Intrarenal aminopeptidase N inhibition augments natriuretic responses to angiotensin III in angiotensin type 1 receptor-blocked rats

The renal angiotensin angiotensin type 2 receptor has been shown to mediate natriuresis, and angiotensin III, not angiotensin II, may be the preferential angiotensin type 2 receptor activator of this response. Angiotensin III is metabolized to angiotensin IV by aminopeptidase N. The present study hypothesizes that inhibition of aminopeptidase N will augment natriuretic responses to intrarenal angiotensin III in angiotension type 1 receptor-blocked rats. Rats received systemic candesartan for 24 hours before the experiment. After a 1-hour control, cumulative renal interstitial infusion of angiotensin III at 3.5, 7, 14, and 28 nmol/kg per minute (each dose for 30 minutes) or angiotensin III combined with aminopeptidase N inhibitor PC-18 was administered into 1 kidney. The contralateral control kidney received renal interstitial infusion of vehicle. In kidneys infused with angiotensin III alone, renal sodium excretion rate increased from 0.05+/-0.01 micromol/min in stepwise fashion to 0.11+/-0.01 micromol/min at 28 nmol/kg per minute of angiotensin III (overall ANOVA F=3.68; P<0.01). In angiotensin III combined with PC-18, the renal sodium excretion rate increased from 0.05+/-0.01 to 0.32+/-0.08 mumol/min at 28 nmol/kg per minute of angiotensin III (overall ANOVA F=6.2; P<0.001). The addition of intrarenal PD-123319, an angiotensin type 2 receptor antagonist, to renal interstitial angiotensin III plus PC-18 inhibited the natriuretic response. Mean arterial blood pressure and renal sodium excretion rate from control kidneys were unchanged by angiotensin III +/- PC-18 + PD-123319. Angiotensin III plus PC-18 induced a greater natriuretic response than Ang III alone (overall ANOVA F=16.9; P=0.0001). Aminopeptidase N inhibition augmented the natriuretic response to angiotensin III, suggesting that angiotensin III is a major agonist of angiotensin type 2 receptor-induced natriuresis.     

Intrarenal angiotensin III is the predominant agonist for proximal tubule angiotensin type 2 receptors

In angiotensin type 1 receptor-blocked rats, renal interstitial (RI) administration of des-aspartyl(1)-angiotensin II (Ang III) but not angiotensin II induces natriuresis via activation of angiotensin type 2 receptors. In the present study, renal function was documented during systemic angiotensin type 1 receptor blockade with candesartan in Sprague-Dawley rats receiving unilateral RI infusion of Ang III. Ang III increased urine sodium excretion, fractional sodium, and lithium excretion. RI coinfusion of specific angiotensin type 2 receptor antagonist PD-123319 abolished Ang III-induced natriuresis. The natriuretic response observed with RI Ang III was not reproducible with RI angiotensin (1-7) alone or together with angiotensin-converting enzyme inhibition. Similarly, neither RI angiotensin II alone or in the presence of aminopeptidase A inhibitor increased urine sodium excretion. In the absence of systemic angiotensin type 1 receptor blockade, Ang III alone did not increase urine sodium excretion, but natriuresis was enabled by the coinfusion of aminopeptidase N inhibitor and subsequently blocked by PD-123319. In angiotensin type 1 receptor-blocked rats, RI administration of aminopeptidase N inhibitor alone also induced natriuresis that was abolished by PD-123319. Ang III-induced natriuresis was accompanied by increased RI cGMP levels and was abolished by inhibition of soluble guanylyl cyclase. RI and renal tissue Ang III levels increased in response to Ang III infusion and were augmented by aminopeptidase N inhibition. These data demonstrate that endogenous intrarenal Ang III but not angiotensin II or angiotensin (1-7) induces natriuresis via activation of angiotensin type 2 receptors in the proximal tubule via a cGMP-dependent mechanism and suggest aminopeptidase N inhibition as a potential therapeutic target in hypertension.     

