosis. Third, I examined a compound found in green tea, L-theanine, a glutamate receptor antagonist that is protective in models of excitotoxic neuronal injury. I found that 24 hour L-theanine treatment reduces the amount of Ca2+ released from neuronal intracellular stores in response to both glutamate stimulation and passive leak through ER channels. An acute 30 minute L-theanine treatment had similar effects. In conclusion, these observations further the understanding of the regulation of Ca2+ signalling in neurons and may lead to novel therapeutic strategies in neurodegenerative disease.

This thesis focuses on the regulation of the ubiquitous second messenger Ca2+ in neuronal physiology and disease. Ca2+ signalling in neurons is regulated by ion channels located in the plasma membrane, as well as in the endoplasmic reticulum (ER) and mitochondrial membranes. Ca2+ signalling is essential for numerous cellular processes, including neuronal excitability, neurotransmitter release, synaptic plasticity, and induction of cell death. Age-related disruptions in Ca2+ signalling may contribute to decline of cognitive function and motor control associated with aging. Furthermore, disruption in neuronal Ca2+ signalling is implicated in several neurodegenerative disorders including Alzheimer’s disease (AD), Parkinson’s disease (PD), Huntington’s disease (HD), and Amyotrophic Lateral Sclerosis (ALS). In this thesis, I studied neuronal Ca2+ signalling and how it is affected in neurodegenerative disease. First, I examined the role of the ER Ca2+ binding protein Calreticulin (CRT) in AD. CRT is involved in regulation of ER Ca2+ signalling and modulation of susceptibility to cell death. I found that there was an increase in the expression of CRT in in vitro and in vivo models of AD. However, increased levels of CRT did not alter susceptibility of neuronal cells to death induced by AD-related stressors. Second, I examined the role of X-Linked Inhibitor of Apoptosis Protein (XIAP) in the modulation of neuronal Ca2+ signalling. I found that overexpression of XIAP in neuronal cells modified Ca2+ signalling by decreasing Ca2+ flux through multiple plasma membrane and ER channels. These effects appear to be independent of caspase inhibition, which is one of the ways that XIAP can inhibit apoptosis. Third, I examined a compound found in green tea, L-theanine, a glutamate receptor antagonist that is protective in models of excitotoxic neuronal injury. I found that 24 hour L-theanine treatment reduces the amount of Ca2+ released from neuronal intracellular stores in response to both glutamate stimulation and passive leak through ER channels. An acute 30 minute L-theanine treatment had similar effects. In conclusion, these observations further the understanding of the regulation of Ca2+ signalling in neurons and may lead to novel therapeutic strategies in neurodegenerative disease.

Made available in DSpace on 2015-04-17T15:02:59Z (GMT). No. of bitstreams: 1 arquivototal.pdf: 1361043 bytes, checksum: 4455c38a98b7073e2147348a625e880c (MD5) Previous issue date: 2013-03-01 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Anxiety is characterized by a state of tension, apprehension and discomfort that arise from internal or external danger imminent and may be a response to stress or environmental stimuli, and can measured by psychological and physiological changes of the individual. The L-theanine (gamma-L-glutamic acid ethylamide) or theanine is an amino acid found in Camellia sinensis, commonly known as "green tea". This amino acid has been used to reduce mental and physical stress, improving memory function and for treating anxiety. Because the anxiety disorders are among the most common disorders seen in both the general population and in the services of primary health care, it is relevant to the investigation of non-pharmacological methods for their treatment. The aim of this study was to investigate the psychophysiological effects of L-theanine on a model of anxiety in healthy humans. The study was an experimental, kind of randomized controlled clinical trial. Thirtie was composed of healthy students, male or female, which were inserted into one of three groups (control, L-theanine 1 or L-theanine 2). The participants of the experimental groups received 200 mg of L-theanine by a capsule as a single dose. The human anxiety was induced by Simulated Public Speaking Test - SPST and was assessed by physiological parameters (blood pressure, heart rate, ending temperature and electrical conductance of the skin) and psychological parameters (anxiety Inventory, trait - STAI-T and state - STAI-E). Statistical tests were considering parametric data the ANOVA (one way) and ANOVA for repeated measures data and considering the nonparametric Kruskal-Wallis and Friedman. Results were considered significant when showed a significance level of 95% (p < 0,05). There was a reduction in the levels of systolic blood pressure and heart rate in group L-Theanine 2 (p <0,05) and the electrical conductance of the skin in group L-theanine 1. There was also a reduction in the score of the STAI-E in both experimental groups since the score remained within the range of anxiety low throughout the test (<40 points). These results suggest that L-theanine has an anxiolytic effect on the physiological and psychological parameters that change in anxiety. Thus, this amino acid may be a non-pharmacological strategy in the treatment thereof. / A ansiedade caracteriza-se por um estado de tensão, apreensão e desconforto, que se originam de um perigo interno ou externo iminente, podendo ser resposta a estresse ou a estímulo ambiental e pode ser mensurada através de medidas psicológicas e fisiológicas do individuo. A L-teanina (ácido gama-etilamida L-glutâmico) ou teanina é um aminoácido encontrado na Camellia sinensis, popularmente conhecida como chá verde . Este aminoácido vem sendo utilizado para reduzir o estresse mental e físico, melhorar a função da memória e no tratamento da ansiedade. Visto que os transtornos da ansiedade estão entre os transtornos mais comumente observados tanto na população geral quanto nos serviços de atenção primária à saúde, é relevante a averiguação de métodos não farmacológicos para o seu tratamento. O objetivo deste estudo foi investigar os efeitos psicofisiológicos da L-teanina sobre um modelo de ansiedade em humanos saudáveis. O estudo teve caráter experimental, do tipo ensaio clínico randomizado e controlado. Foi composto por trinta estudantes saudáveis, do gênero masculino ou feminino, os quais foram inseridos em um dos três grupos experimentais (controle, L-teanina 1 ou L-teanina 2). Os participantes dos grupos experimentais receberam 200 mg de L-teanina na forma de cápsula e em dose única. A ansiedade humana experimental foi induzida pelo Teste de Simulação de Falar em Público (TSFP) e foi avaliada por meio de parâmetros fisiológicos (pressão arterial, frequência cardíaca, temperatura de extremidade e condutância elétrica da pele) e de parâmetros psicológicos (inventário de ansiedade traço IDATE-T e estado IDATE-E). Os testes estatísticos utilizados considerando os dados paramétricos foram o ANOVA (one way) e ANOVA para medidas repetidas e considerando os dados não paramétricos o teste de Kruskal-Wallis e de Friedman. Os resultados foram considerados significativos quando apresentaram um nível de significância de 95% (p < 0,05). Observou-se uma redução nos níveis da pressão arterial sistólica e frequência cardíaca no grupo L-teanina 2 (p < 0,05) e da condutância elétrica da pele no grupo L-teanina 1. Ocorreu também uma redução no escore do IDATE-E nos dois grupos experimentais, uma vez que a pontuação manteve-se dentro da faixa de ansiedade baixa durante todo o teste (< 40 pontos). Estes resultados sugerem que a L-teanina teve um efeito ansiolítico sobre os parâmetros fisiológicos e psicológico que se alteram na ansiedade. Sendo assim, este aminoácido pode ser uma estratégia não farmacológica no tratamento da mesma.