Background-Angiotensin II-induced hypertension is associated with NAD(P)H oxidase-dependent superoxide production in the vessel wall. Vascular superoxide level is also increased in deoxycorticosterone acetate (DOCA)-salt hypertension, which is associated with a markedly depressed plasma renin activity because of sodium retention. However, the mechanisms underlying superoxide production in low-renin hypertension are undefined. Methods and Results-This study investigated (1) whether and how endothelin-1 (ET-1), which is increased in DOCA-salt hypertensive rats, contributes to arterial superoxide generation and (2) the effect of gene transfer of manganese superoxide dismutase and endothelial nitric oxide synthase. Both superoxide and ET-1 levels were significantly elevated in carotid arteries of DOCA-salt rats compared with that of the sham-operated controls. ET-1 concentration-dependently stimulated superoxide production in vitro in carotid arteries of normotensive rats. The increase in arterial superoxide in both ET-1-treated normotensive and DOCA-salt rats was reversed by a selective ET A receptor antagonist, ABT-627, the flavoprotein inhibitor diphenyleneiodonium, and the NADPH oxidase inhibitor apocynin but not by the nitric oxide synthase inhibitor N -L-arginine methyl ester or the xanthine oxidase inhibitor allopurinol. Furthermore, in vivo blockade of ET A receptors significantly reduced arterial superoxide levels, with a concomitant decrease of systolic blood pressure in DOCA-salt rats. Ex vivo gene transfer of manganese superoxide dismutase or endothelial nitric oxide synthase also suppressed superoxide levels in carotid arteries of DOCA-salt rats. Conclusions-These findings suggest that ET-1 augments vascular superoxide production at least in part via an ET A /NADPH oxidase pathway in low-renin mineralocorticoid hypertension.

Development of techniques for the continuous measurement of regional blood flow and vascular resistance in intact small animals has been impeded primarily by the bulkiness of flow probes. The availability of an ultrasonic pulsed Doppler flowmeter system enabled us to construct miniaturized probes using 1-mm-diameter piezoelectric crystals that emit a 20-mHz signal and receive the reflected sound waves from passing blood cells. The finished flow probe is approximately 2.5-4 mm long and 2 mm in cross-sectional diameter with lumen diameters appropriate for the rat, ranging from 0.7 to 1.2 mm. This report describes the materials and methods involved in constructing and implanting the probes in rats to monitor renal, mesenteric, and hindquarter blood flow velocity. The accuracy of the pulsed Doppler method in detecting changes in regional blood flow and vascular resistance was established by the demonstration of a highly significant correlation between velocity recorded from the Doppler unit and volume flow recorded simultaneously. These data indicate that the ultrasonic pulsed Doppler flowmeter provides the opportunity to measure changes in regional blood flow and vascular resistance in a conscious freely moving rat.

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Abstract-Chronic angiotensin II (Ang II) infusion, in rats fed high salt, engages the sympathetic nervous system to increase venomotor tone. The splanchnic sympathetic nervous system is the most important regulator of venous tone, indicating that splanchnic sympathetic nervous system activity may be increased in Ang II salt hypertension. We hypothesized that celiac ganglionectomy (CGx), to selectively disrupt sympathetic innervation to the splanchnic circulation, would attenuate arterial pressure (AP), and venous tone increases in Ang II salt hypertension. Rats fed 2% or 0.4% NaCl were instrumented to allow AP measurement by radiotelemetry at the same time as surgical CGx or sham operation. Ang II was delivered by minipump (150 ng/kg per minute) for 14 days. CGx reduced AP independent of salt diet during control. CGx markedly attenuated Ang II hypertension in rats on 2% NaCl but had little effect in rats fed 0.4% NaCl. To test the possibility that CGx exerted its effects via renal denervation, rats were subjected to the same protocol but received selective bilateral renal denervation. Renal denervation decreased AP during control but had no protective effect on Ang II hypertension and actually tended to exacerbate the pressor response. Finally, separate groups of rats underwent CGx or sham operation and were instrumented to allow repeated measures of mean circulatory filling pressure, an index of venous tone. In addition to attenuating Ang II salt hypertension, CGx completely prevented Ang II salt-induced increases in mean circulatory filling pressure and substantially attenuated depressor responses to acute ganglion blockade. We conclude that, in the presence of high salt, Ang II activates the splanchnic sympathetic nervous system to increase venomotor tone and AP. Key Words: angiotensin II Ⅲ sympathetic nervous system Ⅲ splanchnic circulation Ⅲ venomotor tone Ⅲ renal denervation W e have shown recently, using repeated measures of mean circulatory filling pressure (MCFP) in conscious undisturbed rats, that chronic infusion of angiotensin II (Ang II), only when administered in combination with a high-salt diet, activates the sympathetic nervous system (SNS) to increase venomotor tone. 1 This increase in venomotor tone may contribute to the pathogenesis of Ang II salt hypertension by increasing central blood volume, resulting in a translocation of blood from the highly compliant venous system to the less compliant arterial circulation. 1 This redistribution of blood volume and the well-documented impairment of renal excretory function caused by Ang II 2 would be major factors in increasing arterial pressure (AP) in this model. [3][4][5] Splanchnic veins and venules account for most of the active capacitance responses in the circulation and are richly innervated by the SNS. 6 -8 In fact, it has been estimated that innervation to the nonhepatic splanchnic organs accounts for half of the total norepinephrine (NE) released in the entire body. 9 Therefore, our recent observations in Ang II salt hypertension of neurog...

