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Previous issue date: 2012-08-09 / The clinical depression treatment faces serious obstacles as the disease mechanism is not fully elucidated. In addition, there are no effective means to predict and prevent depression as well as any biological method of diagnosis. The use of antidepressants is still the basis of the treatments for depression. Lithium has been used clinically as effective drug to treat all phases of bipolar disorder, including major depression. The Selective serotonin re-uptake inhibitors (SSRIs), such as fluoxetine and citalopram, and Tricyclic antidepressant (TCA) as clomipramine are drugs constantly used for depression treatment. Recent evidence has shown an involvement of adenosine and its receptors in the pathophysiology of depression. ATP can be stored and co-released with other neurotransmitters like serotonin and can be hydrolyzed by a cell-surface enzyme family known as ectonucleotidases. Among these members, we highlight the nucleoside triphosphate diphosphohtdrolases (NTPDases) and ecto-5'-nucleotidase. They are able to control the availability of ligands such as ATP and adenosine to its specific receptors. Adenosine deaminase (ADA) can promote the hydrolytic deamination of adenosine to inosine, modulating the extracellulr levels of this neuromodulator. In cholinergic signaling, after its release, acetylcholine (ACh) promotes the activation of specific muscarinic or nicotinic receptors and thus, it promotes diverse cellular responses. ACh is hydrolyzed by acetylcholinesterase (AChE) in acetate and choline in synaptic cleft. The zebrafish has been used in research behavioral neuroscience and is also a choice model for elucidating the development and function of neuronal circuitry. Considering the cholinergic and purinergic signaling are important participation in the CNS and these neurotransmitter pathways have been identified and characterized in zebrafish, the objective of this study was to evaluate the effect of antidepressants on ectonucleotidases, ADA and ACh activities, which are essential enzymes in the modulation of these signaling pathways in the zebrafish brain. We evaluated the ex vivo effects of fluoxetine (1-10 ?M), clomipramine (1-10 ?M), citalopram (70-300 ?M) on ectonucleotidases and ADA activities. It has been also analyzed the in vitro (1 to 1000 ?M) and ex vivo (1 to 10mg/L) effect of lithium on ectonucleotidases and AChE activities and gene expression. There was a significant inhibition of ADP hydrolysis after ex vivo exposure to lithium at 5 and 10 mg/L, whereas an inhibitory effect was observed for AMP hydrolysis only at 10 mg/L. The same treatment decreased the AChE activity in a concentration of 10mg/L. Lithium did not induce significant changes in the analysis of gene expression patterns in the concentrations tested. In vivo treatment, there were no significant changes inectonucleotidases and AChE activities. Treatment with clomipramine promotes an inhibition ecto-5'-nucleotidase activities at the concentration of 5?M when compared to the control group. For ADA activity, we also observed a significant inhibition in the treatment with clomipramine at concentrations of 5 and 10 ?M in membrane fractions of zebrafish brain. However, treatment with fluoxetine and citalopram did not alter ectonucleotidases and ADA activities in the zebrafish brain. Our findings may contribute to a better understanding of pharmacology of antidepressants and their interaction with the cholinergic and purinergic neurotransmission. / O tratamento cl?nico da depress?o enfrenta s?rios obst?culos j? que o mecanismo da doen?a n?o ? totalmente elucidado. Al?m disso, n?o existem meios eficazes para prever e prevenir a depress?o bem como nenhum m?todo biol?gico de diagn?stico. O uso de f?rmacos antidepressivos ainda ? a base dos tratamentos para depress?o. O l?tio tem sido usado clinicamente como f?rmaco eficaz para tratar todas as fases do transtorno bipolar, incluindo depress?o aguda. Os