Abstract-Hypertension caused by angiotensin II (Ang II) infusion is associated with oxidative stress in the peripheral vasculature and kidney. The role of redox mechanisms in the central nervous system (CNS), a tissue known to be pivotal in Ang II-dependent hypertension, has not been investigated. ) over a 2-week period in mice caused a gradually developing hypertension that was correlated with marked elevations in O 2 ⅐Ϫ production specifically in the subfornical organ (SFO), a brain region lying outside the blood-brain barrier and known to be a primary sensor for blood-borne Ang II. Adenoviral-mediated delivery of cytoplasmically targeted superoxide dismutase (SOD) selectively to this site prevented the hypertension and the increased O 2 ⅐Ϫ production, whereas gene transfer of SOD targeted to the extracellular matrix had no effect. These data suggest that increased intracellular O 2 ⅐Ϫ production in the SFO is critical in the development of Ang II-induced hypertension. Key Words: reactive oxygen species Ⅲ brain Ⅲ subfornical organ Ⅲ neurons Ⅲ blood pressure V arious forms of hypertension are characterized by an elevation in angiotensin II (Ang II) levels. 1,2 Abundant evidence now suggests that a key mechanism by which Ang II influences blood pressure is via its ability to produce reactive oxygen species (ROS). A decade ago, Griendling et al first discovered that Ang II activates the vascular smooth muscle NAD(P)H oxidase, an important cellular source of ROS. 3 Subsequently, it was shown that hypertension caused by Ang II infusion, but not norepinephrine infusion, increased vascular superoxide (O 2 ⅐Ϫ ) production in vivo 4 and that treatment with liposome-encapsulated superoxide dismutase (SOD) was effective in preventing this form of hypertension. 5 More recent studies demonstrating that genetic disabling of the NADPH oxidase complex attenuates Ang II-induced increases in vascular O 2 ⅐Ϫ production and hypertension further implicates ROS, particularly O 2 ⅐Ϫ

The NADPH oxidase is a multi-subunit enzyme that catalyzes the reduction of molecular oxygen to form superoxide (O(2)(-)). While classically linked to the respiratory burst in neutrophils, recent evidence now shows that O(2)(-) (and associated reactive oxygen species, ROS) generated by NADPH oxidase in nonphagocytic cells serves myriad functions in health and disease. An entire new family of NADPH Oxidase (Nox) homologues has emerged, which vary widely in cell and tissue distribution, as well as in function and regulation. A major concept in redox signaling is that while NADPH oxidase-derived ROS are necessary for normal cellular function, excessive oxidative stress can contribute to pathological disease. This certainly is true in the central nervous system (CNS), where normal NADPH oxidase function appears to be required for processes such as neuronal signaling, memory, and central cardiovascular homeostasis, but overproduction of ROS contributes to neurotoxicity, neurodegeneration, and cardiovascular diseases. Despite implications of NADPH oxidase in normal and pathological CNS processes, still relatively little is known about the mechanisms involved. This paper summarizes the evidence for NADPH oxidase distribution, regulation, and function in the CNS, emphasizing the diversity of Nox isoforms and their new and emerging role in neuro-cardiovascular function. In addition, perspectives for future research and novel therapeutic targets are offered.

Abstract-We have shown that intracellular superoxide (O 2⅐Ϫ ) production in CNS neurons plays a key role in the pressor, bradycardic, and dipsogenic actions of Ang II in the brain. In this study, we tested the hypothesis that a Rac1-dependent NADPH oxidase is a key source of O 2 ⅐Ϫ in Ang II-sensitive neurons and is involved in these central Ang II-dependent effects. We performed both in vitro and in vivo studies using adenoviral (Ad)-mediated expression of dominant-negative Rac1 (AdN17Rac1) to inhibit Ang II-stimulated Rac1 activation, an obligatory step in NADPH oxidase activation. Ang II induced a time-dependent increase in Rac1 activation and O 2 ⅐Ϫ production in Neuro-2A cells, and this was abolished by pretreatment with AdN17Rac1 or the NADPH oxidase inhibitors apocynin or diphenylene iodonium. AdN17Rac1 also inhibited Ang II-induced increases in NADPH oxidase activity in primary neurons cultured from central cardiovascular control regions. In contrast, overexpression of wild-type Rac1 (AdwtRac1) caused more robust NADPH oxidase-dependent O 2 ⅐Ϫ production to Ang II. To extend the in vitro studies, the pressor, bradycardic, and drinking responses to intracerebroventricularly (ICV) injected Ang II were measured in mice that had undergone gene transfer of AdN17Rac1 or AdwtRac1 to the brain. AdN17Rac1 abolished the increase in blood pressure, decrease in heart rate, and drinking response induced by ICV injection of Ang II, whereas AdwtRac1 enhanced these physiological effects. The exaggerated physiological responses in AdwtRac1-treated mice were abolished by O 2 ⅐Ϫ scavenging. These results, for the first time, identify a Rac1-dependent NADPH oxidase as the source of central Ang II-induced O 2 ⅐Ϫ production, and implicate this oxidase in cardiovascular diseases associated with dysregulation of brain Ang II signaling, including hypertension. Key Words: reactive oxygen species Ⅲ dominant-negative Rac1 Ⅲ blood pressure Ⅲ dipsogenic response Ⅲ neurons A ngiotensin II (Ang II), the primary effector peptide of the renin-angiotensin system, acts in the central nervous system (CNS) to modulate neurohumoral pathways involved in water and salt appetite, vasopressin release, and sympathoexcitation. 1 Because dysregulation of central angiotensinergic systems is strongly implicated in cardiovascular diseases such as hypertension and heart failure, 1 elucidating the precise signaling mechanisms of Ang II in the CNS is critical in understanding the pathogenesis of these disorders.We exchange factor-mediated replacement of bound GDP for GTP. 5 Formation and activation of NADPH oxidase allows electrons to be passed from cofactor NADPH to molecular oxygen to produce O 2 ⅐Ϫ . Ang II-induced ROS production via activation of NADPH oxidase is known to be involved in peripheral cell growth, contraction, and inflammation, 6 and dysregulation of redox-mechanisms in the periphery are implicated in the pathogenesis of hypertension caused by systemic Ang II-infusion. 7,8 Recent studies have identified NADPH oxidase components in CN...