Renal angiotensin type 2 receptors mediate natriuresis via angiotensin III in the angiotensin II type 1 receptor-blocked rat

Whereas angiotensin (Ang) II is the major effector peptide of the renin-angiotensin system, its metabolite, des-aspartyl1-Ang II (Ang III), may also have biologic activity. We investigated the effects of renal interstitial (RI) administration of candesartan (CAND), a specific Ang II type 1 receptor (AT1) blocker, with and without coinfusion of PD-123319 (PD), a specific Ang II type 2 receptor (AT2) blocker, on Na+ excretion (UNaV) in uninephrectomized rats. We also studied the effects of unilateral RI infusion of Ang II or Ang III on UNaV with and without systemic infusion of CAND with the noninfused kidney as control. In rats receiving normal Na+ intake, RI CAND increased UNaV from 0.07+/-0.08 to 0.82+/-0.17 micromol/min (P<0.01); this response was abolished by PD. During Na+ restriction, CAND increased UNaV from 0.06+/-0.02 to 0.1+/-0.02 micromol/min (P<0.05); this response also was blocked by PD. In rats with both kidneys intact, in the absence of CAND, unilateral RI infusion of Ang III did not significantly alter UNaV. However, with systemic CAND infusion, RI Ang III increased U(Na)V from 0.08+/-0.01 micromol/min to 0.18+/-0.04 micromol/min (P<0.01) at 3.5 nmol/kg per minute, and UNaV remained elevated throughout the infusion; this response was abolished by PD. However, RI infusion of Ang II did not significantly alter UNaV at any infusion rate (3.5 to 80 nmol/kg per minute) with or without systemic CAND infusion. These results suggest that intrarenal AT1 receptor blockade engenders natriuresis by activation of AT2 receptors. AT2 receptor activation via Ang III, but not via Ang II, mediates the natriuretic response in the presence of systemic AT1 receptor blockade.     

Estrogen regulates adrenal angiotensin type 1 receptors by modulating adrenal angiotensin levels

Estrogen inhibits adrenal angiotensin type 1 receptor (AT(1)R) binding sites and attenuates the adrenal responsivity to angiotensin II (Ang II). Ang II modulates AT(1)R expression. Here, we determined if estrogen-induced down-regulation of adrenal AT(1)Rs involves modulation of adrenal Ang II. Female rats were ovariectomized (OVX) and injected with 17beta-estradiol benzoate (E(2); 40 micro g/kg) or vehicle for 7 d. Adrenal Ang II was separated from other angiotensin peptides by HPLC and measured by RIA. Scatchard analysis of radioligand binding curves showed that E(2) or captopril (Cap; 0.5 g/liter water) significantly reduced adrenal AT(1)R binding (maximum binding capacity) by 22% and 19%, respectively, compared with OVX (276 +/- 2.09 fmol/mg protein). E(2) and Cap lowered adrenal Ang II levels by 39% and 21%, respectively, compared with OVX (4.10 +/- 0.44 pmol/g). E(2) caused no further reductions in adrenal AT(1)R binding or in Ang II levels in Cap-treated OVX rats. High-dose Ang II infusion (1000 ng/kg.min) increased adrenal Ang II levels by 71% and lowered AT(1)R binding by 18%. Under these infusion conditions, E(2) did not reduce adrenal Ang II or AT(1)R binding. No differences in AT(1)R affinity (dissociation constant) were observed among groups. These data suggest that E(2) regulates the number of adrenal AT(1)R binding sites indirectly by modulating adrenal Ang II.     

Brain and peripheral angiotensin II type 1 receptors mediate renal vasoconstrictor and blood pressure responses to angiotensin IV in the rat

