Search

Adenosine can mediate the tubuloglomerular (TGF) response via activation of [A.sub.1] receptors on the afferent arteriole, but both adenosine [A.sub.1] and [A.sub.2] receptors can regulate preglomerular resistance. We tested the hypothesis that adenosine [A.sub.2] receptors offset the effect of [A.sub.1] receptors and modulate the TGF. Maximal TGF responses were measured in male Sprague-Dawley rats as changes in proximal stop-flow pressure ([DELTA][P.sub.SF]) in response to increased perfusion of the loop of Henle (0 to 40 nl/min) with artificial tubular fluid (ATF). The maximal TGF response was studied after 5 rain of intratubular perfusion (10 nl/min) with ATF alone, or with ATF plus the [A.sub.2A] receptor antagonist (ZM-241385; [10.sup.-7] or [10.sup.-5] tool/l), [A.sub.1] receptor antagonist (PSB-36; [10.sup.-8] mol/l), or with a combination of [A.sub.1] (PSB-36; [10.sup.-8] mol/l) and [A.sub.2A] (ZM-241385; [10.sup.-7] mol/1) antagonists. The maximal TGF response ([DELTA][P.sub.SF]) with ATF alone was 11.7 [+ or -] 1.0 mmHg. Specific [A.sub.2] inhibition (low dose) enhanced the maximal TGF response (15.7 [+ or -] 0.8 mmHg; P < 0.01), whereas a high dose (unspecific inhibition) attenuated the response (5.0 [+ or -] 0.4 mmHg; P < 0.001). [A.sub.1] inhibition alone led to a paradoxical TGF response, with an increase in [P.sub.SF] of 3.1 [+ or -] 0.5 mmHg (P < 0.05). Simultaneous application of [A.sub.1] and [A.sub.2] antagonists abolished the TGF response ([DELTA][P..sub.SF]: 0.4 [+ or -] 0.3 mmHg). In conclusion, adenosine [A.sub.2] receptors modulate the TGF response by counteracting the effects of adenosine [A.sub.1] receptors. adenosine receptors; kidney; micropuncture; renal microcirculation doi: 10.1152/ajprenal.00211.2010.

Adenosine 1 receptors (A1AR) in the kidney are expressed in the vasculature and the tubular system. Pharmacological inhibition or global genetic deletion of A1AR causes marked reductions or abolishment of tubuloglomerular feedback (TGF) responses. To assess the function of vascular A1AR in TGF, we generated transgenic mouse lines in which A1AR expression in smooth muscle was augmented by placing A1AR under the control of a 5.38-kb fragment of the rat smooth muscle [alpha]-actin promoter and first intron (12). Two founder lines with highest expression in the kidney [353 [+ or -] 42 and 575 [+ or -] 43% compared with the wild type (WT)] were used in the experiments. Enhanced expression of A1AR at the expected site in these lines was confirmed by augmented constrictor responses of isolated afferent arterioles to administration of the A1AR agonist [N.sup.6]-cyclohexyladenosine. Maximum TGF responses (0-30 nl/min flow step) were increased from 8.4 [+ or -] 0.9 mmHg in WT (n = 21) to 14.2 [+ or -] 0.7 mmHg in A1AR-transgene (tg) 4 (n = 22; P < 0.0001), and to 12.6 [+ or -] 1.2 mmHg in A1AR-tg7 (n = 12; P < 0.02). Stepwise changes in perfusion flow caused greater numerical TGF responses in A1AR-tg than WT in all flow ranges with differences reaching levels of significance in the intermediate flow ranges of 7.5-10 and 10-15 nl/min. Proximal-distal single-nephron glomerular filtration rate (SNGFR) differences (free-flow micropuncture) were also increased in A1AR-tg, averaging 6.25 [+ or -] 1.5 nl/min compared with 2.6 [+ or -] 0.51 nl/min in WT (P = 0.034). Basal plasma renin concentrations as well as the suppression of renin secretion after volume expansion were similar in A1AR-tg and WT mice, suggesting lack of transgene expression in juxtaglomerular cells. These data indicate that A1AR expression in vascular smooth muscle cells is a critical component for TGF signaling and that changes in renal vascular A1AR expression may determine the magnitude of TGF responses. single nephron glomerular filtration rate; smooth muscle; juxtaglomerular apparatus doi: 10.1152/ajprenal.00264.2009.

We reported impaired endotheliumderived relaxation factor/nitric oxide (EDRF/NO) responses and constitutive nitric oxide synthase (cNOS) activity in subcutaneous vessels dissected from patients with essential hypertension (n = 9) compared with normal controls (n = 10). We now test the hypothesis that the patients in this study have increased circulating levels of the cNOS inhibitor, asymmetric dimethylarginine (ADMA), or the lipid peroxidation product of linoleic acid, 13-hydroxyoctadecadienoic acid (HODE), which is a marker of reactive oxygen species. Patients had significantly (P < 0.001) elevated (means [+ or -] SD) plasma levels of ADMA (PADMA, 766 [+ or -] 217 VS. 393 [+ or -] 57 nmol/1) and symmetric dimethylarginine (PsDMA: 644 [+ or -] 140 VS. 399 [+ or -] 70 nmol/1) but similar levels of L-arginine accompanied by significantly (P < 0.015) increased rates of renal ADMA excretion (21 [+ or -] 9 vs. 14 [+ or -] 5 nmol/ [micro] mo1 creatinine) and decreased rates of renal ADMA clearance (18 [+ or -] 3 vs. 28 [+ or -] 5 ml/min). They had significantly increased plasma levels of HODE (PHODE: 309 [+ or -] 30 vs. 226 [+ or -] 24 nmol/1) and renal HODE excretion (433 [+ or -] 93 vs. 299 [+ or -] 67 nmol/[micro]mol creatinine). For the combined group of normal and hypertensive subjects, the individual values for plasma levels of ADMA and HODE were both significantly (P < 0.001) and inversely correlated with microvascular EDRF/NO and positively correlated with mean blood pressure. In conclusion, elevated levels of ADMA and oxidative stress in a group of hypertensive patients could contribute to the associated microvascular endothelial dysfunction and elevated blood pressure. blood pressure; 13-hydroxyoctadecadienoic acid; endothelium-derived relaxation factor/nitric oxide; albuminuria

