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DEHYDRATION-INDUCED carbon monoxide generation: participation of ROS, redox signaling and the NRF2-ARE pathway

Grant number: 17/13653-3
Support type:Scholarships abroad - Research Internship - Doctorate
Effective date (Start): October 01, 2017
Effective date (End): May 31, 2018
Field of knowledge:Biological Sciences - Physiology - Physiology of Organs and Systems
Principal Investigator:José Antunes Rodrigues
Grantee:Juliana Bezerra Medeiros de Lima
Supervisor abroad: Matthew Zimmerman
Home Institution: Faculdade de Medicina de Ribeirão Preto (FMRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil
Local de pesquisa : University of Nebraska Omaha (UNO), United States  
Associated to the scholarship:14/10060-3 - Participation of intracellular pathways modulated by carbon monoxide in the regulation of hydroeletrolitic balance, BP.DR

Abstract

It has been known that heme oxygenase (HO) inhibition compromises not only the carbon monoxide (CO) production but also biliverdin, one of the most important brain antioxidant, and iron, which its homeostasis is implicated with the gene expression. The increased angiotensin II (ANG II) in dehydrated animals (as previous demonstrated by this and several other studies) activates its receptor (AT1R) in hypothalamic areas involved in the hydromineral balance neuroendocrine control. AT1R activation leads to the formation of reactive oxygen species indicating that the redox signaling pathway could be involved in the neuroendocrine responses to dehydration. Therefore, the following hypothesis has been raised, the increase in HO expression and enzymatic activity observed in our previous studies (principal project) is related not only to the glutamatergic stimulation of magnocellular neurons during dehydration but also in response to the dehydration-induced ROS generation. This hypothesis is also supported by our previous data obtained in the principal project that dehydration induces the increase in neuronal nitric oxide synthase (nNOS) expression in the supraoptic nucleus (SON) and in its enzymatic activity in the medial basal hypothalamus (MBH). It is well known that nitric oxide (NO*) can inhibit the mitochondrial electron transport chain leading to the increase in ROS production. Furthermore, CO, one HO byproduct, itself can inhibit the mitochondrial electron transport chain, also increasing ROS generation. We have observed an increase in HO activity in rats submitted to 24 hours of dehydration and an increase in its expression in 48-hour water deprived rats. We have also observed that the nuclear factor erythroid 2-related factor 2 (Nrf-2) expression is increased in the SON of animals submitted to water deprivation, and this is the main transcription factor involved in the antioxidant response.In this way, the current project aims to analyze the redox environment in the SON of mice submitted to dehydration and evaluates the antioxidants s role in the neuroendocrine response to water deprivation (such as plasma AVP e OT concentrations, water intake, plasma protein and osmolality) using a transgenic mouse model (Nrf2-floxed mice) which will be conditionally deleted in SON for the main transcription factor involved in antioxidant response, the Nrf2. The expression of Cu/Zn-superoxide dismutase (SOD1), Mn-superoxide dismutase (SOD2), catalase, NAD(P)H:quinone acceptor oxidoreductase 1 (NQO1), heme oxygenase 1 (HO-1) and heme oxygenase 2 (HO-2) antioxidant enzymes will be evaluated through qPCR and Western Blotting; the superoxide formation will be measured through EPR spectroscopy; plasma AVP and OT will be measured by commercial ELISA kits; and the ingestive behavior (water intake) will be measured in metabolic cages.The internship will last 8 months (from October 2017 to May 2018) under the supervision of Dr. Matthew C. Zimmerman (Associate Professor and Vice-Chair Cellular & Integrative Physiology; Director, Free Radicals in Medicine Program; Director, EPR Spectroscopy Core) and Dr. Irving H. Zucker (F.A.H.A.; F.A.P.S.; Theodore F. Hubbard Professor of Cardiovascular Research; Chairman, Cellular & Integrative Physiology; Editor-In-Chief, The American Journal of Physiology-Heart & Circulatory Physiology) at University of Nebraska Medical Center. These are two prominent researchers whose expertise will guarantee the well-development of the current project with state-of-art technologies (such as conditional gene deletion and electron paramagnetic spectroscopy).