Ang II Type 1 Receptor Activation Research Articles

Vasoconstrictor and Pressor Effects of Des-Aspartate-Angiotensin I in Rat.

This study investigated the vasoactive effects of des-aspartate-angiotensin-I (DAA-I) in male Wistar rats on whole body vascular bed, isolated perfused kidneys, and aortic rings. Dose–response curves to DAA-I were compared with those to angiotensin II (Ang II). The Ang II-type-1 (AT1) receptor blocker, losartan, was used to evaluate the role of AT1 receptors in the responses to DAA-I. Studies were also conducted of the responsiveness in aortic rings after endothelium removal, nitric oxide synthase inhibition, or AT2 receptor blockade. DAA-I induced a dose-related systemic pressor response that was shifted to the right compared with Ang II. Losartan markedly attenuated the responsiveness to DAA-I. DAA-I showed a similar pattern in renal vasculature and aortic rings. In aortic rings, removal of endothelium and nitric oxide inhibition increased the sensitivity and maximal response to DAA-I and Ang II. AT2 receptor blockade did not significantly affect the responsiveness to DAA-I. According to these findings, DAA-I increases the systemic blood pressure and vascular tone in conductance and resistance vessels via AT1 receptor activation. This vasoconstrictor effect of DAA-I participates in the homeostatic control of arterial pressure, which can also contribute to the pathogenesis of hypertension. DAA-I may therefore be a potential therapeutic target in cardiovascular disease.

Angiotensin II Induces Oxidative Stress and Endothelial Dysfunction in Mouse Ophthalmic Arteries via Involvement of AT1 Receptors and NOX2.

Angiotensin II (Ang II) has been implicated in the pathophysiology of various age-dependent ocular diseases. The purpose of this study was to test the hypothesis that Ang II induces endothelial dysfunction in mouse ophthalmic arteries and to identify the underlying mechanisms. Ophthalmic arteries were exposed to Ang II in vivo and in vitro to determine vascular function by video microscopy. Moreover, the formation of reactive oxygen species (ROS) was quantified and the expression of prooxidant redox genes and proteins was determined. The endothelium-dependent artery responses were blunted after both in vivo and in vitro exposure to Ang II. The Ang II type 1 receptor (AT1R) blocker, candesartan, and the ROS scavenger, Tiron, prevented Ang II-induced endothelial dysfunction. ROS levels and NOX2 expression were increased following Ang II incubation. Remarkably, Ang II failed to induce endothelial dysfunction in ophthalmic arteries from NOX2-deficient mice. Following Ang II incubation, endothelium-dependent vasodilation was mainly mediated by cytochrome P450 oxygenase (CYP450) metabolites, while the contribution of nitric oxide synthase (NOS) and 12/15-lipoxygenase (12/15-LOX) pathways became negligible. These findings provide evidence that Ang II induces endothelial dysfunction in mouse ophthalmic arteries via AT1R activation and NOX2-dependent ROS formation. From a clinical point of view, the blockade of AT1R signaling and/or NOX2 may be helpful to retain or restore endothelial function in ocular blood vessels in certain ocular diseases.

Angiotensin-(1-7) and Mas receptor in the brain

The renin-angiotensin system (RAS) is a key regulator of blood pressure and electrolyte homeostasis. Besides its importance as regulator of the cardiovascular function, the RAS has also been associated to the modulation of higher brain functions, including cognition, memory, depression and anxiety. For many years, angiotensin II (Ang II) has been considered the major bioactive component of the RAS. However, the existence of many other biologically active RAS components has currently been recognized, with similar, opposite, or distinct effects to those exerted by Ang II. Today, it is considered that the RAS is primarily constituted by two opposite arms. The pressor arm is composed by Ang II and the Ang II type 1 (AT1) receptor (AT1R), which mediates the vasoconstrictor, proliferative, hypertensive, oxidative and pro-inflammatory effects of the RAS. The depressor arm is mainly composed by Ang-(1-7), its Mas receptor (MasR) which mediates the depressor, vasodilatory, antiproliferative, antioxidant and anti-inflammatory effects of Ang-(1-7) and the AT2 receptor (AT2R), which opposes to the effects mediated by AT1R activation. Central Ang-(1-7) is implicated in the control of the cardiovascular function, thus participating in the regulation of blood pressure. Ang-(1-7) also exerts neuroprotective actions through MasR activation by opposing to the harmful effects of the Ang II/AT1R axis. This review is focused on the expression and regulation of the Ang-(1-7)/MasR axis in the brain, its main neuroprotective effects and the evidence regarding its involvement in the pathophysiology of several diseases at cardiovascular and neurological level.