Acute intravenous Tempol reduces mean arterial pressure (MAP) and heart rate (HR) in spontaneously hypertensive rats. We investigated the hypothesis that the antihypertensive action depends on generation of hydrogen peroxide, activation of heme oxygenase, glutathione peroxidase or potassium conductances, nitric oxide synthase, and/or the peripheral or central sympathetic nervous systems (SNSs). Tempol caused dose-dependent reductions in MAP and HR (at 174 [micro]mol/kg; [DELTA]MAP, -57 [+ or -] 3 mmHg; and [DELTA]HR, -50 [+ or -] 4 beats/min). The antihypertensive response was unaffected by the infusion of a pegylated catalase or by the inhibition of catalase with 3-aminotriazole, inhibition of glutathione peroxidase with buthionine sulfoximine, inhibition of heme oxygenase with tin potassium channels with iberiotoxin. However, the antihypertensive response was significantly (P < 0.01) blunted by 48% by the activation of adenosine 5'-triphosphate-sensitive potassium ([K.sub.ATP]) channels with cromakalim during maintenance of blood pressure with norepinephrine and by 31% by the blockade of these channels with glibenclamide, by 40% by the blockade of nitric oxide synthase with [N.sup.[omega]]-nitro-L-arginine methyl ester (L-NAME), and by 40% by the blockade of ganglionic autonomic neurotransmission with hexamethonium. L-NAME and hexamethonium were additive, but glibenclamide and hexamethonium were less than additive. The central administration of Tempol was ineffective. The acute antihypertensive action of Tempol depends on the independent effects of potentiation of nitric oxide and inhibition of the peripheral SNS that involves the activation of [K.sub.ATP] channels. superoxide dismutase; nitric oxide synthase; sympathetic nervous system; catalase; adenosine 5'-triphosphate-activated potassium channels

Asymmetric ([N.sup.G], [N.sup.G])-dimethylarginine (ADMA) inhibits nitric oxide (NO) synthases (NOS). ADMA is a risk factor for endothelial dysfunction, cardiovascular mortality, and progression of chronic kidney disease. Two isoforms of dimethylarginine dimethylaminohydrolase (DDAH) metabolize ADMA. DDAH-1 is the predominant isoform in the proximal tubules of the kidney and in the liver. These organs extract ADMA from the circulation. DDAH-2 is the predominant isoform in the vasculatare, where it is found in endothelial cells adjacent to the cell membrane and in intracellular vesicles and in vascular smooth muscle cells among the myofibtils and the nuclear envelope. In vivo gene silencing of DDAH-1 in the rat and DDAH +/- mice both have increased circulating ADMA, whereas gene silencing of DDAH-2 reduces vascular NO generation and endothelium-derived relaxation factor responses. DDAH-2 also is expressed in the kidney in the macula densa and distal nephron. Angiotensin type 1 receptor activation in kidneys reduces the expression of DDAH-1 but increases the expression of DDAH-2. This rapidly evolving evidence of isoform-specific distribution and regulation of DDAH expression in the kidney and blood vessels provides potential mechanisms for nephron site-specifiic regulation of NO production. In this review, the recent advances in the regulation and function of DDAH enzymes, their roles in the regulation of NO generation, and their possible contribution to endothelial dysfunction in patients with cardiovascular and kidney diseases are discussed. nitric oxide synthase; hypertension; diabetes mellitus; chronic kidney disease; asymmetric dimethylarginine

Angiotensin (ANG) II activating type 1 receptors (ATIRs) enhances superoxide anion ([O.sub.2.sup.*-]) and arachidonate (AA) formation. AA is metabolized by cyclooxygenases (COXs) to [PGH.sub.2], which is metabolized by thromboxane (Tx)[A.sub.2] synthase to Tx[A.sub.2] or oxidized to 8-isoprostane [PGF.sub.2[alpha]] (8-Iso) by [O.sub.*-]. [PGH.sub.2], Tx[A.sub.2], and 8-Iso activate thromboxane-prostanoid receptors (TPRs). We investigated whether blood pressure in a rat model of early (3 wk) two-kidney, one-clip (2K,1C) Goldblatt hypertension is maintained by [AT.sub.1]Rs or [AT.sub.2]Rs, driving COX-1 or -2-dependent products that activate TPRs. Compared with sham-operated rats, 2K,1C Goldblatt rats had increased mean arterial pressure (MAP; 120 [+ or -] 4 vs. 155 [+ or -] 3 mmHg; P < 0.001), plasma renin activity (PRA; 22 [+ or -] 7 vs. 48 [+ or -] 5 ng x [m1.sup.-1] x [h.sup.-l]; P < 0.01), plasma malondialdehyde (1.07 [+ or -] 0.05 vs. 1.58 [+ or -] 0.16 nmol/1; P < 0.01), and Tx[B.sub.2] excretion (26 [+ or -] 4 vs. 51 [+ or -] 7 ng/24 h; P < 0.01). Acute graded intravenous doses of benazeprilat (angiotensin-converting enzyme inhibitor) reduced MAP at 20 min (-36 [+ or -] 5 mmHg; P < 0.001) and excretion of Tx[A.sub.2] metabolites. Indomethacin (nonselective COX antagonist) or SC-560 (COX-1 antagonist) reduced MAP at 20 min (-25 [+ or -] 5 and -28 [+ or -] 7 mmHg; P < 0.001), whereas valdecoxib (COX-2 antagonist) was ineffective (-9 [+ or -] 5 mmHg; not significant). Losartan ([ATI.sub.1]R antagonist) or SQ-29548 (TPR antagonist) reduced MAP at 150 min (-24 [+ or -] 6 and -22 [+ or -] 3 mmHg; P < 0.001), whereas PD-123319 ([AT.sub.2]R antagonist) was ineffective. Acute blockade of TPRs, COX-1, or COX-2 did not change PRA, but Tx[B.sub.2] generation by the clipped kidney was reduced by blockade of COX-1 and increased by blockade of COX-2.2K, 1C hypertension in rats activates renin, [O.sub.2.sup*-], and vasoconstrictor PGs. Hypertension is maintained by [AT.sub.1]Rs and by COX-1, but not COX-2, products that activate TPRs. renovascular hypertension; angiotensin receptor blocker; thromboxane [A.sub.2]; isoprostane; angiotensin-converting enzyme inhibition

