Angiotensin AT1 Receptor Research Articles

Vascular effects of a tripeptide fragment of novokinine in hypertensive rats: Mechanism of the hypotensive action

BackgroundActivation of angiotensin AT2 receptors (AT2R) counteracts vasoconstrictor effects of AT1R stimulation and contributes to blood pressure control. We examined effects on mean arterial pressure (MAP) and renal hemodynamics of LKP, a tripeptide fragment of novokinine, an established AT2R agonist. MethodsEffects of intravenous LKP infusion and then superimposed losartan (AT1R antagonist) on MAP, total renal (RBF, Transonic probe) and renal medullary blood flows (laser-Doppler), and on renal excretion, were examined in anesthetized (1) Wistar rats with acute norepinephrine-induced hypertension, untreated or pretreated with AT2R antagonist (PD 123319) and (2) spontaneously hypertensive rats (SHR) maintained on standard or high-sodium (HS) diet. ResultsIn Wistar rats LKP decreased MAP (−4%, p<0.01) and increased renal medullary perfusion, these effects were abolished in rats pretreated with PD 123319 in which a post-LKP increase in MAP (+6%, p<0.02) occurred. LKP did not alter MAP in SHR; in those on HS diet RBF decreased (−14%, p<0.02), the response that was reverted by losartan. Addition of losartan always decreased or tended to decrease MAP. ConclusionsLKP lowered MAP in norepinephrine-induced hypertension, probably via activation of AT2R. At reduced availability of AT2R, as in SHR, LKP appeared to bind to different receptors, possibly AT1, and induced systemic or renal vasoconstriction. Compared to the parent novokinine, a smaller LKP molecule might be easier absorbed after oral application and more useful in therapy.

Probiotics (VSL#3) prevent endothelial dysfunction in rats with portal hypertension: role of the angiotensin system.

AimsPortal hypertension characterized by generalized vasodilatation with endothelial dysfunction affecting nitric oxide (NO) and endothelium-dependent hyperpolarization (EDH) has been suggested to involve bacterial translocation and/or the angiotensin system. The possibility that ingestion of probiotics prevents endothelial dysfunction in rats following common bile duct ligation (CBDL) was evaluated.MethodsRats received either control drinking water or the probiotic VSL#3 solution (50 billion bacteria.kg body wt−1.day−1) for 7 weeks. After 3 weeks, rats underwent surgery with either resection of the common bile duct or sham surgery. The reactivity of mesenteric artery rings was assessed in organ chambers, expression of proteins by immunofluorescence and Western blot analysis, oxidative stress using dihydroethidium, and plasma pro-inflammatory cytokine levels by flow cytometry.ResultsBoth NO- and EDH-mediated relaxations to acetylcholine were reduced in the CBDL group compared to the sham group, and associated with a reduced expression of Cx37, Cx40, Cx43, IKCa and SKCa and an increased expression of endothelial NO synthase (eNOS). In aortic sections, increased expression of NADPH oxidase subunits, angiotensin converting enzyme, AT1 receptors and angiotensin II, and formation of ROS and peroxynitrite were observed. VSL#3 prevented the deleterious effect of CBDL on EDH-mediated relaxations, vascular expression of connexins, IKCa, SKCa and eNOS, oxidative stress, and the angiotensin system. VSL#3 prevented the CBDL-induced increased plasma TNF-α, IL-1α and MCP-1 levels.ConclusionsThese findings indicate that VSL#3 ingestion prevents endothelial dysfunction in the mesenteric artery of CBDL rats, and this effect is associated with an improved vascular oxidative stress most likely by reducing bacterial translocation and the local angiotensin system.

AT2 receptors targeting cardiac protection post-myocardial infarction.

The angiotensin AT2-receptor mediates tissue protective actions. Its regenerative potential has been tested in multiple disease models including models of myocardial infarction. These studies used different experimental approaches in order to detect AT2-receptor-related effects such as AT2-receptor deficiency or overexpression, treatment with an AT1-receptor blocker leading to indirect stimulation of the unopposed AT2-receptor, or studies using AT2-receptor agonists. It is a common finding in these studies that the AT2-receptor improves cardiac function in the early phase post-MI, and that this effect is preserved over periods of up to four months. Depending on the experimental protocol, the AT2R also attenuates post-MI left ventricular remodeling or protects the heart from early left ventricular thinning and rupture. In combination with AT1-receptor blockade or deficiency, post-MI cardiac hypertrophy is reduced. This article reviews studies on the role of the AT2-receptor in myocardial infarction with an emphasis on the most recent data obtained in studies using AT2-receptor agonists.