Blood Pressure, Vascular Reactivity, and SOD Expression/Activity in Mas1 Receptor Knockout Rats in the Dahl Salt‐Sensitive Genetic Background

Dahl salt‐sensitive (SS) rats are the most widely used rodent genetic model of human salt‐sensitive hypertension. Earlier studies have established the importance of impaired regulation of the renin gene in contributing to endothelial dysfunction in SS rats and the protective effect of low dose angiotensin II (ANG II) infusion in restoring endothelial function via the ANG II type1 receptor (AT1R) in the SS genetic background. Activation of the Mas1 receptor (Mas1R) for angiotensin (1‐7) [ANG (1‐7)] generally opposes ANG II‐mediated effects of AT1R activation. In Sprague‐Dawley rats fed high salt diet, binding of ANG (1‐7) to the Mas1R can activate NRF2 defenses and restore impaired endothelial function. The goal of the present study was to provide initial phenotyping of a Mas1R knockout (Mas1R KO) rat in the Dahl SS genetic background. Mas1R KO rats and wild type (WT) controls (12–15 weeks old) were maintained on a low salt (LS; 0.4% NaCl) or switched to a high salt (HS; 4% NaCl) diet for 3 days or 10 days. Arterial pressure measured via telemetry was similar in Mas1R KO rats (121±7 mmHg, n=3) and WT rats fed LS diet (124 mmHg, n=2) and increased in WT (138 mmHg, n=2) and Mas1R KO rats (132±5 mmHg, n=3) fed HS diet for 10 days. In rats fed LS diet, Mas1R KO eliminated vasodilator responses to ANG (1‐7) and AVE0991 (Mas receptor agonist) in middle cerebral arteries (MCA) and mesenteric resistance arteries (MRA) and reduced acetylcholine (ACh) reactivity in MCA. Because superoxide dismutase (SOD) enzymes contribute to antioxidant defense by catalyzing the dismutation of superoxide radicals and help to prevent diseases related to oxidative stress and endothelial dysfunction, we measured superoxide dismutase 1 (SOD1) and superoxide dismutase 2 (SOD2) activity in the liver of WT and Mas1R KO rats after 3 days of HS diet. High salt diet increased SOD1 activity (WT‐LS 1.04±0.16 U/ml vs WT‐HS 1.65±0.12 U/ml) and SOD2 activity (WT‐LS 0.76±0.03 U/ml vs WT‐HS 1.06±0.11 U/ml) in WT rats, but not in Mas1R KO rats. SOD1 and SOD2 mRNA expression (evaluated via RT‐PCR) tended to increase in WT rats fed HS diet vs. LS diet (1.3 – 1.8 fold change, p > 0.05). SOD1 mRNA expression was significantly higher in HS‐fed Mas1R KO rats vs. Mas1R KO rats fed LS diet (2.8 fold change, p = 0.01) and SOD2 mRNA tended to increase as well (1.7 fold change; p = 0.1). SOD1 and SOD2 protein expression measured by Western blot analysis was similar in WT and Mas1R KO rats fed either LS or HS diet. The present study using the Mas1 receptor knockout in the Dahl SS rat genetic background suggests that Mas1R activation by its agonists (AVE 0991) and ANG (1‐7) improves endothelial function and may provide novel therapeutic strategies for the treatment of low‐renin salt‐sensitive hypertension.Support or Funding InformationNIH #HL‐128242 and NIH #R21‐OD024781

Activation of Bone Marrow-Derived Cells Angiotensin (Ang) II Type 1 Receptor by Ang II Promotes Atherosclerotic Plaque Vulnerability.