OBJECTIVES: Angiotensin (Ang) IV was reported to increase renal cortical blood flow (CBF) via putative angiotensin IV receptor (AT4) stimulation but reduce total renal blood flow (RBF) via angiotensin II type 1 (AT1) receptors. We investigated the effect of Ang IV on simultaneously measured mean arterial pressure (MAP), RBF, and CBF. The possible involvement of AT1 or AT4 receptors, the possible natriuretic effect, and responses to central administration were also explored. METHODS AND RESULTS: Intravenous injections of Ang IV dose dependently increased MAP and decreased CBF and RBF; these effects were abolished by AT1 receptor blockade. These reductions in CBF and RBF highly correlated as did renal vascular responses to Ang II and fenoldopam. Ang IV did not induce renal vasodilation even following AT1 receptor blockade. Intrarenal Ang IV infusion reduced CBF and RBF but had no natriuretic effect. Central Ang IV administration induced an AT1-mediated immediate increase in MAP and renal vascular resistance and a secondary increase in RBF. AT4 selective ligands, LVV-hemorphin-7 and AT4-16 (intravenous, intrarenal or intracerebroventricular), had no effects on MAP, RBF or urinary sodium excretion. Additional in-vitro experiments indicated that the majority of the Ang IV-sensitive aminopeptidase activity in kidney membranes is attributed to aminopeptidase-N. CONCLUSION: Insulin-regulated aminopeptidase (IRAP)/AT4 receptors are involved in neither the regulation of RBF or CBF nor in the handling of renal sodium. Ang IV increases MAP and induces renal vasoconstriction via stimulation of brain and peripheral AT1 receptors and may be involved in the regulation of renal blood flow and blood pressure.     

Angiotensin III stimulates aldosterone secretion from adrenal gland partially via angiotensin II type 2 receptor but not angiotensin II type 1 receptor

Angiotensin II (Ang II) and Ang III stimulate aldosterone secretion by adrenal glomerulosa, but the angiotensin receptor subtypes involved and the effects of Ang IV and Ang (1-7) are not clear. In vitro, different angiotensins were added to rat adrenal glomerulosa, and aldosterone concentration in the medium was measured. Ang II-induced aldosterone release was blocked (30.3 ± 7.1%) by an Ang II type 2 receptor (AT2R) antagonist, PD123319. Candesartan, an Ang II type 1 receptor (AT1R) antagonist, also blocked Ang II-induced aldosterone release (42.9 ± 4.8%). Coadministration of candesartan and PD123319 almost abolished the Ang II-induced aldosterone release. A selective AT2R agonist, CGP42112, was used to confirm the effects of AT2R. CGP42112 increased aldosterone secretion, which was almost completely inhibited by PD123319. In addition to Ang II, Ang III also induced aldosterone release, which was not blocked by candesartan. However, PD123319 blocked 22.4 ± 10.5% of the Ang III-induced aldosterone secretion. Ang IV and Ang (1-7) did not induce adrenal aldosterone secretion. In vivo, both Ang II and Ang III infusion increased plasma aldosterone concentration, but only Ang II elevated blood pressure. Ang IV and Ang (1-7) infusion did not affect blood pressure or aldosterone concentration. In conclusion, this report showed for the first time that AT2R partially mediates Ang III-induced aldosterone release, but not AT1R. Also, over 60% of Ang III-induced aldosterone release may be independent of both AT1R and AT2R. Ang III and AT2R signaling may have a role in the pathophysiology of aldosterone breakthrough.     

Sex differences in renal medullary endothelin receptor function in angiotensin II hypertensive rats

We hypothesized that angiotensin (Ang) II hypertensive rats have impaired natriuresis after renal medullary endothelin (ET) B receptor stimulation that would be more evident in male versus female rats. Acute intramedullary infusion of the ET(B) agonist sarafotoxin 6c in normotensive male rats increased sodium excretion from 0.51±0.11 μmol/min during baseline to 1.64±0.19 μmol/min (P<0.05) after S6c. After 2 weeks of Ang II infusion (260 ng/kg per minute SC), male rats had an attenuated natriuretic response to S6c of 0.62±0.16 μmol/min during baseline versus 0.95±0.07 μmol/min after S6c. In contrast, ET(B)-dependent natriuresis was similar in female hypertensive rats (0.48±0.07 versus 1.5±0.18 μmol/min; P<0.05) compared with normotensive controls (1.05±0.07 versus 2.14±0.24 μmol/min; P<0.05). Because ET(A) receptors also mediate natriuresis in normotensive female rats, we examined ET(A) receptor function in female Ang II hypertensive rats. Intramedullary infusion of ET-1 increased sodium excretion in both hypertensive and normotensive female rats, which was partially blocked by the ET(A) antagonist BQ-123. Maximum ET(B) receptor binding in inner medullary membrane preparations was comparable between vehicle and Ang II hypertensive females; however, maximum ET(B) binding was significantly lower in male hypertensive rats (1952±251 versus 985±176 fmol/mg; P<0.05). These results indicate that renal ET(B) function is impaired in male Ang II hypertension attributed, at least in part, to a reduced number of ET(B) binding sites. Furthermore, renal ET receptor function is preserved in female rats during chronic Ang II infusion, suggesting that renal ET receptor function could serve to limit hypertension in females compared with males.     

