Exercise training attenuates renovascular hypertension partly via RAS- ROS- glutamate pathway in the hypothalamic paraventricular nucleus

www.nature.com/scientificreports OPEN received: 17 May 2016 accepted: 27 October 2016 Published: 24 November 2016 Exercise training attenuates renovascular hypertension partly via RAS- ROS- glutamate pathway in the hypothalamic paraventricular nucleus Yan Zhang1,*, Xiao-Jing Yu1,*, Wen-Sheng Chen2, Hong-Li Gao1, Kai-Li Liu1, Xiao-Lian Shi3, Xiao-Yan Fan1, Lin-Lin Jia1, Wei Cui4, Guo-Qing Zhu5, Jin-Jun Liu1 & Yu-Ming Kang1,* Exercise training (ExT) has been reported to benefit hypertension; however, the exact mechanisms involved are unclear. We hypothesized that ExT attenuates hypertension, in part, through the reninangiotensin system (RAS), reactive oxygen species (ROS), and glutamate in the paraventricular nucleus (PVN). Two-kidney, one-clip (2K1C) renovascular hypertensive rats were assigned to sedentary (Sed) or treadmill running groups for eight weeks. Dizocilpine (MK801), a glutamate receptor blocker, or losartan (Los), an angiotensin II type1 receptor (AT1-R) blocker, were microinjected into the PVN at the end of the experiment. We found that 2K1C rats had higher mean arterial pressure (MAP) and renal sympathetic nerve activity (RSNA). These rats also had excessive oxidative stress and overactivated RAS in PVN. Eight weeks of ExT significantly decreased MAP and RSNA in 2K1C hypertensive rats. ExT inhibited angiotensin-converting enzyme (ACE), AT1-R, and glutamate in the PVN, and angiotensin II (ANG II) in the plasma. Moreover, ExT attenuated ROS by augmenting copper/zinc superoxide dismutase (Cu/Zn-SOD) and decreasing p47phox and gp91phox in the PVN. MK801or Los significantly decreased blood pressure in rats. Together, these findings suggest that the beneficial effects of ExT on renovascular hypertension may be, in part, through the RAS-ROS-glutamate pathway in the PVN. Recent studies indicate that exercise training (ExT) is beneficial to hypertension in patients and animals1,2. The favourable effect of exercise training is due, in part, to decreased sympathetic activity and improved autonomic function3,4. Evidence suggests that ExT is associated with neuronal plasticity in the brain, which regulates blood pressure5,6. The role of ExT on glutamate within the rostral ventrolateral medulla (RVLM) and the associated improvement in sympathetic outflow has been extensively demonstrated in hypertension7. In other studies, ExT restores the balance between excitatory and inhibitory neurotransmitters and between pro- and anti-inflammatory cytokines, attenuates total reactive oxygen species (ROS) and superoxide production, and increases antioxidants within the paraventricular nucleus (PVN) of spontaneously hypertensive rats (SHR)8,9. The rennin-angiotensin system (RAS) is involved in the pathophysiology of renovascular hypertension10–12. It has been reported that 2K1C (two-kidney, one-clip) renovascular hypertensive rats show a significant increase in mRNA and protein expression of the angiotensin II type 1 receptor (AT1-R) and angiotensin-converting enzyme 1 Department of Physiology and Pathophysiology, Key Laboratory of Environment and Genes Related to Diseases, Xi’an Jiaotong University School of Basic Medical Sciences, Xi’an Jiaotong University Health Science Center, Xi’an Jiaotong University Cardiovascular Research Center, Xi’an, 710061, China. 2Department of Cardiovascular Surgery, Xijing Hospital, Fourth Military Medical University, Xi’an, 710032, China. 3Department of Pharmacology, School of Basic Medical Sciences, Xi’an Jiaotong University Health Science Center, Xi’an, 710061, China. 4Department of Endocrinology and Metabolism, The First Affiliated Hospital of Xi’an Jiaotong University, Xi’an Jiaotong University Health Science Center, Xi’an, 710061, China. 5Key Laboratory of Cardiovascular Disease and Molecular Intervention, Department of Physiology, Nanjing Medical University, Nanjing, 210029, China. *These authors contributed equally to this work. Correspondence and requests for materials should be addressed to X.-J.Y. (email: xiaojingyu621@163. com) or J.-J.L. (email: ) or Y.-M.K. (email: ) Scientific Reports | 6:37467 | DOI: 10.1038/srep37467 1 www.nature.com/scientificreports/ (ACE) within the PVN13. Recent studies indicate that RAS in the PVN exerts its actions mainly via interaction with AT1-R and ACE, thereby contributing to sympathoexcitation and hypertensive response in hypertension13. Our study, along with others, has shown that AT1-R in the PVN induces mitochondria dysfunction and produces excessive amounts of ROS in peripheral angiotensin II (ANGII)-induced hypertension rats14,15. Glutamate is a well-known excitatory neurotransmitter, which participates in regulating neuronal excitation in the central nervous system (CNS). Neuronal activity in the PVN is regulated by glutamate and other excitatory neurotransmitters16,17. Previous studies show that oxidative stress contributes to modulating glutamatergic output in the PVN in hypertension rats18. These data suggest that RAS induces gene transcription of ROS, which leads to further glutamatergic output, and eventually to accelerated progression of hypertension. PVN is a key site for the central control of sympathetic outflow and a predominant region for coordinating nervous system signals that regulate blood pressure, which plays a crucial role in renovascular hypertension10,19. Few studies on ExT in 2K1C hypertension models have focused on RAS, ROS, or glutamate within the PVN. Here, we test the hypothesis that ExT decreases blood pressure in renovascular hypertensive rats. Furthermore, we hypothesize that the favourable effect of exercise will be, in part, associated with RAS, ROS, and glutamate within the PVN of renovascular hypertensive rats. Methods Animal care. Experiments were performed in male Sprague-Dawley rats (eight weeks old and weighing 180–210 g). All rats were housed in a condition-controlled (21–23 °C, with the lights on from 7 pm to 7 am) room. They were permitted free access to standard rat chow and tap water. The rats were treated in accordance with the principles of the National Institutes of Health Guide for the Care and Use of Laboratory Animals (the US National Institutes of Health Publication No. 85–23, revised 1996). All protocols were approved by the Animal Care and Use Committee at Xi’an Jiaotong University. Renal artery clipping. Eight-week-old rats were anesthetized with xylazine (10 mg/kg) and ketamine (90 mg/kg) through intraperitoneal (i.p.) injection. Then, the rats were secured on the operating table, a right-flank incision was made in the abdomen, a silver clip (0.2 mm) was placed around the right renal artery, and then the flank incision was closed. Sham-clipped (Sham) rats underwent identical surgery without the silver clip. At the end of surgery, each rat received butorphanol tartrate (0.2 mg/kg subcutaneously) for an analgesic and penicillin for disinfection19,20. Exercise training. Four or five days after sham or renal artery clipping, the rats were r (...truncated)