Ang II action within the kidneys is implicated in may models of human hypertension. When infused at initially subpressor doses, Ang II causes a slow pressor response with potent constriction of the renal afferent arteriole that increases over time. We will examine the concept that the reactivity of the renal afferent arteriole to Ang II is dependent on the generation of reactive oxygen species (ROS) derived from p47/phox- dependent NAD(P)H oxidase. These effects of oxidative stress are countered by oxidant defense mechanisms mediated via extracellular superoxide dismutase (EC-SOD) and intracellularly via a signaling cascade that is regulated by the constitutively-active dopamine 5 receptor (D5-R), We will test whether the functional effect of oxidative stress in the renal afferent arteriole is to enhance tone by reducing eNOS-derived NO. In contrast, the tone of the mesenteric resistance vessels may be blunted by an endothelium-dependent hyperpolarizing factor (EDHF) whose response may actually be mediated by an ROS. These differential effects of oxidative stress could provide for selective effects of Ang II mediated via ROS, on renal afferent arterioles. Studies will contrast isolated afferent and mesenteric resistance vessels from mice. Measurement of contraction or relaxation will be related to measurements of vascular [NO] and [ROS] assess by fluorescence microscopy and to measurements of the mRNA and protein expression of key mediators in these vessels.
The first aim will utilize p47/phox NAD(P)H oxidase and eNOS Knockout mice to define the roles of these systems in the acute microvascular responses to Ang II.
The second aim will assess the expression of key oxidases and antioxidant and pathways in microvessels during prolonged Ang II infusion.
The third aim will examine the functional consequences of finding sin Aim 2. It will contrast acetylcholine-induced relaxation in afferent and mesenteric arterioles, and study the effect of O2-dependent changes in ROS generation. Arterioles will be dissected from mice during prolonged infusion of Ang II or vehicle to related relaxation to vascular [NO] and [ROS] in models deleted in eNOS, p47/phox, EC-SOD and D5-R. These protocols are part of an integrated approach to studying the roles of ROS in the renal mechanism of Ang II-induced hypertension.
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