Mechanisms of dopamine D(1) and angiotensin type 2 receptor interaction in natriuresis

Renal dopamine D(1)-like receptors (D(1)Rs) and angiotensin type 2 receptors (AT(2)Rs) are important natriuretic receptors counterbalancing angiotensin type 1 receptor-mediated tubular sodium reabsorption. Here we explore the mechanisms of D(1)R and AT(2)R interactions in natriuresis. In uninephrectomized, sodium-loaded Sprague-Dawley rats, direct renal interstitial infusion of the highly selective D(1)R agonist fenoldopam induced a natriuretic response that was abolished by the AT(2)R-specific antagonist PD-123319 or by microtubule polymerization inhibitor nocodazole but not by actin polymerization inhibitor cytochalasin D. By confocal microscopy and immunoelectron microscopy, fenoldopam translocated AT(2)Rs from intracellular sites to the apical plasma membranes of renal proximal tubule cells, and this translocation was abolished by nocodazole. Because D(1)R activation induces natriuresis via an adenylyl cyclase/cAMP signaling pathway, we explored whether this pathway is responsible for AT(2)R recruitment and AT(2)R-mediated natriuresis. Renal interstitial coinfusion of the adenylyl cyclase activator forskolin and 3-isobutly-1-methylxanthine induced natriuresis that was abolished either by PD-123319 or nocodazole but was unaffected by specific the D(1)R antagonist SCH-23390. Coadministration of forskolin and 3-isobutly-1-methylxanthine also translocated AT(2)Rs to the apical plasma membranes of renal proximal tubule cells; this translocation was abolished by nocodazole but was unaffected by SCH-23390. The results demonstrate that D(1)R-induced natriuresis requires AT(2)R recruitment to the apical plasma membranes of renal proximal tubule cells in a microtubule-dependent manner involving an adenylyl cyclase/cAMP signaling pathway. These studies provide novel insights regarding the mechanisms whereby renal D(1)Rs and AT(2)Rs act in concert to promote sodium excretion in vivo.     

Role of the angiotensin AT(1) receptor in rat aortic and cardiac PAI-1 gene expression

Although the renin-angiotensin system has been implicated in increasing plasminogen activator inhibitor-1 (PAI-1) expression, the role of the angiotensin type 1 (AT(1)) receptor is controversial. This report examines the effects of angiotensin peptides, angiotensin-converting enzyme inhibition, and AT(1) antagonism on rat aortic and cardiac PAI-1 gene expression. In vitro, angiotensin (Ang) I, Ang II, and angiotensin Arg(2)-Phe(8) (Ang III) were potent agonists of PAI-1 mRNA expression in rat aortic smooth muscle cells (RASMCs), and stimulation of PAI-1 by these peptides was blocked by the AT(1) antagonist candesartan. Angiotensin Val(3)-Phe(8) (Ang IV) and angiotensin Asp(1)-Pro(7) (Ang [1-7]) did not affect PAI-1 expression in RASMCs. In neonatal rat cardiomyocytes, Ang II increased PAI-1 mRNA expression by 4-fold (P<0.01), and this response was completely blocked by AT(1) receptor antagonism. Continuous intrajugular infusion of Ang II into Sprague-Dawley rats for 3 hours increased aortic and cardiac PAI-1 mRNA expression by 17- and 9 fold, respectively, and these Ang II responses were completely blocked by coinfusion with candesartan. Aortic and cardiac PAI-1 expressions were compared in spontaneously hypertensive rats and Wistar-Kyoto rats. PAI-1 expression in the aorta and heart from spontaneously hypertensive rats was 5.8-fold and 2-fold higher, respectively, than in control Wistar-Kyoto rats (P<0.05). Candesartan treatment for 1 week reduced aortic and cardiac PAI-1 expression in spontaneously hypertensive rats by 94% and 72%, respectively (P<0.05), but did not affect vascular PAI-1 levels in Wistar-Kyoto rats. These results demonstrate a role for the AT(1) receptor in mediating the effects of Ang II on aortic and cardiac PAI-1 gene expression.     