Tempol catalyzes the formation of [H.sub.2][O.sub.2] from superoxide and relaxes blood vessels. We tested the hypothesis that the generation of [H.sub.2][O.sub.2] by tempol in vascular smooth muscle cells during oxidative stress contributes to the vasorelaxation. Tempol and nitroblue tetrazolium (NBT) both metabolize superoxide in vascular smooth muscle cells, but only tempol generates [H.sub.2][O.sub.2]. Rat pressurized mesenteric arteries were exposed for 20 min to the thromboxane-prostanoid receptor agonist, U-46619, or norepinephrine. During U-46619, tempol caused a transient dilation (22 [+ or -] 2%), whereas NBT was ineffective (2 [+ or -] 1%), and neither dilated vessels constricted with norepinephrine, which does not cause vascular oxidative stress. Neither endothelium removal nor blockade of [K.sup.+] channels with 40 mM KCl affected the tempol-induced dilation, but catalase blunted the tempol dilation by 53 [+ or -] 7%. Tempol, but not NBT, increased [H.sub.2][O.sub.2] in rat mesenteric vessels detected with dichlorofluorescein. To test physiological relevance in vivo, topical application of tempol caused a transient dilation (184 [+ or -] 20%) of mouse cremaster arterioles exposed to angiotensin II for 30 min, which was not seen with NBT (9 [+ or -] 4%). The vasodilation to tempol was reduced by 68 [+ or -] 6% by catalase. We conclude that the transient relaxation of blood vessels by tempol after prolonged exposure to U-46619 or angiotensin II is mediated in part via production of [H.sub.2][O.sub.2] and is largely independent of the endothelium and potassium channels. superoxide dismutase; hypertension; angiotensin II; thromboxane

Angiotensin II (ANG II) infusion increases renal superoxide ([O.sup.-.sub.2]) and enhances renal vasoconstriction via macula densa (MD) regulation of tubuloglomerular feedback, but the mechanism is unclear. We targeted the [p22.sup.phox] subunit of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) with small-interfering RNA (siRNA) to reduce NADPH oxidase activity and blood pressure response to ANG II in rats. We compared single nephron glomerular filtration rate (SNGFR) in samples collected from the proximal tubule (PT), which interrupts delivery to the MD, and from the distal tubule (DT), which maintains delivery to the MD, to assess MD regulation of GFR. SNGFR was measured in control and ANG II-infused rats (200 ngx[kg.sup.-1]x[min.suup.-1] for 7 days) 2 days after intravenous injection of vehicle or siRNA directed to [p22.sup.phox] to test the hypothesis that [p22.sup.phox] mediates MD regulation of SNGFR during ANG II. The regulation of SNGFR by MD, determined by PT SNGFR-DT SNGFR, was not altered by siRNA in control rats (control + vehicle, 13 [+ or -] 1, n = 8; control + siRNA, 12 [+ or -] 2 nl/min, n = 8; not significant) but was reduced by siRNA in ANG II-treated rats (ANG II + vehicle, 13 [+ or -] 2, n = 7; ANG II + siRNA, 7 [+ or -] 1 nl/min, n = 8; P < 0.05). We conclude that [p22.sup.phox] and NADPH oxidase regulate the SNGFR during ANG II infusion via MD-dependent mechanisms. renal function; oxidative stress; hypertension; tubuloglomerular feedback; glomerular filtration rate

Tempol is an amphipathic radical nitroxide (N) that acutely reduces blood pressure (BP) and heart rate (HR) in the spontaneously hypertensive rat (SHR). We investigated the hypothesis that the response to nitroxides is determined by SOD mimetic activity or lipophilicity. Groups (n = 6-10) of anesthetized SHRs received graded intravenous doses of Ns: tempol (T), 4-amino-tempo (AT), 4-oxotempo (OT), 4-trimethylammonium-2,2,6,6-tetramethylpiperidine-1-oxyl iodide (CAT-1), 3-carbamoyl-proxyl (3-CP), or 3-carboxyproxyl (3-CTPY). Others received native or liposomal (L) Cu/Zn SOD. T and OT are uncharged, AT is positively charged and cell-permeable, and CAT-1 is positively charged and cell-impermeable. 3-CP and 3-CTPY have five-member pyrrolidine rings, whereas T, AT, OT, and CAT-1 have six-member piperidine rings. T and AT reduced mean arterial pressure (MAP) similarly (-48 [+ or -] 2 mmHg and -55 [+ or -] 8 mmHg) but more (P < 0.05) than OT and CAT-1. 3-CP and 3-CTPY were ineffective. The group mean change in MAP with piperidine Ns correlated with SOD activity (r = -0.94), whereas their E[D.sub.50] correlated with lipophilicity (r = 0.89). SOD and L-SOD did not lower BP acutely but reduced it after 90 min (-32 [+ or -] 5 and -31 [+ or -] 6 mmHg; P < 0.05 vs. vehicle). Pyrrolidine nitroxides are ineffective antihypertensive agents. The antihypertensive response to piperidine Ns is predicted by SOD mimetic action, and the sensitivity of response is by hydrophilicity. SOD exerts a delayed hypotensive action that is not enhanced by liposome encapsulation, suggesting it must diffuse to an extravascular site. tempol; superoxide dismutase; blood pressure; nitric oxide; hypertension.

There is growing evidence that oxidative stress contributes to hypertension. Oxidative stress can precede the development of hypertension. In almost all models of hypertension, there is oxidative stress that, if corrected, lowers BP, whereas creation of oxidative stress in normal animals can cause hypertension. There is overexpression of the [p22.sup.phox] and Nox-1 components of NADPH oxidase and reduced expression of extracellular superoxide dismutase (EC-SOD) in the kidneys of ANG II-infused rodents, whereas there is overexpression of [p47.sup.phox] and [gp91.sup.phox] and reduced expression of intracellular SOD with salt loading. Several mechanisms have been identified that can make oxidative stress self-sustaining. Reactive oxygen species (ROS) can enhance afferent arteriolar tone and reactivity both indirectly via potentiation of tubuloglomerular feedback and directly by microvascular mechanisms that diminish endothelium-derived relaxation factor/nitric oxide responses, generate a cyclooxygenase-2-dependent endothelial-derived contracting factor that activates thromboxane-prostanoid receptors, and enhance vascular smooth muscle cells reactivity. ROS can diminish the efficiency with which the kidney uses [O.sub.2] for [Na.sup.+] transport and thereby diminish the P[o.sub.2] within the kidney cortex. This may place a break on further ROS generation yet could further enhance vasculopathy and hypertension. There is a tight relationship between oxidative stress in the kidney and the development and maintenance of hypertension. reactive oxygen species; superoxide anion; nitric oxide synthase; thromboxaneprostanoid receptors; salt sensitivity

