Efeito do estresse crônico imprevisível no metabolismo de nucleotídeos e nucleosídeos em encéfalo de zebrafish (Danio rerio)

• Autor(a) principal: Zimmermann, Fernanda Francine
• Data de Publicação: 2013
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: Biblioteca Digital de Teses e Dissertações da PUC_RS
• Texto Completo: http://tede2.pucrs.br/tede2/handle/tede/5464
• topic: BIOLOGIA CELULAR; BIOLOGIA MOLECULAR; EXPERIMENTAÇÃO ANIMAL; ESTRESSE; ADENOSINA; CNPQ::CIENCIAS BIOLOGICAS::BIOLOGIA GERAL

According to data from the World Health Organization (WHO), stress affects more than 90% of the world's population and is considered a global epidemy. Stress has become an integral part of human life and organisms are constantly subjected to stressful stimuli that change various physiological processes, representing a risk factor for many diseases, such as cancer, cardiovascular, metabolic, infectious and gastrointestinal deseases and depression. The zebrafish (Danio rerio) has many similarities with mammalian in terms of overall organization and the functioning of their stress regulatory systems. The zebrafish hypothalamic-pituitary-interrenal (HPI), analogous to the mammalian hypothalamic-pituitary-adrenal (HPA), has been found in zebrafish and the activation of this pathway is essential for the maintenance of vital functions in response to stressful events. Despite extensive knowledge regarding stress responses in mammals, data on the relationship between chronic unpredictable stress and its effects on purinergic signaling is limited. Purinergic signaling is mediated by ATP and adenosine receptor activation through P2 and P1, respectively. The catabolism of extracellular ATP to adenosine is promoted by the ectonucleotidases, such as Ecto-nucleoside triphosphate diphosphohydrolases (E-NTPDases) and ecto-5 '-nucleotidases (E-5´NT). Adenosine is deaminated by adenosine deaminase (ADA) producing inosine. Thus, these enzymes control the levels of extracellular nucleotides and nucleosides, exerting an influence on purinergic signaling. Considering some models of stress can affect the signalling systems, the objective of this study was to verify whether the unpredictable chronic stress changes ectonucleotidases and adenosine deaminase activities in zebrafish brain. In addition, we analyzed the metabolism of ATP as well as the expression of genes related to ADA, ada1, ada2.1, ada2.2, adaL, adaasi and the effects of agonists and antagonists of adenosine receptors in an unpredictable chronic stress model in zebrafish. Our results demonstrated that there are no changes on ATP (P = 0.1499), ADP (P = 0.3615) and AMP (P = 0.8987) hydrolysis in zebrafish brain membranes submitted to unpredictable chronic stress. In contrast, ecto-ADA activity significantly decreased (26.8%; 8,164 ± 0.78 NH3 min &#8722; 1 mg&#8722; 1 of protein; p 0.05; n = 5 <) in brain membranes of animals exposed to the unpredictable chronic stress when compared to the control group (11.15 ± 2.16 NH3 min &#8722; 1 mg &#8722; 1 of protein; p 0.05; n = 5). However, the activity of cytosolic ADA was not changed after the unpredictable chronic stress. Quantitative analysis by RT-qPCR did not show significant changes after chronic unpredictable stress exposure on gene expression in the genes ada1, ada2.1, ada2.2, adaL, adaasi. ATP metabolism in brain membrane preparations of zebrafish in the control group and stressed was measured by high-performance liquid chromatography (HPLC) and showed a significant increase of adenosine in stressed animals. Considering adenosine A1 receptor agonists have an anxyolitic activity and the A2A receptor antagonists can exert neuroprotective effects, we investigated the effect of administration of an agonist of A1 receptor, N&#8310;-ciclopentiladenosine (CPA), and an antagonist of A2A receptors, (ZM241385) on behavioral parameters of animals submitted to unpredictable chronic stress to check a possible relationship between the adenosinergic system and the stress. As a result we showed that these compounds have not been able to reverse the anxiogenic effect caused by unpredictable chronic stress. Since adenosine has neuromodulatory and anxiolytic effects, changes in its levels could play a role in counteracting the stress, which could be related to a compensatory mechanism in order to restore the homeostasis.