Angiotensin (Ang) II triggers vulnerable atherosclerotic plaque development. Bone marrow (BM)-derived cells are key players in atherogenesis but whether Ang II induces plaque vulnerability directly through Ang II type 1 receptor (AT1R) activation on these cells remains to be clarified. In the present study, we investigated whether a lack of AT1R on BM-derived cells might affect Ang II-mediated vulnerable plaque development. The 2-kidney, 1-clip (2K1C) model (Ang II-dependent mouse model of advanced atherosclerosis and vulnerable plaques) was generated in ApoE−/− mice transplanted with AT1aR−/− or AT1aR+/+ BM. Plasma cholesterol as well as hepatic mRNA expression levels of genes involved in cholesterol metabolism were significantly lower in 2K1C mice transplanted with AT1aR−/− BM than in controls. Atherosclerotic lesions were significantly smaller in AT1aR−/− BM 2K1C mice (−79% in the aortic sinus and −71% in whole aorta compared to controls). Plaques from AT1aR−/− BM 2K1C mice exhibited reduced lipid core/fibrous cap and macrophage/smooth muscle cells ratios (−82% and −88%, respectively), and increased collagen content (+70%), indicating a more stable phenotype. Moreover, aortic mRNA levels of pro-inflammatory cytokines IL-12p35, IL-1β, and TNF-α were significantly reduced in AT1aR−/− BM 2K1C mice. No significant differences in either the number of circulating Ly6Chigh inflammatory monocytes and Ly6Clow resident anti-inflammatory monocyte subsets, or in mRNA levels of aortic M1 or M2 macrophage markers were observed between the two groups. No significant differences were observed in splenic mRNA levels of T cell subsets (Th1, Th2, Th17 and Treg) markers between the two groups. In conclusion, direct AT1R activation by Ang II on BM-derived cells promotes hepatic mRNA expression of cholesterol-metabolism-related genes and vascular mRNA expression of pro-inflammatory cytokines that may lead to plaque instability.

Angiotensin II facilitates GABAergic neurotransmission at postsynaptic sites in rat amygdala neurons

The central nucleus of the amygdala (CeA) is critical in the regulation of sodium appetite. Angiotensin II (Ang II) is important in the generation of sodium appetite and may function as a neurotransmitter or modulator to affect the synaptic transmission and the excitability of neurons. However, the role of Ang II in the CeA remains unclear. In this study, we determined the effects of Ang II on the excitatory and inhibitory synaptic inputs to the CeA neurons in brain slices with whole-cell patch-clamp recordings. Ang II (0.5–5 μM) significantly potentiated the amplitude of spontaneous GABAergic inhibitory postsynaptic currents (IPSCs) in a concentration-dependent manner. Ang II (2 μM) significantly increased the amplitude of miniature GABAergic inhibitory postsynaptic currents (mIPSCs) without affecting the frequency. This effect was blocked by Ang II type 1 (AT1) receptor antagonist, losartan. One mM guanosine 5′-O-(-2-thiodiphosphate) (GDP-β-s) in the pipette internal solution eliminated the facilitatory effect of Ang II on GABAergic synaptic transmission. In contrast, Ang II had no effect on the spontaneous glutamatergic excitatory postsynaptic currents (EPSCs) and did not alter the frequency and amplitude of miniature EPSCs at concentrations that facilitated IPSCs. Furthermore, Ang II decreased the firing activity of CeA neurons, and this effect was abolished by losartan and GDP-β-s. In addition, Ang II failed to inhibit CeA neurons in the presence of bicuculline. These data provide substantial new evidence that Ang II inhibits the CeA neurons by facilitation of GABAergic synaptic input efficacy through activation of postsynaptic AT1 receptors.