Angiotensinergic neurotransmission in the paraventricular nucleus of the hypothalamus modulates the pressor response to acute restraint stress in rats

We tested the hypothesis that the angiotensinergic neurotransmission, specifically in the paraventricular nucleus of the hypothalamus (PVN), is involved in the cardiovascular modulation during acute restraint stress (RS) in rats. The intravenous pretreatment with the angiotensin AT1 receptor antagonist losartan (5mg/kg) inhibited the pressor response to RS, but did not affect the concomitant RS-evoked tachycardiac response. Because similar effects were observed after the PVN pretreatment with CoCl2, and considering the high density of angiotensin receptors reported in the PVN, we studied the effect of the pretreatment of the PVN with either losartan or the angiotensin-converting enzyme (ACE) inhibitor lisinopril on the RS-evoked cardiovascular response. The bilateral microinjection of losartan (0.5nmol/100nL) or lisinopril (0.5nmol/100nL) into the PVN inhibited the RS-related pressor response without affecting the tachycardiac response, suggesting that the PVN angiotensinergic neurotransmission modulates the vascular component of the stress response. Finally, to exclude the possibility that centrally injected drugs could be leaking to the circulation and acting on peripheral vascular receptors, we tested the effect of the intravenous pretreatment with either losartan (0.5nmol/animal) or lisinopril (0.5nmol/animal), assuming the hypothesis of a total spread of drugs from the CNS to the peripheral circulation. When animals were pretreated with such doses of either losartan or lisinopril, the cardiovascular RS-evoked response was not affected, thus indicating that even if there were a complete leakage of the drug to the periphery, it would not affect the cardiovascular response to RS. This observation favors the idea that the effect of the intravenous injection of 5mg/kg of losartan on the RS-related cardiovascular response would be explained by an action across the blood–brain barrier, possibly in the PVN. In conclusion, the results suggest that an angiotensinergic neurotransmission in the PVN acting on AT1-receptors modulates the vascular component of the RS-evoked cardiovascular response.

GRK5 knockout mice created by TALEN‐mediated gene targeting (576.2)

G protein‐coupled receptor kinase 5 (GRK5) is one of the seven GRK family members whose primary function is to desensitize G protein‐coupled receptors (GPCRs). In recently, GRK5 deficiency has been linked to early Alzheimer disease (AD), but the mechanism by which GRK5 deficiency may accelerate to AD pathogenesis remains elusive. The GRK5 mRNA is expressed widely in brain and peripheral tissues, with highest expression evident heart, lung, and placenta. In cellular model systems, GRK5 can phosphorylate several neuronal GPCRs including ß2 adrenergic, M2‐muscarinic, secretin, angiotensin AT1, and thyroid stimulating hormone receptors. Transcription activator‐like effector nucleases (TALENs) are programmable nucleases that join FokI endonucleases with the modular DNA‐binding domain of TALEs and highly effective in inducing mutations at specific genome loci. TALEN‐mediated mutagenesis in zygotes is a potential alternative to conventional gene targeting in mice. In the presented study, we report the generation of mice with genetic knockout of the GRK gene using TALENs. We designed TALEN vectors for exon 1, 3 and 5 of mouse GRK5 gene and tested their ability to alter the each surrogate vector in 293T cells. We prepared of mRNAs for the linearized TALEN using the mMessage mMachine T7 Ultra kit. mRNAs (4ng/μl) was injected into cytoplasm of 180 one‐cell embryos. After incubation for 24 hours, the selected two‐cell embryos transferred into the oviduct of seven pseudopregnant C57BL/6 mice. We confirmed the genotype of Fo mice by sequencing and T7E1 assay.Grant Funding Source: Supported by BK 21 plus program, the Ministry of Education, Science and Technology, Korea

Maternal vasculature changes produced by experimental gestational diabetes mellitus by obesity (1051.10)