We tested whether chronic ANG II infusion into rats affects descending vasa recta (DVR) contractility, synthesis of superoxide, or synthesis of nitric oxide (NO). Rats were infused with ANG II at 250 ng x [kg.sup.-1] x [min.sup.-1] for 11-13 days. DVR were loaded with dihydroethidium (DHE) to measure superoxide and 3-amino-4-aminomethyl-2',7'-difluorofluorescein (DAFFM) to measure NO. Acute constriction of DVR by ANG II (0.1, 1, and 10 nM) was diminished, and NO generation rate was raised by chronic ANG II infusion. DHE oxidation by DVR from ANG II-infused rats was similar to controls and was significantly higher when NO synthesis was prevented with [N.sup.[omega]]-nitro-L-arginine methyl ester (L-NAME). The superoxide dismutase mimetic Tempol (1 mM) increased NO generation compared with controls. The increased synthesis of NO by chronic ANG II-treated vessels persisted in the presence of Tempol. DVR endothelial cytoplasmic [Ca.sup.2+] response to ACh was diminished by chronic ANG II treatment, but the capacity of ACh to increase NO generation was unaltered. We conclude that DVR generation of superoxide is not affected by chronic ANG II exposure but that basal NO synthesis is increased. DVR superoxide is unlikely to be an important mediator of chronic ANG II slow pressor hypertension in rats. medulla; kidney; microcirculation; calcium; oxidative stress

We tested the hypothesis that superoxide anion ([O.sup.-.sub.2]*) generated in the kidney by prolonged angiotensin II (ANG II) reduces renal cortical P[o.sub.2] and the use of [O.sub.2] for tubular sodium transport ([T.sub.Na]:Q[O.sub.2]). Groups (n = 8-11) of rats received angiotensin II (ANG II, 200 ng x [kg.sup.-1] x [min.sup.-1] sc) or vehicle for 2 wk with concurrent infusions of a permeant nitroxide SOD mimetic 4-hydroxy-2,2,6,6-tetramethylpiperidine 1-oxyl (Tempol, 200 nmol x [kg.sup.-1] x [min.sup.-1]) or vehicle. Rats were studied under anesthesia with measurements of renal oxygen usage and P[O.sub.2] in the cortex and tubules with a glass electrode. Compared with vehicle, ANG II increased mean arterial pressure (107 [+ or -] 4 vs. 146 [+ or -] 6 mmHg; P < 0.001), renal vascular resistance (42 [+ or -] 3 vs. 65 [+ or -] 7 mmHg x [ml.sup.-1] x [min.sup.-1] x 100 [g.sup.-1] ; P < 0.001), renal cortical NADPH oxidase activity (2.3 [+ or -] 0.2 vs. 3.6 [+ or -]] 0.4 nmol [O.sup.-.sub.2]* x [min.sup.-1] x [mg.sup.-1] protein: P < 0.05), mRNA and protein expression for p[22.sup.phox] (2.1- and 1.8-fold respectively; P < 0.05) and reduced the mRNA for extracellular (EC)-SOD (-1.8 fold; P < 0.05). ANG II reduced the P[O.sub.2] in the proximal tubule (39 [+ or -] 1 vs. 34 [+ or -] 2 mmHg; P < 0.05) and throughout the cortex and reduced the [T.sub.Na]:Q[O.sub.2] (17 [+ or -] 1 vs. 9 [+ or -] 2 [micro]mol/[micro]mol; P < 0.001). Tempol blunted or prevented all these effects of ANG II. The effects of prolonged ANG II to cause hypertension, renal vasoconstriction, renal cortical hypoxia, and reduced efficiency of [O.sub.2] usage for [Na.sup.+] transport, activation of NADPH oxidase, increased expression of p[22.sup.phox], and reduced expression of EC-SOD can be ascribed to [O.sup.-.sub.2]* generation because they are prevented by an SOD mimetic. reactive oxygen species; superoxide dismutase; TEMPOL; nitroxides; NADPH oxidase; hypertension

TP receptors regulate renal hemodynamics during angiotensin II slow pressor response. Am J Physiol Renal Physiol 287: F753-F759, 2004. First published June 22, 2004; 10.1152/ajprenal.00423.2003.--We investigated the hypothesis that thromboxane [A.sub.2] (Tx[A.sub.2])-prostaglandin [H.sub.2] receptors (TP-Rs) mediate the hemodynamic responses and increase in reactive oxygen species (ROS) to ANG II (400 ng x [kg.sub.-1] x [min.sub.-1] sc for 14 days) using TP-R knockout (TP -/-) and wild-type (+/+) mice. TP -/- had normal basal mean arterial blood pressure (MAP) and glomerular filtration rate but reduced renal blood flow and increased filtration fraction (FF) and renal vascular resistance (RVR) and markers of ROS (thiobarbituric acid-reactive substances and 8-isoprostane PG[F.sub.2[alpha]]) and nitric oxide (NOx). Infusion of ANG II into TP +/+ increased ROS and thromboxane [B.sub.2] (Tx[B.sub.2]) and increased RVR and FF. ANG II infusion into TP -/- mice reduced ANG I and increased aldosterone but caused a blunted increase in MAP (TP -/-: +6 [+ or -] 2 vs. TP +/+: +15 [+ or -] 3 mmHg) and failed to increase FF, ROS, or Tx[B.sub.2] but increased NOx and paradoxically decreased RVR (-2.1 [+ or -] 1.7 vs. +2.6 [+ or -] 0.8 mmHg*[ml.sup.-1]*[min.sup.-1]*[g.sup.-1]). Blockade of A[T.sub.1] receptor of TP -/- mice infused with ANG II reduced MAP (-8 mmHg) and aldosterone but did not change the RVR or ROS. In conclusion, during an ANG II slow pressor response, A[T.sub.1] receptors activate TP-Rs that generate ROS and prostaglandins but inhibit NO. TP-Rs mediate all of the increase in RVR and FF, part of the increase in MAP, but are not implicated in the suppression of ANG I or increase in aldosterone. TP -/- mice have a basal increase in RVR and FF associated with ROS. thromboxane [A.sub.2]-prostaglandin [H.sub.2] receptor; isoprostane; hypertension; renal vascular resistance

Oxidative stress accompanies angiotensin (ANG) II infusion, but the role of ANG type 1 vs. type 2 receptors (A[T.sub.1]-R and A[T.sub.2]-R, respectively) is unknown. We infused ANG II subcutaneously in rats for 1 wk. Excretion of 8-isoprostaglandin [F.sub.2[alpha]] (8-Iso) and malonyldialdehyde (MDA) were related to renal cortical mRNA abundance for subunits of NADPH oxidase and superoxide dismutases (SODs) using real-time PCR. Subsets of ANG II-infused rats were given the A[T.sub.1]-R antagonist candesartan cilexetil (Cand) or the A[T.sub.2]-R antagonist PD-123,319 (PD). Compared to vehicle (Veh), ANG II increased 8-Iso excretion by 41% (Veh, 5.4 [+ or -] 0.8 vs. ANG II, 7.6 [+ or -] 0.5 pg/24 h; P < 0.05). This was prevented by Cand (5.6 [+ or -] 0.5 pg/24 h; P < 0.05) and increased by PD (15.8 [+ or -] 2.0 pg/24 h; P < 0.005). There were similar changes in MDA excretion. Compared to Veh, ANG II significantly (P < 0.005) increased the renal cortical mRNA expression of p[22.sup.phox] (twofold), Nox-1 (2.6-fold), and Mn-SOD (1.5-fold) and decreased expression of Nox-4 (2.1-fold) and extracellular (EC)-SOD (2.1-fold). Cand prevented all of these changes except for the increase in Mn-SOD. PD accentuated changes in p[22.sup.phox] and Nox-1 and increased p[67.sup.phox]. We conclude that ANG II infusion stimulates oxidative stress via A[T.sub.1]-R, which increases the renal cortical mRNA expression of p[22.sup.phox] and Nox-1 and reduces abundance of Nox-4 and EC-SOD. This is offset by strong protective effects of A[T.sub.2]-R, which are accompanied by decreased expression of p[22.sup.phox], Nox-1, and p[67.sup.phox]. candesartan; isoprostane; angiotensin receptor blocker; PD-123,319; malonyldialdehyde