Objective-Obesity and hypertension are comorbid in epidemic proportion, yet their biological connection is largely a mystery. The peptide chemerin is a candidate for connecting fat deposits around the blood vessel (perivascular adipose tissue) to arterial contraction. We presently tested the hypothesis that chemerin is expressed in perivascular adipose tissue and is vasoactive, supporting the existence of a chemerin axis in the vasculature. Approach and Results-Real-time polymerase chain reaction, immunohistochemistry, and Western analyses supported the synthesis and expression of chemerin in perivascular adipose tissue, whereas the primary chemerin receptor ChemR23 was expressed both in the tunica media and endothelial layer. The ChemR23 agonist chemerin-9 caused receptor, concentration-dependent contraction in the isolated rat thoracic aorta, superior mesenteric artery, and mesenteric resistance artery, and contraction was significantly amplified (more than 100%) when nitric oxide synthase was inhibited and the endothelial cell mechanically removed or tone was placed on the arteries. The novel ChemR23 antagonist CCX832 inhibited phenylephrine-induced and prostaglandin F2α-induced contraction (+perivascular adipose tissue), suggesting that endogenous chemerin contributes to contraction. Arteries from animals with dysfunctional endothelium (obese or hypertensive) demonstrated a pronounced contraction to chemerin-9. Finally, mesenteric arteries from obese humans demonstrate amplified contraction to chemerin-9. Conclusions-These Watts et al Chemerin as a Vasoconstrictor 1321also play a role in obesity. Additionally, chemerin regulates adipocyte differentiation [18][19][20] and production of several proinflammatory cytokines. We hypothesized that a chemerin axis exists in blood vessels. We propose that chemerin and the primary receptor for chemerin, ChemR23, are present and mediate contraction in the vasculature. Materials and MethodsMaterials and Methods are available in the online-only Supplement. Results Arterial Chemerin AxisIsolated rat arteries express chemerin protein in the PVAT ( Figure 1A). Real-time polymerase chain reaction supports the expression of chemerin (RARRES2) mRNA in the rat thoracic aortic PVAT (whole PVAT; threshold cycle [C T ] =22.78±0.35; β2-microglobulin as control = 19.32±0.27; n=6). Chemerin signal does not wholly derive from resident mast cells because there was negligible CD68 staining in PVAT ( Figure 1B, + control below), and staining for chemerin was, in many places, not punctate. Positive staining was observed within the cytoplasm of the fat cell, outside the rounded lipid droplet. The predominant receptor for chemerin, ChemR23, is expressed in the tunica media and endothelial cell layer ( Figure 1C) and is observed as 3 dominant bands in homogenates (−PVAT) of the thoracic aorta and superior mesenteric artery cleaned of PVAT ( Figure 1D and 1E). Two bands (at arrows) are consistent with that observed in a JAR (choriocarcinoma) positive control and were 42 kDa (expected size for...

