Yasuaki Koga

Mitochondria-derived reactive oxygen species mediate sympathoexcitation induced by angiotensin II in the rostral ventrolateral medulla.

Objectives Reactive oxygen species (ROS) in the central nervous system are thought to contribute to sympathoexcitation in cardiovascular diseases such as hypertension and heart failure. Nicotinamide adenine dinucleotide phosphate oxidase is a major source of ROS in the central nervous system, which acts as a key mediator (mediators) of angiotensin II (AngII). It is not clear, however, whether mitochondria-derived ROS in the central nervous system also participate in sympathoexcitation. Methods In an in-vivo study, we investigated whether the AngII-elicited pressor response in the rostral ventrolateral medulla, which controls sympathetic nerve activity, is attenuated by adenovirus-mediated gene transfer of a mitochondria-derived antioxidant (Mn-SOD). In an in-vitro study, using differentiated PC-12 cells with characteristics similar to those of sympathetic neurons, we examined whether AngII increases mitochondrial ROS production. Results Overexpression of Mn-SOD attenuated the AngII-induced pressor response and also suppressed AngII-induced ROS production, as evaluated by microdialysis in the rostral ventrolateral medulla. Using reduced MitoTracker red, we showed that AngII increased mitochondrial ROS production in differentiated PC-12 cells in vitro. Overexpression of Mn-SOD and rotenone, a mitochondrial respiratory complex I inhibitor, suppressed AngII-induced ROS production. Depletion of extracellular Ca2+ with ethylene glycol bis-N,N,N′,N′-tetraacetate (EGTA) and administration of p-trifluoromethoxycarbonylcyanide phenylhydrazone, which prevents further Ca2+ uptake into the mitochondria, blocked AngII-elicited mitochondrial ROS production. Conclusion These results indicate that AngII increases the intracellular Ca2+ concentration and that the increase in mitochondrial Ca2+ uptake leads to mitochondrial ROS production.

Inhibition of Rac1-Derived Reactive Oxygen Species in Nucleus Tractus Solitarius Decreases Blood Pressure and Heart Rate in Stroke-Prone Spontaneously Hypertensive Rats

Reactive oxygen species (ROS) in the brain are thought to contribute to the neuropathogenesis of hypertension by enhancing sympathetic nervous system activity. The nucleus tractus solitarius (NTS), which receives afferent input from baroreceptors, has an important role in cardiovascular regulation. reduced nicotinamide-adenine dinucleotide phosphate oxidase is thought to be a major source of ROS in the NTS. Rac1 is a small G protein and a key component of reduced nicotinamide-adenine dinucleotide phosphate oxidase. The role of Rac1-derived ROS in the NTS in cardiovascular regulation of hypertension is unknown. Therefore, we examined whether inhibition of Rac1 in the NTS decreases ROS generation, thereby reducing blood pressure in stroke-prone spontaneously hypertensive rats (SHRSPs). The basal Rac1 activity level in the NTS was greater in SHRSPs than in Wistar-Kyoto rats. Inhibition of Rac1, induced by transfecting adenovirus vectors encoding dominant-negative Rac1 into the NTS, decreased blood pressure, heart rate, and urinary norepinephrine excretion in SHRSPs but not in Wistar-Kyoto rats. Inhibition of Rac1 also reduced nicotinamide-adenine dinucleotide phosphate oxidase activity and ROS generation. In addition, Cu/Zn-superoxide dismutase activity in the NTS of SHRSPs was decreased compared with that of Wistar-Kyoto rats, despite the increased ROS generation. Overexpression of Cu/Zn-superoxide dismutase in the NTS decreased blood pressure and heart rate in SHRSPs. These results indicate that the activation of Rac1 in the NTS generates ROS via reduced nicotinamide-adenine dinucleotide phosphate oxidase in SHRSPs, and this mechanism might be important for the neuropathogenesis of hypertension in SHRSPs.

High Salt Intake Enhances Blood Pressure Increase during Development of Hypertension via Oxidative Stress in Rostral Ventrolateral Medulla of Spontaneously Hypertensive Rats

High salt intake increases blood pressure (BP) in spontaneously hypertensive rats (SHR), and central neural mechanisms are suggested to be involved. Increased generation of reactive oxygen species (ROS) in the rostral ventrolateral medulla (RVLM) contributes to the neural mechanism of hypertension in SHR. We sought to examine whether high salt intake increases hypertension in SHR and whether the increased ROS in the RVLM contributes to this mechanism. Male SHR and Wistar-Kyoto rats (WKY) (6 weeks old) were fed a high-salt diet (8%: HS-S; HS-W) or a regular-salt diet (0.5%: RS-S; RS-W) for 6 weeks. Systolic BP was significantly higher in HS-S than in RS-S at 12 weeks of age (244±5 vs. 187±7 mmHg, n=8; p<0.05). Urinary norepinephrine excretion was significantly higher in HS-S than in RS-S. Thiobarbituric acid-reactive substances levels in the RVLM were significantly higher in HS-S than in RS-S (9.9±0.5 vs. 8.1±0.6 μmol/g wet wt, n=5; p<0.05). Microinjection of tempol or valsartan into the RVLM induced significantly greater BP reduction in HS-S than in RS-S. The increase in angiotensin II type 1 receptor (AT1R) expression and the increase in reduced nicotinamide-adenine dinucleotide phosphate (NAD(P)H) oxidase activity in the RVLM were significantly greater in HS-S than in RS-S. These findings indicate that high salt intake exacerbates BP elevation and sympathetic nervous system activity during the development of hypertension in SHR. These responses are mediated by increased ROS generation that is probably due to upregulation of AT1R/NAD(P)H oxidase in the RVLM.

Azelnidipine decreases sympathetic nerve activity via antioxidant effect in the rostral ventrolateral medulla of stroke-prone spontaneously hypertensive rats

The long-acting dihydropyridine calcium channel blocker, azelnidipine, is suggested to inhibit sympathetic nerve activity. We previously demonstrated that oxidative stress in the rostral ventrolateral medulla (RVLM) activates sympathetic nerve activity. The aim of the present study was to determine whether oral administration of azelnidipine inhibits sympathetic nerve activity and if so to determine whether the effect is mediated by antioxidant effect in the RVLM. Azelnidipine, hydralazine, or vehicle was orally administered for 28 days to stroke-prone spontaneously hypertensive rats. Reductions in systolic blood pressure were similar in azelnidipine and hydralazine groups. Heart rate was significantly higher in the hydralazine group than in the control, but not altered in the azelnidipine group. Urinary norepinephrine excretion as an indicator of sympathetic nerve activity was significantly lower in the azelnidipine group, whereas it was significantly higher in the hydralazine group than in the control. Levels of thiobarbituric acid-reactive substances and nicotinamide adenine dinucleotide phosphate oxidase activity were significantly lower in the azelnidipine group than in control. Superoxide dismutase activity was significantly increased in the azelnidipine group more than in the control. These results suggest that azelnidipine decreases an indicator of sympathetic nerve activity by antioxidant effect mediated through inhibition of nicotinamide adenine dinucleotide phosphate oxidase activity and activation of superoxide dismutase in the RVLM of stroke-prone spontaneously hypertensive rats.