A TP receptor (TP-R) mimetic causes salt-sensitive hypertension and renal afferent arteriolar vasoconstriction. TP-Rs mediate effects of ANG II on renal vascular resistance and drinking. Therefore, we investigated the hypothesis that thromboxane [A.sub.2] synthase (Tx[A.sub.2]-S) and/or TP-R expression is regulated by salt and/or ANG II. Rats (n = 6) received high-salt (HS) or low-salt (LS) diets. Additional HS-diet rats received ANG II while other HS- and LS-diet rats received the A[T.sub.1] receptor (A[T.sub.1]-R) antagonist losartan. Excretion of thromboxane [B.sub.2] by conscious rats was increased with the HS diet compared with the LS diet (126 [+ or -] 10 vs. 48 [+ or -] 5 pmol/24 h, respectively; P < 0.01). The mRNA abundance for TP-Rs (relative to [beta]-actin) in the kidney cortex was enhanced 30% by the HS diet (P < 0.001) and was reduced 50% by the addition of ANG II (P < 0.001). However, during losartan administration, the effects of salt were reversed; mRNA more than doubled during the LS diet (P < 0.001). Similarly, the mRNA abundance for TP-Rs in the brain stem was reduced by 50% with the addition of ANG II (P < 0.001) and during losartan administration was almost doubled by the LS diet (P < 0.001). The mRNA abundance for Tx[A.sub.2]-S in the kidney cortex also was increased many times with the HS diet (P < 0.001). In contrast, the mRNA for Tx[A.sub.2]-S in the brain was unaffected by salt. ANG II did not affect Tx[A.sub.2]-S at either site. During losartan administration, Tx[A.sub.2]-S increased modestly in the brain stem with the LS diet. mRNA abundance for TP-Rs in the kidney cortex and brain stem is suppressed, by ANG II acting on A[T.sub.1]-Rs. In the absence of A[T.sub.1]-Rs, expression of TP-Rs at both sites is enhanced by LS intake. In contrast, ANG II does not affect the mRNA abundance for Tx[A.sub.2]-S. Expression of Tx[A.sub.2]-S is enhanced by HS intake in the kidney cortex but by LS intake in the brain stem only during losartan administration. Thus TP-Rs are strongly dependent on ANG II acting on AT1-Rs, whereas TxA2-S is regulated differentially in the kidney cortex and brain stem by salt intake. thromboxane [A.sub.2]; prostaglandins; angiotensin II; losartan; angiotensin receptor blocker; thromboxane prostanoid receptor

Central angiotensin II (ANG II) regulates thirst. Because thromboxane [A.sub.2]-prostaglandin [H.sub.2] (TP) receptors are expressed in the brain and mediate some of the effects of ANG II in the vasculature, we investigated the hypothesis that TP receptors mediate the drinking response to intracerebroventricular (icv) injections of ANG II. Pretreatment with the specific TP-receptor antagonist ifetroban (Ifet) decreased water intake with 50 ng/kg icy ANG II (ANG II + Veh, 7.2 [+ or -] 0.7 ml vs. ANG II + Ifet, 2.8 [+ or -] 0.8 ml; n = 5 rats; P < 0.001) but had no effect on water intake induced by hypertonic saline (NaCl + Veh, 8.4 [+ or -] 1.1 ml vs. NaCl + Ifet, 8.9 [+ or -] 1.8 ml; n = 5 rats; P = not significant). Administration of 0.6 [micro]g/kg icy of the TP-receptor agonist U-46,619 did not induce drinking when given alone but did increase the dipsogenic response to a near-threshold dose of 15 ng/kg icv ANG II (ANG II + Veh, 1.1 [+ or -] 0.7 vs. ANG II + U-46,619, 4.5 [+ or -] 0.9 ml; n = 5 rats; P < 0.01). We conclude that central TP receptors contribute to the dipsogenic response to ANG II. drinking; ifetroban; TP receptors; U-46,619

The spontaneously hypertensive rat (SHR) has an increased tubuloglomerular feedback (TGF) and a reduced buffering by juxtaglomerular apparatus (JGA)-derived NO. The hypothesis that these effects are caused by reductions in nitric oxide synthase (NOS) expression or limited availability of L-arginine or tetrahydrobiopterin (BH4). Findings showed that there is a decreased role for NO from neuronal NOS in blunting TGF in SHR which cannot be caused by limited NOS expression or availability of substrate or BH4.

The synthesis of nitric oxide by the neuronal nitric oxide synthase during salt loading in rats is investigated using enzyme inhibition studies and diuresis manipulation using furosemide. The generation of nitric oxide was monitored by measuring the excretion of metabolites. Results reveal that the increase in nitric oxide synthesis is brought about by an increase in the activity of neuronal nitric oxide synthase and solute reabsorption in the macula densa.

Research shows that thromboxane A2 (TxA2) and prostaglandin endoperoxide (PGH2) receptor activity is boosted by high-salt intake in rats. Experiments designed to assess tubuloglomerular feedback responses to TXA2/PGH2 mimetic using male Sprague-Dawley rats are described. TxA2 activity measurements in relation to low and high salt intake are illustrated. Renal vasoconstriction responses to receptor feedback are discussed.

A study was conducted on the role of arginine vasopressin (AVP) in pressor responses during activation of central TxA2/PGH2 receptors. The method involved the injection of an agonist into the lateral cerebral ventricle of rats. The findings suggest that TxA2/PGH2-receptor agonist U-46619 increases blood pressure and releases AVP through brain receptor activation. Moreover, AVP is responsible for one-half of the pressor response.