Gestational diabetes mellitus is an increasing incidence disease whose short‐term consequences in the maternal vasculature are poorly understood. The purpose of this study was to determine whether an experimental model of obesity induced gestational diabetes mellitus can modify the vasoconstrictor response of isolated arteries and if so, to explore the possible causes. Wistar rats were subjected to a high calorie diet for 7 weeks, of which the last three coincided with the period of gestation. GDM was defined by an abnormal glucose tolerance test. GDM group was compared with a control, obese, and healthy pregnant group. Dose response curves to phenylephrine and angiotensin II were done in isolated aortic rings. Also, plasma levels of angiotensin II and expression of the COX‐1, COX‐2, iNOS and AT1 receptors for angiotensin II were measured. Results showed that GDM group presented impaired glucose tolerance and an increase in the vasoconstrictor response in the presence of losartan or prazosin associated with an increase in the protein expression of AT‐1 receptor, COX‐2 and iNOS. These results provide evidence that obesity occurs DMG deleterious changes in the short‐term maternal vasculature.

Kinin Receptors in Vascular Biology and Pathology

Endogenous kinins are important vasoactive peptides whose effects are mediated by two G-Protein-coupled receptors (R), named B2R (constitutive) and B1R (inducible). They are involved in vascular homeostasis, ischemic pre- and post- conditioning, but also in cardiovascular diseases. They contribute to the therapeutic effects of angiotensin-1 converting enzyme inhibitors (ACEI) and angiotensin AT1 receptor blockers. Benefits derive primarily from vasodilatory, antiproliferative, antihypertrophic, antifibrotic, antithrombic and antioxidant properties, which are associated with the release of endothelial factors such as nitric oxide, prostacyclin and tissue plasminogen activator. Uncontrolled production of kinins or the inhibition of their metabolism may lead to unwanted pro-inflammatory side effects. Thus, B2R antagonism is salutary in angioedema, septic shock, stroke, and Chagas vasculopathy. B1R is virtually absent in healthy tissues, yet this receptor is induced by the cytokine pathway and the oxidative stress via the transcriptional nuclear factor NF-κB. The B1R may play a compensatory role for the lack of B2R, and its up-regulation during tissue damage may be a useful mechanism of host defense. Activation of both receptors may be beneficial, notably in neovascularisation, angiogenesis, heart ischemia and diabetic nephropathy. At the same time, B1R is a potent activator of inducible nitric oxide and NADPH oxidase, which are associated with vascular inflammation, increased permeability, insulin resistance, endothelial dysfunction and diabetic complications. The dual beneficial and deleterious effects of kinin receptors and, particularly B1R, raise an unsettled issue on the therapeutic value of B1R/B2R agonists versus antagonists in cardiovascular diseases. Hence, the Janus-face of kinin receptors needs to be seriously addressed in the upcoming clinical trials for each pathological setting.

The role of brain angiotensin AT2 receptors and NO in the renal sympatho‐inhibitory response to acute volume expansion in conscious rats (858.2)

This study investigated the role of AT2 receptor and NO on the renal sympatho‐inhibitory response to volume expansion (VE). Conscious rats were subjected to VE (0.25% bwt/min saline for 10min IV) following intracerebroventricular (I.C.V.) infusion of saline, angiotensin II (Ang II) or a combination of Ang II with either losartan, PD123319 (PD), an AT2 receptor antagonist or L‐NAME, an NO synthase inhibitor. I.C.V. losartan, PD or L‐NAME did not change baseline mean arterial pressure (MAP), heart rate (HR) or renal sympathetic nerve activity (RSNA). However, I.C.V. Ang II increased MAP and decreased both HR and RSNA (113±2 vs. 107±2 mmHg, 365±7 vs. 379±5 beat/min, 1.03±0.13 vs. 1.29±0.15 µV.s, respectively, all P&lt;0.05). VE decreased RSNA in all groups by some 27±2% (P&lt;0.05) at the end of the 10min. Ang II, losartan or PD had no effect on the magnitude of renal sympatho‐inhibitory response to VE. However, L‐NAME decreased the sympatho‐inhibitory response compared to saline (14±3 vs. 30±5%, P&lt;0.05). I.C.V. Ang II in combination with losartan enhanced (41±3 vs. 29±5%) but with PD decreased (15±2 vs. 28±4%, all P&lt;0.05) the renal sympatho‐inhibition compared to Ang II. The sympatho‐inhibitory response to VE following Ang II plus L‐NAME was similar to Ang II alone. These findings suggest that activation of central AT2 receptors enhances the renal sympatho‐inhibitory response to VE but this effect is not dependent on NO.Grant Funding Source: Funded by Wellcome Trust