inibidores seletivos da recapta??o de serotonina (ISRSs), como fluoxetina e citalopram, e os antidepressivos tric?clicos (TCA), como a clomipramina, s?o f?rmacos constantemente utilizados para o tratamento da depress?o. Evid?ncias recentes mostram o envolvimento da adenosina e seus receptores na patofisiologia da depress?o. O ATP pode ser armazenado e co-liberado, juntamente com outros neurotransmissores, como serotonina e noradrenalina, e pode ser hidrolisado at? adenosina por uma fam?lia de enzimas de superf?cie celular, conhecidas como ectonucleotidases. Dentre elas, destacam-se as Nucleos?deo Trifosfato Difosfoidrolases (NTPDases) e a ecto-5?-nucleotidase, capazes de controlar a disponibilidade de ligantes como ATP e adenosina aos seus receptores espec?ficos. A Adenosina desaminase (ADA) pode promover a desamina??o hidrol?tica da adenosina em inosina, modulando os n?veis extracelulares deste neuromodulador. Na sinaliza??o colin?rgica, a acetilcolina (ACh), ap?s liberada, promove a ativa??o de receptores muscar?nicos ou nicot?nicos, e desta maneira a ACh promove diversas respostas celulares. A ACh que permanece na fenda sin?ptica ? hidrolisada pela acetilcolinesterase (AChE) em acetato e colina. O peixe-zebra tem sido utilizado na pesquisa da neuroci?ncia comportamental, sendo tamb?m um modelo de escolha para elucidar o desenvolvimento e a fun??o do circuito neuronal. Considerando que as sinaliza??es purin?rgica e colin?rgica t?m importante participa??o no sistema nervoso central e que essas vias de neurotransmiss?o j? est?o caracterizadas em peixe-zebra, o objetivo desse estudo foi avaliar o efeito de f?rmacos antidepressivos na atividade das ectonucleotidases, ADA e AChE, enzimas essenciais na modula??o destas vias de sinaliza??o em c?rebro de peixe-zebra. Foram avaliados os efeitos ex vivo da fluoxetina (1-10 ?M), clomipramina (1-10 ?M) e citalopram (70-300 ?M) na atividade das ectonucleotidases e ADA. Foi analisado tamb?m o efeito in vitro (1 a 1000 ?M) e ex vivo (1-10mg/L) do l?tio sobre a atividade e express?o g?nica das ectonucleotidases e AChE.A exposi??o ao l?tio inibiu a hidr?lise de ADP nas concentra??es de 5 e 10mg/L e inibiu a hidr?lise de AMP na concentra??o de 10mg/L quando comparado ao grupo controle. Este mesmo tratamento diminuiu a atividade da AChE na concentra??o de 10mg/L.O l?tio n?o induziu altera??es significativas na an?lise do padr?o de express?o g?nica. No tratamento in vitro, n?o foram observadas altera??es na atividade das ectonucleotidases e AChE. O tratamento com a clomipramina mostrou uma inibi??o na atividade da ecto-5?-nucleotidase na concentra??o de 5 ?M quando comparado ao grupo controle. Na atividade da ADA tamb?m observamos uma inibi??o significativa no tratamento com a clomipramina nas concentra??es de 5 e 10 ?M em fra??es de membrana de c?rebro de peixe-zebra. Entretanto, o tratamento com fluoxetina e citalopram n?o alterou a atividade das ectonucleotidases e ADA no c?rebro do peixe-zebra. Nossos resultados podem contribuir para uma melhor compreens?o da farmacologia dos f?rmacos antidepressivos e a sua intera??o com a neurotransmiss?o colin?rgica e purin?rgica.
| Identifer | oai:union.ndltd.org:IBICT/oai:tede2.pucrs.br:tede/5447 |
| Date | 09 August 2012 |
| Creators | Oliveira, Renata da Luz |
| Contributors | Bonan, Carla Denise |
| Publisher | Pontif?cia Universidade Cat?lica do Rio Grande do Sul, Programa de P?s-Gradua??o em Biologia Celular e Molecular, PUCRS, BR, Faculdade de Bioci?ncias |
| Source Sets | IBICT Brazilian ETDs |
| Language | Portuguese |
| Detected Language | English |
| Type | info:eu-repo/semantics/publishedVersion, info:eu-repo/semantics/masterThesis |
| Format | application/pdf |
| Source | reponame:Biblioteca Digital de Teses e Dissertações da PUC_RS, instname:Pontifícia Universidade Católica do Rio Grande do Sul, instacron:PUC_RS |
| Rights | info:eu-repo/semantics/openAccess |
| Relation | 8198246930096637360, 600, 600, 36528317262667714 |
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