Proximal tubular fluid angiotensin II levels in angiotensin II-induced hypertensive rats

BACKGROUND: It has been shown that infusions of low-dose angiotensin II (Ang II) for 2 weeks lead to impaired pressure natriuresis and autoregulatory capability. Although intrarenal renin content and renin mRNA levels are markedly reduced, whole-kidney Ang II content has been shown to be increased. However, the intrarenal distribution of the increased intrarenal Ang II has not been established. OBJECTIVE: To determine the concentrations of Ang II in the proximal tubule fluid achieved in hypertensive rats (n = 16) infused with Ang II, previously prepared by infusion with Ang II at 60 ng/min via osmotic minipump for 13 days. METHODS: Rats were anesthetized with pentobarbital sodium and prepared for micropuncture, and then several free-flow proximal tubular fluid collections were obtained and pooled for each rat. At the end of each experiment, a blood sample was collected and the micropunctured kidney was excised and homogenized in chilled methanol. All samples were extracted immediately after collection and stored at 20 degrees C until the day of Ang II radioimmunoassay. RESULTS: Mean arterial blood pressure averaged 179 +/- 3 mmHg, renal plasma flow was 1.89 +/- 0.15 ml/min per g, and glomerular filtration rate averaged 0.58 +/- 0.04 ml/min per g. The Ang II concentration in proximal tubular fluid averaged 4.5 +/- 1.1 pmol/ml, a value substantially greater than the Ang II concentrations in plasma (0.17 +/- 0.03 pmol/ml), urine (0.06 +/- 0.01 pmol/ml), or total kidney tissue (0.40 +/- 0.10 pmol/g). Plasma renin activity (1.0 +/- 0.21 ng Ang I/ml per h) was markedly suppressed, as observed previously.CONCLUSIONS These findings indicate that Ang II concentrations in proximal tubular fluid collected from kidneys of anesthetized hypertensive rats infused with Ang II are in the nanomolar range, similar to those observed in normotensive rats. The inappropriate maintenance of nanomolar concentrations of Ang II in proximal tubular fluid of Ang II-infused hypertensive rats, even at markedly increased arterial pressures, may contribute to the impaired pressure natriuresis capability previously reported and, thereby, to the development and maintenance of hypertension in this model.     

Role of angiotensin receptor subtypes in mesenteric vascular proliferation and hypertrophy.

The aim of this study was to explore the regulation of angiotensin receptors after chronic infusion with angiotensin II (Ang II) and to clarify the relative roles of the angiotensin type 1 (AT(1)) and type 2 (AT(2)) receptors in the mediation of Ang II-induced mesenteric vascular hypertrophy. In male Sprague-Dawley rats, Ang II infusion at a dose of 58.3 ng/min by subcutaneous osmotic minipumps for 14 days led to increased mesenteric weight and wall:lumen ratio of the vessels and proliferation of smooth muscle cells. These vascular changes were attenuated by either valsartan, an AT(1) receptor antagonist, at a dose of 30 mg. kg(-1). d(-1) by gavage, or PD123319, an AT(2) receptor antagonist, at a dose of 830 ng/min by intraperitoneally implanted osmotic minipumps. Ang II infusion was associated with hypertension, which was prevented by valsartan, but not PD123319. (125)I-Sar(1), Ile(8) Ang II binding to mesenteric vasculature was increased after Ang II infusion. Valsartan treatment was associated with reduced Ang II binding to both receptor subtypes, whereas PD123319 was associated with reduced Ang II binding to only the AT(2) receptor subtype. These findings suggest that the trophic and proliferative effects of Ang II on the mesenteric vasculature are mediated by both AT(1) and AT(2) receptors.     