We present direct surface force measurements performed using smooth and robust hydrophobic monolayers deposited on mica. From atomic force microscopy measurements we show that these surfaces are composed of large polymerized domains surrounded by a less densely packed, probably gaseous, phase. They exhibit electrical neutrality and stability on exposure to water and aqueous electrolyte solution and are, we suggest, relatively defect free. Despite these characteristics, these novel surfaces do not show the typical longrange hydrophobic interaction that is oRen associated with good quality hydrophobic films. However, we believe that the quality of these surfaces provides the clearest characterization of the interaction between macroscopic hydrophobic surfaces to date and that the novel observation of spontaneous cavitation occurring between these hydrocarbon films is further proof ofthe suitability of these surfaces in such an investigation. IntroductionThe interaction between hydrophobic moieties in water

. Nox2-containing NADPH oxidase and Akt activation play a key role in angiotensin II-induced cardiomyocyte hypertrophy. Physiol Genomics 26: 180 -191, 2006. First published May 2, 2006 doi:10.1152/physiolgenomics.00029.2005.-Angiotensin II (ANG II) has profound effects on the development and progression of pathological cardiac hypertrophy; however, the intracellular signaling mechanisms are not fully understood. In this study, we used genetic tools to test the hypothesis that increased formation of superoxide (O 2 Ϫ ⅐) radicals from a Rac1-regulated Nox2-containing NADPH oxidase is a key upstream mediator of ANG II-induced activation of serine-threonine kinase Akt, and that this signaling cascade plays a crucial role in ANG II-dependent cardiomyocyte hypertrophy. ANG II caused a significant time-dependent increase in Rac1 activation and O 2 Ϫ ⅐ production in primary neonatal rat cardiomyocytes, and these responses were abolished by adenoviral (Ad)-mediated expression of a dominant-negative Rac1 (AdN17Rac1) or cytoplasmic Cu/ZnSOD (AdCu/ZnSOD). Moreover, both AdN17Rac1 and AdCu/ZnSOD significantly attenuated ANG II-stimulated increases in cardiomyocyte size. Quantitative real-time PCR analysis demonstrated that Nox2 is the homolog expressed at highest levels in primary neonatal cardiomyocytes, and small interference RNA (siRNA) directed against it selectively decreased Nox2 expression by Ͼ95% and abolished both ANG II-induced O 2 Ϫ ⅐ generation and cardiomyocyte hypertrophy. Finally, ANG II caused a time-dependent increase in Akt activity via activation of AT 1 receptors, and this response was abolished by Ad-mediated expression of cytosolic human O 2 Ϫ ⅐ dismutase (AdCu/ ZnSOD). Furthermore, pretreatment of cardiomyocytes with dominant-negative Akt (AdDNAkt) abolished ANG II-induced cellular hypertrophy. These findings suggest that O 2 Ϫ ⅐ generated by a Nox2-containing NADPH oxidase is a central mediator of ANG II-induced Akt activation and cardiomyocyte hypertrophy, and that dysregulation of this signaling cascade may play an important role in cardiac hypertrophy. superoxide radicals; renin-angiotensin system; dominant-negative Rac1; small interference RNA; oxidative stress CARDIAC HYPERTROPHY is one of the most significant sequelae of ischemic heart disease, hypertension, and valvular disease (14). Initially an adaptive response that preserves cardiac output and minimizes wall tension, cardiac hypertrophy predisposes to ischemia, arrhythmia, and heart failure. Signal transduction pathways important in the pathogenesis of hypertrophy have been the target of many recent investigations, with angiotensin II (ANG II) emerging as a key molecule both in compensatory cardiac hypertrophy and in the pathogenesis of progressive myocardial dysfunction leading to heart failure (48, 54). Despite the importance of ANG II in cardiac hypertrophy, the signaling cascades downstream of ANG II receptor activation remain to be fully elucidated.The small GTP-binding protein Rac1, a member of the Rho family of GTPases (51), has eme...