SUMMARYThe effect of surgical ablation of the area postrema on acute (5-10 minutes) and chronic (5-10 days) increases in mean arterial pressure produced by intravenous infusion of angiotensin II hi conscious, instrumented rats was studied. In agreement with previous studies, pressor responses of area postrema-ablated rats (a = 11) to acute angiotensin II infusion were identical to those of control sham-lesioned rats (n = 13). In these same rats, however, a 5-day infusion of angiotensin II produced a sustained hypertension hi the sham-lesioned group whereas mean arterial pressure was increased only transiently (1-3 days) hi the area postrema-ablated rats. No differences before infusion of arterial pressure, heart rate, water intake, urinary sodium excretion, and urinary potassium excretion were observed between sham-lesioned and area postrema-ablated rats; only arterial pressure was changed significantly during angiotensin II infusion hi either group. Twenty-four hours after terminating angiotensin II infusion, mean arterial pressure was within the normotensive range hi both shamlesioned and area postrema-ablated rats. In a separate group of sham-lesioned (n = 13) and area postrema-ablated (n = 12) rats, angiotensin n was infused intravenously for a 10-day period; mean arterial pressure was increased significantly over the entire 10-day infusion hi sham-lesioned rats, but for only 1 day hi area postrema-ablated rats. An intact area postrema appears necessary for the development of chronic, but not acute, hypertension during intravenous infusion of angiotensin II hi the rat. Supported by National Heart, Lung, and Blood Institute Grants HL24111 andHL32981.Address for reprints: Dr. Gregory D. Fink, Dept. of Pharmacology and Toxicology, Michigan State University, East Lansing, MI 48824.Received January 27, 1986; accepted October 20, 1986. perimental hypertension. A detailed analysis 1 of many of these studies led to the conclusion that ANG II does not cause chronic hypertension by direct vascular constriction, but rather by an ability to stimulate a more slowly developing (hours to days) pressor mechanism: a mechanism whose sensitivity to ANG II increases with prolonged exposure to the hormone. Actions of ANG II on aldosterone secretion, 2 renal handling of salt and water,

1. The peptide hormone angiotensin II (AngII) is acknowledged to be an important factor in the pathophysiology of hypertension. This is particularly the case in hypertension caused by luminal narrowing of one renal artery, (i.e. renovascular hypertension). The primary mechanism by which AngII raises blood pressure, however, is disputed. Strong arguments can be made supporting either vascular contraction, effects on renal excretion of sodium and water, or trophic actions on cardiovascular structures as the key element. In this paper I review evidence that AngII influences blood pressure by modulating autonomic nervous system activity. Modulation can occur at both the peripheral and central aspects of the autonomic system, but I focus on brain pathways involved in determining sympathetic nervous system activity. 2. Experimental and clinical investigations are cited to support the hypothesis that sympathetically mediated pressor effects are increased by both circulating and brain-derived AngII in hypertension. Recent work points specifically to sympathetic pre-motor neurons in the rostral ventrolateral medulla (RVLM) as a critical site of action of brain AngII in normotensive and hypertensive animals. 3. This same set of neurons appears to be an important relay in the sympatho-excitatory response to circulating AngII initiated at circumventricular organs, particularly the area postrema. AngII has important effects on the baroreflex. These do not mediate the sympatho-excitation elicited by circulating AngII, but rather mask its expression. 4. Substantial data support the hypothesis that increased blood concentrations of AngII in renovascular hypertension elevate blood pressure by causing neurogenic vasoconstriction mediated through the area postrema and RVLM.

Renal denervation has been shown to lower arterial pressure in some hypertensive patients, yet it remains unclear whether this is due to ablation of afferent or efferent renal nerves. To investigate the role of afferent renal nerves in arterial pressure regulation, previous studies have used methods that disrupt both renal and nonrenal afferent signaling. The present study was conducted to develop and validate a technique for selective ablation of afferent renal nerves that does not disrupt other afferent pathways. To do this, we adapted a technique for sensory denervation of the adrenal gland by topical application of capsaicin and tested the hypothesis that exposure of the renal nerves to capsaicin (renal-CAP) causes ablation of afferent but not efferent renal nerves. Renal-CAP had no effect on renal content of the efferent nerve markers tyrosine hydroxylase and norepinephrine; however, the afferent nerve marker, calcitonin gene-related peptide was largely depleted from the kidney 10 days after intervention, but returned to roughly half of control levels by 7 wk postintervention. Moreover, renal-CAP abolished the cardiovascular responses to acute pharmacological stimulation of afferent renal nerves. Renal-CAP rats showed normal weight gain, as well as cardiovascular and fluid balance regulation during dietary sodium loading. To some extent, renal-CAP did blunt the bradycardic response and increase the dipsogenic response to increased salt intake. Lastly, renal-CAP significantly attenuated the development of deoxycorticosterone acetate-salt hypertension. These results demonstrate that renal-CAP effectively causes selective ablation of afferent renal nerves in rats.