Renal angiotensin II type-2 receptors are upregulated and mediate the candesartan-induced natriuresis/diuresis in obese Zucker rats

Recently, there has been a growing interest in studying the role of angiotensin II type-2 (AT(2)) receptor in renal/cardiovascular function in pathological conditions. The present study was designed to determine the functional role of the AT(2) receptors on natriuresis/diuresis and compare the level of the tubular AT(2) receptor expression in obese and lean Zucker rats (12 weeks old). Under anesthesia, candesartan (angiotensin II type 1 [AT(1)]-specific antagonist; 100 microg/kg bolus) produced natriuresis/diuresis to a greater degree in obese than in lean rats. The specific AT(2) antagonist PD123319 (50 microg/kg per minute) after candesartan administration abolished the natriuretic/diuretic effects of candesartan in obese rats but not in lean rats. Infusion of AT(2) receptor agonist, CGP-42112A (1 microg/kg per minute), produced greater increase in sodium and urine excretion over basal in obese than in lean rats. The presence of the AT(2) receptor expression in the brush-border and basolateral membranes was confirmed by Western blotting using specific antibody and antigen-blocking peptide. Densitometric analysis of the bands revealed approximately 1.5- to 2.0-fold increase in the AT(2) receptor proteins in both membranes of obese compared with lean rats. Our results suggest upregulation of the AT(2) receptors, which play a role in mediating the natriuretic/diuretic effects of AT(1) receptor blockers in obese Zucker rats. We speculate that AT(2) receptors, by promoting sodium excretion, may protect obese Zucker rats against blood pressure increase associated with sodium and water retention.     

Role of angiotensin type-1 and angiotensin type-2 receptors in the expression of vascular integrins in angiotensin II-infused rats

Angiotensin II plays an important role in vascular remodeling through effects that involve, in part, interactions of vascular smooth muscle cells with extracellular matrix via integrins, which belong to a family of transmembrane receptors. We hypothesized that angiotensin (Ang) II regulates expression of vascular integrins and their ligands in experimental hypertension. Rats were infused subcutaneously with Ang II and received angiotensin type-1 (AT1) receptor blocker losartan, the AT1/angiotensin type-2 (AT2) [Sar1-Ile8]-Ang II, or the vasodilator hydralazine for 7 days. Osteopontin and integrin subunit expression were evaluated immunohistochemically. Ang II enhanced vascular alpha8, beta1, beta3 integrins and osteopontin expression, which were significantly reduced by losartan, [Sar1-Ile8]-Ang II, and hydralazine. Although Ang II increased vascular alpha5 subunit expression, this was additionally increased by losartan. Losartan was the only treatment that induced alpha1 subunit expression. These results demonstrate that AT1 and AT2 receptors have countervailing effects on vascular integrin subunit expression that may influence their effects on vascular remodeling and extracellular matrix composition.     

Evidence for renal vascular remodeling in angiotensin II-induced hypertension

OBJECTIVES: To determine whether 'slow pressor' hypertension from systemic angiotensin (Ang II) infusion was associated with renal vascular structural remodeling of the renal resistance vessels and glomerulus. METHODS: Ang II (4.5-10 ng/kg per min) or vehicle was infused for 10 days. Renal resistance vascular lumen changes were assessed at 10 days as changes in renal pressure flow and pressure-glomerular filtration rate (GFR) and pressure-Na+ excretion in maximally dilated, isotonically perfused kidneys. RESULTS: Low-dose, initially subpressor Ang II infusion for 10 days increased conscious arterial pressure by 27 mmHg compared to vehicle-infused rats (140 +/- 7 and 113 +/- 2 mmHg, respectively). There was no change in the pressure-flow relationship but the slope of the pressure-GFR relationship was reduced in the rats treated with Ang II. These changes are consistent with equal and opposite pre-and post-glomerular effects (i.e., increased pre-glomerular vessel resistance and reduced post-glomerular vessel resistance) and reduced glomerular ultrafiltration coefficient. There was also a significant reduction in pressure-dependent Na+ excretion. CONCLUSIONS: Slow pressor Ang II-induced hypertension was associated with apparent pro-hypertensive changes in the kidney involving pre/post-glomerular vessel remodeling as indicated by an apparent reduction in pre-glomerular lumen dimensions, a reduced glomerular filtration capacity and a reduction in the pressure natriuresis relationship.     