An investigation of the sympathetic nervous system and hypertension during prolonged thromboxane/prostaglandin H2 receptor activation was conducted to determine its effects on the sympathetic nervous system of rats. The mean arterial pressure (MAP) was measured using implanted flow probes. Results of the study indicated that hypertension during prolonged infusions of U-46619 into salt-loaded rats leads to an increase in total and renal vascular resistance.

The presence of thromboxane A2 (TxA2) and prostaglandin H2 (PGH2) receptors in rat neural and glial cells was analyzed to determine blood pressure responses to the TxA2 mimetic, U-46,619. Intracerebrovascular administration of U-46,619 enhanced the pressor responses of rats fed on a high-salt diet. However, intravenous and intracerebrovascular administration of ifetroban attenuated the effects of U-49,619 on pressor response. The effects of U-46,619 indicated the presence of TxA2/PGH2 receptor messenger RNAs in rat neurons, glial and brain stem.

The hypertensive action of angiotensin II type I (AT1) and thromboxane A2/prostaglandin endoperoxide (TxA2/PGH2) receptor antagonists was analyzed in two-kidney,one-clip (2k,1C) Goldblatt rat models of hypertension. A marked reduction in blood pressure during the early, intermediate and late phase of renovascular hypertension was detected in 2K,1C Goldblatt rats after the three-day administration of AT1 or TxA2/PGH2 receptors antagonist. However, hypertension from the early to the late phase was reversed by lorsartan and ifetroban.

Renal blood flow (RBF) rates measured using miniature ultrasonic transit-time flow probes that are calibrated using excised arteries at different hematocrit (Hct) and flow rates exhibit a high correlation with true flow rates at subnormal and normal Hct values. In vivo RBF measurements with the excretion and extraction of P-aminohippuric acid correlate well over a broad range of flow rates, indicating that the use of ultrasonic transit-time flow meter enables accurate measurement of RBF in anesthetized, instrumented rats.

Kitiyakara, Chagriya, Tina Chabrashvili, Pedro Jose, William J. Welch, and Christopher S. Wilcox. Effects of dietary salt intake on plasma arginine. Am J Physiol Regulatory Integrative Comp Physiol 280: R1069-R1075, 2001.--Because L-arginine is degraded by hepatic arginase to ornithine and urea and is transported by the regulated 2A cationic amino acid [y.sup.+] transporter (CAT2A), hepatic transport may regulate plasma arginine concentration. Groups of rats (n = 6) were fed a diet of either low salt (LS) or high salt (HS) for 7 days to test the hypothesis that dietary salt intake regulates plasma arginine concentration and renal nitric oxide (NO) generation by measuring plasma arginine and ornithine concentrations, renal NO excretion, and expression of hepatic CAT2A, and arginase. LS rats had lower excretion of NO metabolites and cGMP, lower plasma arginine concentration (LS: 83 [+ or -] 7 vs. HS: 165 [+ or -] 10 [micro]mol/l, P [is less than] 0.001), but higher plasma ornithine concentration (LS: 82 [+ or -] 6 vs. HS: 66 [+ or -] 4 [micro]mol/l, P [is less than] 0.05) and urea excretion. However, neither the in vitro hepatic arginase activity nor the mRNA for hepatic arginase I was different between groups. In contrast, LS rats had twice the abundance of mRNA for hepatic CAT2A (LS: 3.4 [+ or -] 0.4 vs. HS: 1.6 [+ or -] 0.5, P [is less than] 0.05). The reduced plasma arginine concentration with increased plasma ornithine concentration and urea excretion during LS indicates increased arginine metabolism by arginase. This cannot be ascribed to changes in hepatic arginase expression but may be a consequence of increased hepatic arginine uptake via CAT2A. nitric oxide; system [y.sup.+] transport; cationic amino acid transporter; ornithine; urea

Schnackenberg, Christine G., William J. Welch, and Christopher S. Wilcox. TP receptor-mediated vasoconstriction in microperfused afferent arterioles: roles of [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] and NO. Am J Physiol Renal Physiol 279: F302-F308, 2000.--Thromboxane [A.sub.2] (Tx[A.sub.2]) preferentially constricts the renal afferent arteriole. Nitric oxide (NO) modulates vasoconstriction and is rapidly degraded by superoxide radical ([MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII]). We investigated the roles of NO and [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] in rabbit isolated, perfused renal afferent arteriole responses to the Tx[A.sub.2]/prostaglandin [H.sub.2] (TP) receptor agonist U-46,619. U-46,619 ([10.sup.-10] - [10.sup.-6] M) dose-dependently reduced afferent arteriolar luminal diameter ([ED.sub.50] = 7.5 [+ or -] 5.0 nM), which was blocked by the TP receptor antagonist ifetroban ([10.sup.-6] M). Tempol ([10.sup.-3] M) pretreatment, which prevented paraquat-induced vasoconstriction in afferent arterioles, blocked the vasoconstrictor responses to U-46,619. To test whether U-46,619 stimulates NO and whether tempol prevents U-46,619-induced vasoconstriction by enhancing the biological activity of NO, we examined the luminal diameter response to U-46,619 in arterioles pretreated with [N.sup.w]-nitro-L-arginine methyl ester (L-NAME, [10.sup.-4] M) or L-NAME + tempol. During L-NAME, the sensitivity and maximal responses of the afferent arteriole to U-46,619 were significantly (P [is less than] 0.05) enhanced. Moreover, L-NAME restored a vasoconstrictor response to U-46,619 in vessels pretreated with tempol. In conclusion, in isolated perfused renal afferent arterioles TP receptor activation stimulates NO production, which buffers the vasoconstriction, and stimulates [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] production, which mediates the vasoconstriction, in part, through interaction with NO. thromboxane [A.sub.2]; nitric oxide; superoxide; afferent arteriole; tempol