Apelin-13 stimulates angiogenesis by promoting cross-talk between AMP-activated protein kinase and Akt signaling in myocardial microvascular endothelial cells

Currently, there is major interest in the functions of apelin-13, an endogenous ligand for the orphan G-protein coupled receptor APJ, a receptor that closely resembles the angiotensin receptor AT1. In the present study, the role of apelin-13 in angiogenesis and its mechanism as a novel angiogenic factor in myocardial microvascular endothelial cells (MMVECs) was investigated. It was revealed that apelin-13 can promote proliferation, migration and tube formation in MMVECs. In addition, apelin-13 dose dependently stimulated the phosphorylation of AMP-activated protein kinase (AMPK) and endothelial nitric oxide synthase (eNOS) at Thr-172 and Ser-1179, respectively. The treatment with the AMPK (compound C) and protein kinase Akt/protein kinase B (Akt; LY294002) inhibitor significantly suppressed the apelin‑13-induced AMPK, Akt and eNOS phosphorylation. They also inhibited the apelin13‑stimulated endothelial cell migration and tube formation. Therefore, we hypothesize that apelin-13 promotes angiogenesis through the modulation of AMPK and Akt signaling in MMVECs.

Angiotensin AT2 receptors and the baroreflex control of renal sympathetic nerve activity

Angiotensin AT2 receptors and the baroreflex control of renal sympathetic nerve activity

Autoimmune Mechanisms Activating the Angiotensin AT1 Receptor in 'Primary' Aldosteronism

The mechanisms causing excessive aldosterone production and hypertension in primary aldosteronism (PA) are complex and often incompletely recognized. Autoantibodies to the angiotensin AT1 receptor (AT1R) have been reported in some PA patients with an aldosterone-producing adenoma but not with idiopathic adrenal hyperplasia. We investigated whether these autoantibodies will activate AT1R and thereby potentially contribute to the pathophysiology of PA. AT1R autoantibody activity in sera and/or IgG purified from 13 biochemically confirmed PA patients was measured using AT1R-transfected cells, and their contractile effects were assayed using perfused rat cremaster arterioles. Aldosterone stimulation was measured in vitro using isolated human adrenal carcinoma (HAC15) adrenal cells. These data were compared with sera obtained from a group of normotensive control subjects who were expected to have negligible AT1R autoantibodies. Sera from each of the 13 PA patients significantly increased AT1R activation in AT1R-transfected cells compared with 20 control subjects, and this activity was inhibited by the selective AT1R blocker losartan. Sera and IgG purified from AT1R autoantibody-positive sera demonstrated significant vasoconstrictive effects in isolated rat cremaster arterioles and were blocked by losartan. Moreover, the AT1R autoantibody-positive IgG directly stimulated aldosterone production in the cultured adrenal cells and enhanced angiotensin-induced aldosterone production in these cells, and these effects were blocked by candesartan. These data support a probable pathophysiological role for AT1R autoantibodies in PA and thereby raise important etiological and therapeutic implications.

Functional coupling of renal K+ and Na+ handling causes high blood pressure in Na+ replete mice

A network of kinases, including WNKs, SPAK and Sgk1, is critical for the independent regulation of K+ and Na+ transport in the distal nephron. Angiotensin II is thought to act as a key hormone in orchestrating these kinases to switch from K+ secretion during hyperkalaemia to Na+ reabsorption during intravascular volume depletion, thus keeping disturbances in electrolyte and blood pressure homeostasis at a minimum. It remains unclear, however, how K+ and Na+ transport are regulated during a high Na+ intake, which is associated with suppressed angiotensin II levels and a high distal tubular Na+ load. We therefore investigated the integrated blood pressure, renal, hormonal and gene and protein expression responses to large changes of K+ intake in Na+ replete mice. Both low and high K+ intake increased blood pressure and caused Na+ retention. Low K+ intake was accompanied by an upregulation of the sodium-chloride cotransporter (NCC) and its activating kinase SPAK, and inhibition of NCC normalized blood pressure. Renal responses were unaffected by angiotensin AT1 receptor antagonism, indicating that low K+ intake activates the distal nephron by an angiotensin-independent mode of action. High K+ intake was associated with elevated plasma aldosterone concentrations and an upregulation of the epithelial sodium channel (ENaC) and its activating kinase Sgk1. Surprisingly, high K+ intake increased blood pressure even during ENaC or mineralocorticoid receptor antagonism, suggesting the contribution of aldosterone-independent mechanisms. These findings show that in a Na+ replete state, changes in K+ intake induce specific molecular and functional adaptations in the distal nephron that cause a functional coupling of renal K+ and Na+ handling, resulting in Na+ retention and high blood pressure when K+ intake is either restricted or excessively increased.