Loss of biphasic effect on Na/K-ATPase activity by angiotensin II involves defective angiotensin type 1 receptor-nitric oxide signaling

Oxidative stress causes changes in angiotensin (Ang) type 1 receptor (AT1R) function, which contributes to hypertension. Ang II affects blood pressure via maintenance of sodium homeostasis by regulating renal Na(+) absorption through its effects on Na/K-ATPase (NKA). At low concentrations, Ang II stimulates NKA; higher concentrations inhibit the enzyme. We examined the effect of oxidative stress on renal AT1R function involved in biphasic regulation of NKA. Male Sprague-Dawley rats received tap water (control) and 30 mmol/L of L-buthionine sulfoximine (BSO), an oxidant, with and without 1 mmol/L of Tempol (antioxidant) for 2 weeks. BSO-treated rats exhibited increased oxidative stress, AT1R upregulation, and hypertension. In proximal tubules from control rats, Ang II exerted a biphasic effect on NKA activity, causing stimulation of the enzyme at picomolar and inhibition at micromolar concentrations. However, in BSO-treated rats, Ang II caused stimulation of NKA at both of the concentrations. The effect of Ang II was abolished by the AT1R antagonist candesartan and the mitogen-activated protein kinase inhibitor UO126, whereas the Ang type 2 receptor antagonist PD-123319 and NO synthase inhibitor N(G)-nitro-L-arginine methyl ester had no effect. The inhibitory effect of Ang II was sensitive to candesartan and N(G)-nitro-L-arginine methyl ester, whereas PD-123319 and UO126 had no effect. In BSO-treated rats, Ang II showed exaggerated stimulation of NKA, mitogen-activated protein kinase, proline-rich-tyrosine kinase 2, and NADPH oxidase but failed to activate NO signaling. Tempol reduced oxidative stress, normalized AT1R signaling, unmasked the biphasic effect on NKA, and reduced blood pressure in BSO-treated rats. In conclusion, oxidative stress-mediated AT1R upregulation caused a loss of NKA biphasic response and hypertension. Tempol normalized AT1R signaling and blood pressure.     

Sex differences in the pressor and tubuloglomerular feedback response to angiotensin II

Awareness of sex differences in the pathology of cardiovascular disease is increasing. Previously, we have shown a role for the angiotensin type 2 receptor (AT(2)R) in the sex differences in the arterial pressure response to Ang II. Tubuloglomerular feedback (TGF) contributes in setting pressure-natriuresis properties, and its responsiveness is closely coupled to renal Ang II levels. We hypothesize that, in females, the attenuated pressor response to Ang II is mediated via an enhanced AT(2)R mechanism that, in part, offsets Ang II-induced sensitization of the TGF mechanism. Mean arterial pressure was measured via telemetry in male and female wild-type (WT) and AT(2)R knockout (AT(2)R-KO) mice receiving Ang II (600 ng/kg per minute SC). Basal 24-hour mean arterial pressure did not differ among the 4 groups. After 10 days of Ang II infusion, mean arterial pressure increased in the male WT (28±6 mm Hg), male AT(2)R-KO (26±2 mm Hg), and female AT(2)R-KO (26±4 mm Hg) mice, however, the response was attenuated in female WT mice (12±4 mm Hg; P between sex and genotype=0.016). TGF characteristics were determined before and during acute subpressor Ang II infusion (100 ng/kg per minute IV). Basal TGF responses did not differ between groups. The expected increase in maximal change in stop-flow pressure and enhancement of TGF sensitivity in response to Ang II was observed in the male WT, male AT(2)R-KO, and female AT(2)R-KO but not in the female WT mice (P between sex and genotype <0.05; both). In conclusion, these data indicate that an enhanced AT(2)R-mediated pathway counterbalances the hypertensive effects of Ang II and attenuates the Ang II-dependent resetting of TGF activity in females. Thus, the enhancement of the AT(2)R may, in part, underlie the protection that premenopausal women demonstrate against cardiovascular disease.     