Roles of NO and oxygen radicals in tubuloglomerular feedback in SHR. Am J Physiol Renal Physiol 278: F769-F776, 2000.--The spontaneously hypertensive rat (SHR) has enhanced tubuloglomerular feedback (TGF) responses and diminished buffering by juxtaglomerular apparatus (JGA)-derived nitric oxide (NO) despite enhanced expression of NO synthase (NOS) isoforms in the JGA. We tested the hypothesis that the enhanced TGF response is due to inactivation of NO by oxygen radicals ([MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII]). SHR had significantly (P [is less than] 0.05) greater expression of the peroxynitrate reaction product, nitrotyrosine, in renal cortex. A membrane-permeant, metal-independent superoxide dismutase mimetic, tempol, was used to test the functional role of [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII]. Maximum TGF responses, assessed from changes in proximal stop-flow pressure ([P.sub.SF]) during orthograde loop of Henle (LH) perfusion of artificial tubular fluid (ATF), were enhanced in SHR [Wistar-Kyoto rat (WKY) 8.8 [+ or -] 0.4 (n = 30 nephrons) vs. SHR 10.8 [+ or -] 0.4 mmHg (n = 39 nephrons), P [is less than] 0.001]. TGF responses of SHR were unresponsive to microperfusion of 7-nitroindazole (7-NI, [10.sup.-4] M), which is an inhibitor of neuronal NOS (nNOS) [WKY 8.3 [+ or -] 0.3 to 10.8 [+ or -] 0.4 (n = 8, P [is less than] 0.001) vs. SHR 10.0 [+ or -] 0.7 to 10.5 [+ or -] 0.8 mmHg (n = 8; not significant)]. Microperfusion of tempol ([10.sup.-4] M) into the efferent arteriole (EA) supplying the peritubular capillaries (PTC) blunted TGF. The response to tempol was significantly (P [is less than] 0.05) greater in SHR [[Delta] TGF in WKY 19 [+ or -] 6% (n = 10) vs. SHR 32 [+ or -] 3% (n = 10)]. Microperfusion of the NO donor compound S-nitroso-N-acetyl-penicillamine (SNAP, [10.sup.-7]-[10.sup.-4] M) via the LH blunted TGF, but the sensitivity of the response was impaired significantly (P [is less than] 0.05) in SHR nephrons. PTC perfusion of tempol ([10.sup.-4] M) normalized the response to loop perfusion of both SNAP and 7-NI in SHR nephron to levels in WKY (during tempol, [[Delta] [P.sub.SF]] with 7-NI in WKY 8.9 [+ or -] 0.6 to 11.4 [+ or -] 0.8; n = 12 vs. SHR 9.5 [+ or -] 0.5 to 12.5 [+ or -] 0.4 mmHg; n = 16). In conclusion, TGF responses are enhanced in SHR, in part due to a diminished role for NO from nNOS in blunting TGF due to enhanced [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] formation. [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] in the JGA enhances TGF responses by inactivation of locally generated NO. 7-nitroindazole; superoxide dismutase; tempol; S-nitroso-N-acetyl-penicillamine; nitrotyrosine; nitric oxide synthase

Nitric oxide prevents hypertension yet enhances proximal tubule Na+ reabsorption. Nitric oxide synthase is inhibited by asymmetric dimethylarginine (ADMA) that is metabolized by dimethylarginine dimethylaminohydrolase (DDAH) whose type 1 isoform is expressed abundantly in the proximal tubule (PT). We hypothesize that ADMA metabolized by DDAH-1 inhibits fluid reabsorbtion (Jv) by the proximal tubule. S2 segments of the PT were microperfused between blocks in vivo to assess Jv in anesthetized rats. Compared with vehicle, microperfusion of ADMA or Nω -nitro-l-arginine methyl ester (l-NAME) in the proximal tubule reduced Jv dose dependently. At 10 -4 mol/l both reduced Jv by ~40% (vehicle: 3.2 ± 0.7 vs. ADMA: 2.1 ± 0.5, P < 0.01 vs. l-NAME: 1.9 ± 0.4 nl·min-1·mm-1, P < 0.01; n = 10). Selective inhibition of DDAH-1 in rats with intravenous L-257 (60 mg/kg) given 2 h before and L-257 (10-5 mol/l) perfused in the proximal tubule for 5 min reduced Jv by 32 ± 4% (vehicle: 3.2 ± 0.5 vs. L-257: 2.2 ± 0.5 nl·min-1·mm-1; P < 0.01) and increased plasma ADMA by ≈50% (vehicle: 0.46 ± 0.03 vs. L-257: 0.67 ± 0.03 µmol/l, P < 0.0001) without changing plasma symmetric dimethylarginine. Compared with nontargeted control small-interference RNA, knock down of DDAH-1 in mice by 60% with targeted small-interference RNAs (siRNA) reduced Jv by 29 ± 5% (nontargeted siRNA: 2.8 ± 0.20 vs. DDAH-1 knockdown: 1.9 ± 0.31 nl·min-1·mm-1, P < 0.05). In conclusion, fluid reabsorption in the proximal tubule is reduced by tubular ADMA or by blocking its metabolism by DDAH-1. L-257 is a novel regulator of proximal tubule fluid reabsorption. [ABSTRACT FROM AUTHOR]

Nuclear factor erythyroid factor 2 (Nrf2) transcribes genes in cultured endothelial cells that reduce reactive oxygen species (ROS) and generate nitric oxide (NO) or metabolize asymmetric dimethylarginine (ADMA), which inhibits NO synthase (NOS). Therefore, we undertook a functional study to test the hypothesis that activation of Nrf2 by tertbutylhydroquinone (tBHQ) preserves microvascular endothelial function during oxidative stress. Wild-type CB57BL/6 (wt), Nrf2 wt (+/+), or knockout (-/-) mice received vehicle (Veh) or tBHQ (0.1%; activator of Nrf2) during 14-day infusions of ANG II (to induce oxidative stress) or sham. MAP was recorded by telemetry. Mesenteric resistance arterioles were studied on isometric myographs and vascular NO and ROS by fluorescence microscopy. ANG II increased the mean arterial pressure (112 ± 5 vs. 145 ± 5 mmHg; P < 0.01) and excretion of 8-isoprostane F2α (2.8 ± 0.3 vs. 3.8 ± 0.3 ng/mg creatinine; P < 0.05) at 12–14 days. However, 12 days of ANG II reduced endothelium-derived relaxation (27 ± 5 vs. 17 ± 3%; P < 0.01) and NO (0.38 ± 0.07 vs. 0.18 ± 0.03 units; P < 0.01) but increased microvascular remodeling, endothelium-derived contractions (7.5 ± 0.5 vs. 13.0 ± 1.7%; P < 0.01), superoxide (0.09 ± 0.03 vs. 0.29 ± 0.08 units; P < 0.05), and contractions to U-46,619 (87 ± 6 vs. 118 ± 3%; P < 0.05), and endothelin-1(89 ± 4 vs. 123 ± 12%; P < 0.05). tBHQ prevented all of these effects of ANG II at 12–14 days in Nrf2+/+ mice but not in Nrf2-/- mice. In conclusion, tBHQ activates Nrf2 to prevent microvascular endothelial dysfunction, remodeling, and contractility, and moderate ADMA and hypertension at 12–14 days of ANG II infusion, thereby preserving endothelial function and preventing hypertension. [ABSTRACT FROM AUTHOR]