Erratum in “Antioxidant Activity-Mediated Neuroprotective Effects of an Antagonist of AT1 Receptors, Candesartan, Against Cerebral Ischemia and Edema in Rats,”

We examined the effects of post-ischemic blockade of angiotensin AT1 receptors by candesartan on cerebral infarction and formation of edema. Male Sprague–Dawley rats were divided into three groups, sham, control ischemic, and candesartan-treated (0.3 mg/kg) ischemic. Transient focal cerebral ischemia was induced by 90-min-long occlusion of the left middle cerebral artery followed by 24-h-long reperfusion. Neurological deficit score was evaluated at the end of the reperfusion period. Thereafter, the animals were randomly selected and used for three projects: (i) Measurement of the infarct volumes, (ii) investigation of ischemic brain edema formation using a wet/dry method, and (iii) assessment of the malondialdehyde (MDA) and reduced glutathione (GSH) concentrations using a HPLC technique. Induction of cerebral ischemia in the control group produced considerable infarctions in the cortex and striatum in conjunction with severely impaired motor functions. Candesartan treatment significantly reduced the infarct volumes and improved the above functions. The water content in the left (lesioned) hemisphere was considerably elevated in the control ischemic group. Candesartan treatment significantly lowered the water content in the ischemic lesioned hemisphere, retained tissue GSH level, and led to a lower MDA production. AT1 receptor blockade by candesartan treatment can noticeably decrease ischemic brain injury and attenuate edema formation, likely via increasing the antioxidant activity.

Vasopressor meets vasodepressor: The AT1–B2 receptor heterodimer

The AT1 receptor for the vasopressor angiotensin II is one of the most important drug targets for the treatment of cardiovascular diseases. Sensitization of the AT1 receptor system is a common feature contributing to the pathogenesis of many cardiovascular disorders but underlying mechanisms are not fully understood. More than a decade ago, evidence was provided for control of AT1R activation by heterodimerization with the B2 receptor for the vasodepressor peptide, bradykinin, a physiological counterpart of the vasoconstrictor angiotensin II. AT1–B2 receptor heterodimerization was shown to enhance AT1R-stimulated signaling under pathophysiological conditions such as experimental and human pregnancy hypertension. Notably, AT1R signal sensitization of patients with preeclampsia hypertension was attributed to AT1R–B2R heterodimerization. Vice versa, transgenic mice lacking the AT1–B2 receptor heterodimer due to targeted deletion of the B2R gene showed a significantly reduced AT1R-stimulated vasopressor response compared to transgenic mice with abundant AT1R–B2R heterodimerization. Biophysical methods such as BRET and FRET confirmed those data by demonstrating efficient AT1–B2 receptor heterodimerization in transfected cells and transgenic mice. Recently, a study on AT1R-specific biased agonism directed the focus to the AT1–B2 receptor heterodimer again. The β-arrestin-biased [Sar1,Ile4,Ile8]-angiotensin II promoted not only the recruitment of β-arrestin to the AT1R but also stimulated the down-regulation of the AT1R-associated B2 receptor by co-internalization. Thereby specific targeting of the AT1R–B2R heterodimer became feasible and could open the way to a new class of drugs, which specifically interfere with pathological angiotensin II-AT1 receptor system activation.