Angiotensinergic stimulation of vascular endothelium in mice causes hypotension, bradycardia, and attenuated angiotensin response

It is not clear whether endothelial cell (EC) activation by the hormone angiotensin II (Ang II) modulates contraction of vascular smooth muscle cells (VSMCs) in the vasculature and whether impairment of this regulation in vivo contributes to hypertension. Delineation of the actions of Ang II through the type 1 receptor (AT1R) on ECs in the blood vessels has been a challenging problem because of the predominance of the AT1R functions in VSMCs that lie underneath the endothelium. We have obviated this limitation by generating transgenic (TG) mice engineered to target expression of the constitutively active N111G mutant AT1R only in ECs. In these TG mice, the enhanced angiotensinergic signal in ECs without infusion of Ang II resulted in hypotension and bradycardia. The pressor response to acute infusion of Ang II was significantly reduced. Increased expression of endothelial nitric oxide synthase and production of hypotensive mediators, nitric oxide and cyclic guanosine monophosphate, cause these phenotypes. Hypotension and bradycardia observed in the TG mice could be rescued by treatment with an AT1R-selective antagonist. Our results imply that the Ang II action by means of EC-AT1R is antagonistic to vasoconstriction in general, and it may moderate the magnitude of functional response to Ang II in VSMCs. This control mechanism in vivo most likely is a determinant of altered hemodynamic regulation involved in endothelial dysfunction in hypertensive cardiovascular disease.     

Effect of pregnancy and diabetes on vascular receptors for angiotensin II

Both gestation and diabetes mellitus (DM) are conditions associated with an increased participation of renin-angiotensin system (RAS) as well as with changes in the vascular response to angiotensin II (Ang II). We sought to establish the impact of gestational diabetes mellitus (GDM) on Ang II- induced vasoconstriction and its correlation with the expression of angiotensin II receptors (AT(1)R, AT(2) R). Female Wistar rats, virgin, or on day 3 after mating, received randomly streptozotocin (STZ) 60 mg/kgor saline ip.Streptozotocin-treated animals developed hyperglycemia (25.6 ± 1.42 mM). Ang II-induced vasoconstriction was evaluated on isolated aortas with (e+) and without (e-) endothelium and the protein expression of AT(1)R and AT(2)R was measured by western blot. Gestational diabetes mellitus significantly increased vasoconstriction with respect to all the other groups. Changes were observed only on e+ vessels. Interestingly, GDM moved AT(1)R: AT(2)R balance towards AT(1) R, while both pregnancy and diabetes towards AT(2)R expression. In conclusion, GDM increases the possibility of an hypertensive complication by an increased AT(1)R expression.     

Reversal of low dose angiotension hypertension by angiotensin receptor antagonists

During acute angiotension II (Ang II) infusion (200 ng/kg/min i.v.) into anesthetized rats, mean arterial pressure rose from 124 +/- 1 to 154 +/- 2 mm Hg. The peptidic Ang II antagonist saralasin lowered arterial pressure in a dose-dependent manner. The maximal decrease in pressure was similar to that observed after the Ang II infusion was discontinued. The nonpeptide Ang II antagonist, 4'-[( 2-butyl-4-chloro-5-(hydroxymethyl)-1H-imidazole-1-yl] methyl) [1,1'-biphenyl] -2-carboxylic acid (SC-48742), lowered acutely elevated arterial pressure to a level similar to that on discontinuation of the angiotensin infusion. Chronic (8 days) infusion of Ang II (20 ng/kg/min i.v.) increased mean arterial pressure from 116 +/- 3 to 164 +/- 7 mm Hg, which then decreased to 121 +/- 6 mm Hg on termination of the infusion. Saralasin (10 micrograms/kg/min, a maximally effective dose during acute angiotensin infusion) decreased mean arterial pressure from 168 +/- 7 to 141 +/- 3 mm Hg, a pressure significantly higher (p less than 0.05) than the pressure observed after the angiotensin infusion was discontinued. SC-48742 decreased mean arterial pressure from 167 +/- 7 to 127 +/- 3 mm Hg, a pressure not statistically different from the minimum pressure observed after the angiotensin infusion was terminated. The mechanism of blood pressure elevation during acute high dose or chronic low dose Ang II infusion is different, the latter having a significant neural component as measured by the response to trimethaphan. The peptidic antagonist saralasin was fully effective in lowering acute angiotensin hypertension but only partially effective during chronic hypertension.(ABSTRACT TRUNCATED AT 250 WORDS)