The Na+-K+-2Cl- cotransporter (NKCC2) on the loop of Henle is the site of action of furosemide. Because outer medullary potassium channel (ROMK) inhibitors prevent reabsorption by NKCC2, we tested the hypothesis that ROMK inhibition with a novel selective ROMK inhibitor (compound C) blocks tubuloglomerular feedback (TGF) and reduces vascular resistance. Loop perfusion of either ROMK inhibitor or furosemide caused dose-dependent blunting of TGF, but the response to furosemide was 10-fold more sensitive (IC50 = 10-6 M for furosemide and IC50 = 10-5 M for compound C). During systemic infusion, both diuretics inhibited TGF, but ROMK inhibitor was 10-fold more sensitive (compound C: 63% inhibition; furosemide: 32% inhibition). Despite blockade of TGF, 1 h of constant systemic infusion of both diuretics reduced the glomerular filtration rate (GFR) and renal blood flow (RBF) by 40 -60% and increased renal vascular resistance (RVR) by 100 - 200%. Neither diuretic altered blood pressure or hematocrit. Proximal tubule hydrostatic pressures (PPT) increased transiently with both diuretics (compound C: 56% increase; furosemide: 70% increase) but returned to baseline. ROMK inhibitor caused more natriuresis (3,400 vs. 1,600% increase) and calciuresis (1,200 vs. 800% increase) but less kaliuresis (33 vs. 167% increase) than furosemide. In conclusion, blockade of ROMK or Na+-K+-2Cl- transport inhibits TGF yet increases renal vascular resistance. The renal vasoconstriction was independent of volume depletion, blood pressure, TGF, or PPT. [ABSTRACT FROM AUTHOR]

Myogenic contraction is the principal component of renal autoregulation that protects the kidney from hypertensive barotrauma. Contractions are initiated by a rise in perfusion pressure that signals a reduction in membrane potential (Em) of vascular smooth muscle cells to activate voltage-operated Ca2+ channels. Since ROS have variable effects on myogenic tone, we investigated the hypothesis that superoxide (O2·-) and H2O2 differentially impact myogenic contractions. The myogenic contractions of mouse isolated and perfused single afferent arterioles were assessed from changes in luminal diameter with increasing perfusion pressure (40-80 mmHg). O2·-, H2O2, and Em were assessed by fluorescence microscopy during incubation with paraquat to increase O2·- or with H2O2. Paraquat enhanced O2·- generation and myogenic contractions (-42 ± 4% vs. -19 ± 4%, P < 0.005) that were blocked by SOD but not by catalase and signaled via PKC. In contrast, H2O2 inhibited the effects of paraquat and reduced myogenic contractions (-10 ± 1% vs. -19 ± 2%, P < 0.005) and signaled via PKG. O2·- activated Ca2+-activated Cl- channels that reduced Em, whereas H2O2 activated Ca2+-activated and voltage-gated K+ channels that increased Em. Blockade of voltage-operated Ca2+ channels prevented the enhanced myogenic contractions with paraquat without preventing the reduction in Em. Myogenic contractions were independent of the endothelium and largely independent of nitric oxide. We conclude that O2·- and H2O2 activate different signaling pathways in vascular smooth muscle cells linked to discreet membrane channels with opposite effects on Em and voltage-operated Ca2+ channels and therefore have opposite effects on myogenic contractions. [ABSTRACT FROM AUTHOR]

Adenosine 1 receptors (A1AR) have been shown in previous experiments to play a major role in the tubuloglomerular feedback (TGF) constrictor response of afferent arterioles (AA) to increased loop of Henle flow. Overexpression studies have pointed to a critical role of vascular A1AR, but it has remained unclear whether selective deletion of A1AR from smooth muscle cells is sufficient to abolish TGF responsiveness. To address this question, we have determined TGF response magnitude in mice in which vascular A1AR deletion was achieved using the loxP recombination approach with cre recombinase being controlled by a smooth muscle actin promoter (SmCre/A1ARff). Effective vascular deletion of A1AR was affirmed by absence of vasoconstrictor responses to adenosine or cyclohexyl adenosine (CHA) in microperfused AA. Elevation of loop of Henle flow from 0 to 30 nl/min caused a 22.1 ± 3.1% reduction of stop flow pressure in control mice and of 7.2 ± 1.5% in SmCre/A1ARff mice (P < 0.001). Maintenance of residual TGF activity despite absence of A1AR-mediated responses in AA suggests participation of extravascular A1AR in TGF. Support for this notion comes from the observation that deletion of A1ARff by nestin-driven cre causes an identical TGF response reduction (7.3 ± 2.4% in NestinCre/A1ARff vs. 20.3 ± 2.7% in controls), whereas AA responsiveness was reduced but not abolished. A1AR on AA smooth muscle cells are primarily responsible for TGF activation, but A1AR on extravascular cells, perhaps mesangial cells, appear to contribute to the TGF response. [ABSTRACT FROM AUTHOR]

We tested the hypothesis that reactive oxygen species (ROS) contributed to renal hypoxia in C57BL/6 mice with 5/6 surgical reduction of renal mass (RRM). ROS can activate the mitochondrial uncoupling protein 2 (UCP-2) and increase O2 usage. However, UCP-2 can be inactivated by glutathionylation. Mice were fed normal (NS)- or high-salt (HS) diets, and HS mice received the antioxidant drug tempol or vehicle for 3 mo. Since salt intake did not affect the tubular Na+ transport per O2 consumed (TNa/QO2), further studies were confined to HS mice. RRM mice had increased excretion of 8-isoprostane F2α and H2O2, renal expression of UCP-2 and renal O2 extraction, and reduced TNa/QO2 (sham: 20 ± 2 vs. RRM: 10 ± 1 μmol/μmol; P < 0.05) and cortical PO2 (sham: 43 ± 2, RRM: 29 ± 2 mmHg; P < 0.02). Tempol normalized all these parameters while further increasing compensatory renal growth and glomerular volume. RRM mice had preserved blood pressure, glomeruli, and patchy tubulointerstitial fibrosis. The patterns of protein expression in the renal cortex suggested that RRM kidneys had increased ROS from upregulated p22phox, NOX-2, and -4 and that ROS-dependent increases in UCP-2 led to hypoxia that activated transforming growth factor-β whereas erythroid-related factor 2 (Nrf- 2), glutathione peroxidase-1, and glutathione-S-transferase mu-1 were upregulated independently of ROS. We conclude that RRM activated distinct processes: a ROS-dependent activation of UCP-2 leading to inefficient renal O2 usage and cortical hypoxia that was offset by Nrf-2-dependent glutathionylation. Thus hypoxia in RRM may be the outcome of NADPH oxidase-initiated ROS generation, leading to mitochondrial uncoupling counteracted by defense pathways coordinated by Nrf-2. [ABSTRACT FROM AUTHOR]