Minireview: More Than Just a Hammer: Ligand “Bias” and Pharmaceutical Discovery

Conventional orthosteric drug development programs targeting G protein-coupled receptors (GPCRs) have focused on the concepts of agonism and antagonism, in which receptor structure determines the nature of the downstream signal and ligand efficacy determines its intensity. Over the past decade, the emerging paradigms of "pluridimensional efficacy" and "functional selectivity" have revealed that GPCR signaling is not monolithic, and that ligand structure can "bias" signal output by stabilizing active receptor states in different proportions than the native ligand. Biased ligands are novel pharmacologic entities that possess the unique ability to qualitatively change GPCR signaling, in effect creating "new receptors" with distinct efficacy profiles driven by ligand structure. The promise of biased agonism lies in this ability to engender "mixed" effects not attainable using conventional agonists or antagonists, promoting therapeutically beneficial signals while antagonizing deleterious ones. Indeed, arrestin pathway-selective agonists for the type 1 parathyroid hormone and angiotensin AT1 receptors, and G protein pathway-selective agonists for the GPR109A nicotinic acid and μ-opioid receptors, have demonstrated unique, and potentially therapeutic, efficacy in cell-based assays and preclinical animal models. Conversely, activating GPCRs in "unnatural" ways may lead to downstream biological consequences that cannot be predicted from prior knowledge of the actions of the native ligand, especially in the case of ligands that selectively activate as-yet poorly characterized G protein-independent signaling networks mediated via arrestins. Although much needs to be done to realize the clinical potential of functional selectivity, biased GPCR ligands nonetheless appear to be important new additions to the pharmacologic toolbox.

Aliskiren Reduces Myocardial Ischemia–Reperfusion Injury by a Bradykinin B 2 Receptor– and Angiotensin AT 2 Receptor–Mediated Mechanism

Angiotensin-converting enzyme inhibitors and angiotensin AT1 receptor blockers reduce myocardial ischemia-reperfusion injury via bradykinin B2 receptor- and angiotensin AT2 receptor-mediated mechanisms. The renin inhibitor aliskiren increases cardiac tissue kallikrein and bradykinin levels. In the present study, we investigated the effect of aliskiren on myocardial ischemia-reperfusion injury and the roles of B2 and AT2 receptors in this effect. Female Sprague-Dawley rats were treated with aliskiren (10 mg/kg per day) and valsartan (30 mg/kg per day), alone or in combination, together with the B2 receptor antagonist icatibant (0.5 mg/kg per day) or the AT2 receptor antagonist PD123319 (30 mg/kg per day), for 4 weeks before myocardial ischemia-reperfusion injury. Aliskiren increased cardiac bradykinin levels and attenuated valsartan-induced increases in plasma angiotensin II levels. In vehicle-treated rats, myocardial infarct size (% area at risk, mean±SEM, n=7-13) was 43±3%. This was reduced to a similar extent by aliskiren, valsartan, and their combination to 24±3%, 25±3%, and 22±2%, respectively. Icatibant reversed the cardioprotective effects of aliskiren and the combination of aliskiren plus valsartan, but not valsartan alone, indicating that valsartan-induced cardioprotection was not mediated by the B2 receptor. PD123319 reversed the cardioprotective effects of aliskiren, valsartan, and the combination of aliskiren plus valsartan. Aliskiren protects the heart from myocardial ischemia-reperfusion injury via a B2 receptor- and AT2 receptor-mediated mechanism, whereas cardioprotection by valsartan is mediated via the AT2 receptor. In addition, aliskiren attenuates valsartan-induced increases in angiotensin II levels, thus preventing AT2 receptor-mediated cardioprotection by valsartan.

Participation of 5-HT and AT1Receptors within the Rostral Ventrolateral Medulla in the Maintenance of Hypertension in the Goldblatt 1 Kidney-1 Clip Model

The hypothesis that changes in neurotransmission within the rostral ventrolateral medulla (RVLM) are important to maintain the high blood pressure (BP) was tested in Goldblatt one kidney-one clip hypertension model (1K-1C). Male Wistar rats were anesthetized (urethane 1.2 g/kg, i.v.), and the effects of bilateral microinjections into the RVLM of the following drugs were measured in 1K-1C or control groups: glutamate (0.1 mol/L, 100 nL) and its antagonist kynurenic acid (0.02 mol/L, 100 nL), the angiotensin AT1 receptor antagonist candesartan (0.01 mol/L, 100 nL), and the nonselective 5-HT receptor antagonist methiothepin (0.06 mol/L, 100 nL). Experiments in 1K-1C rats were performed 6 weeks after surgery. In anesthetized rats glutamate response was larger in hypertensive than in normotensive rats (H: Δ67 ± 6.5; N: Δ43 ± 3.54 mmHg). In contrast, kynurenic acid microinjection into the RVLM did not cause any change in BP in either group. The blockade of either AT1 or 5-HT receptors within the RVLM decreased BP only in 1K-1C rats. A largest depressor response was caused by 5-HT receptor blockade. The data suggest that 5-HT and AT1 receptors act tonically to drive RVLM in 1K-1C rats, and these actions within RVLM contribute to the pathogenesis of this model of hypertension.

Natriorexigenic effect of DAMGO is decreased by blocking AT1 receptors in the central nucleus of the amygdala

μ-Opioid receptor (μ-OR) activation with agonist [D-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin (DAMGO) in the central nucleus of the amygdala (CeA) induces sodium (0.3M NaCl) intake in rats. The purpose of this study was to examine the effects of pre-injections of losartan (AT1 angiotensin receptor antagonist) into the CeA on 0.3M NaCl and water intake induced by DAMGO injected bilaterally in the same area in rats submitted to water deprivation–partial rehydration (WD–PR) and in rats treated with the diuretic furosemide (FURO) combined with a low dose of the angiotensin-converting enzyme inhibitor captopril (CAP) injected subcutaneously (FURO/CAP). Male Sprague–Dawley rats with stainless steel cannulas implanted bilaterally into the CeA were used. In WD–PR rats, bilateral injections of DAMGO (2nmol in 0.5μL) into the CeA induced 0.3M NaCl and water intake, and pre-treatment with losartan (108nmol in 0.5μL) injected into the CeA reduced 0.3M NaCl and water intake induced by DAMGO. In FURO/CAP rats, pre-treatment with losartan (108nmol in 0.5μL) injected into the CeA attenuated the increase in 0.3M NaCl and water intake induced by DAMGO (2nmol in 0.5μL) injected into the same site. The results suggest that the natriorexigenic effect of DAMGO injected into the CeA is facilitated by endogenous angiotensin II acting on AT1 receptors in the CeA, which drives rats to ingest large amounts of hypertonic NaCl.

Antihypertensive Mechanism of Lactoferrin-Derived Peptides: Angiotensin Receptor Blocking Effect

Looking for antihypertensive mechanisms beyond ACE inhibition, we assessed whether lactoferrin (LF)-derived peptides can act as receptor blockers to inhibit vasoconstriction induced by angiotensin II or endothelin-1. The lactoferricin B (LfcinB)-derived peptide LfcinB20-25 (RRWQWR), the low molecular weight LF hydrolysate (LFH < 3 kDa), and two peptides identified in LFH < 3 kDa (LIWKL and RPYL) were tested in ex vivo assays of vasoactive responses. The peptide RPYL was tested in radioligand receptor binding assays. Both LFH < 3 kDa and individual peptides inhibited angiotensin II-induced vasoconstriction. RPYL showed the highest ex vivo inhibitory effect and also inhibited binding of [(125)I]-(Sar(1),Ile(8))-angiotensin II to AT1 receptors. By contrast, neither LFH < 3 kDa nor RPYL inhibited endothelin-1 and depolarization-induced vasoconstrictions. In conclusion, LF-derived peptides selectively inhibit angiotensin II-induced vasoconstriction by blocking angiotensin AT1 receptors. Therefore, inhibition of angiotensin II-induced vasocontriction is suggested as a mechanism contributing along with ACE inhibition to the antihypertensive effect of some LF-derived peptides.

Avosentan is protective in hypertensive nephropathy at doses not causing fluid retention

Multiple studies indicate that endothelin antagonism may have a protective effect for chronic kidney disease. Despite that, clinical studies using avosentan have been halted due to adverse effects including fluid overload. Therefore, we aimed at investigating whether avosentan may have protective effects against hypertensive nephropathy at doses below those inducing fluid-retention. We used double transgenic rats (dTGR), overexpressing both the human renin and angiotensinogen gene, which develop malignant hypertension. Effects of avosentan alone or in combination with low-dose of valsartan (angiotensin AT1 receptor antagonist) on end-organ damage were studied. Avosentan induced a decrease of diuresis (18.3%) with a consequent decrease in hematocrit (8.3%) only at the highest dose investigated (100mg/kg). Treatment with the combination of avosentan and valsartan (10 and 0.1mg/kg, once daily by gavage, respectively) decreased albuminuria to a greater extent than each compound given alone (avosentan: 19.6mg/24h; valsartan: 12.9mg/24h; avosentan+valsartan: 1.7mg/24h, data are median values). Histological severity score also showed a drastic reduction of kidney damage. Furthermore, avosentan alone or in combination therapy dramatically decreased mortality compared to the 100% in untreated animals. These data support a therapeutic effect of avosentan at doses below those inducing fluid overload.