Activation of angiotensin II type 1 receptors and contractile activity in human sigmoid colon in vitro.

To analyse the effects of angiotensin II (Ang II) on the contractility of human sigmoid colon, and to characterize the subtype(s) of receptor(s) involved and the related action mechanism. The contractility of sigmoid colon circular muscle strips was recorded isometrically. RT-PCR and immunohistochemistry were used to reveal the eventual existence of a local renin-angiotensin system (RAS) and the distribution of Ang II receptors. Transcripts encoding for the Ang II type 1 (AT1 ) and the Ang II type 2 (AT2 ) receptor subtypes and for the angiotensin-converting enzyme in the whole-thickness muscular wall were observed. Ang II caused a concentration-dependent contractile response, which is antagonized by losartan, AT1 receptor antagonist, but not by PD123319, AT2 receptor antagonist. The joint application of losartan and PD123319 did not produce any additive effect. The contractile response to Ang II was partially reduced by tetrodotoxin, Na(+) voltage-gated neural channel blocker, and to some extent by SR48968, tachykinin NK2 receptor antagonist. However, hexamethonium, nicotinic receptor antagonist, atropine, cholinergic muscarinic receptor antagonist and SR140333, tachykinin NK1 receptor antagonist, were ineffective. Immunohistochemical analysis showed that AT1 receptors were expressed on the smooth muscle layers and myenteric plexus. Ang II positively modulates the spontaneous contractile activity of human sigmoid colon via activation of post-junctional and pre-junctional AT1 receptors, the latter located on the enteric nerves that modulate the release of tachykinins. The presence of the components of RAS in the human colon suggests that Ang II can be also locally generated to control colonic motility.

Role of Angiotensin II type 1 receptor on renal NAD(P)H oxidase, oxidative stress and inflammation in nitric oxide inhibition induced-hypertension

AimsActivation of the renin–angiotensin system (RAS), renal oxidative stress and inflammation are constantly present in experimental hypertension. Nitric oxide (NO) inhibition with Nw-nitro-L-arginine methyl ester (L-NAME) has previously been reported to produce hypertension, increased expression of Angiotensin II (Ang II) and renal dysfunction. The use of Losartan, an Ang II type 1 receptor (AT1R) antagonist has proven to be effective reducing hypertension and renal damage; however, the mechanism by which AT1R blockade reduced kidney injury and normalizes blood pressure in this experimental model is still complete unknown. The current study was designed to test the hypothesis that AT1R activation promotes renal NAD(P)H oxidase up-regulation, oxidative stress and cytokine production during L-NAME induced-hypertension. Main methodsMale Sprague–Dawley rats were distributed in three groups: L-NAME, receiving 70mg/100ml of L-NAME, L-NAME+Los, receiving 70mg/100ml of L-NAME and 40mg/kg/day of Losartan; and Controls, receiving water instead of L-NAME or L-NAME and Losartan. Key findingsAfter two weeks, L-NAME induced high blood pressure, renal overexpression of AT1R, NAD(P)H oxidase sub-units gp91, p22 and p47, increased levels of oxidative stress, interleukin-6 (IL-6) and interleukin-17 (IL-17). Also, we found increased renal accumulation of lymphocytes and macrophages. Losartan treatment abolished the renal expression of gp91, p22, p47, oxidative stress and reduced NF-κB activation and IL-6 expression. SignificanceThese findings indicate that NO induced-hypertension is associated with up-regulation of NADPH oxidase, oxidative stress production and overexpression of key inflammatory mediators. These events are associated with up-regulation of AT1R, as evidenced by their reversal with AT1R blocker treatment.

Angiotensin II Impairs Endothelial Progenitor Cell Number and Function In Vitro and In Vivo

Endothelial progenitor cells (EPCs) contribute to endothelial regeneration. Angiotensin II (Ang II) through Ang II type 1 receptor (AT(1)-R) activation plays an important role in vascular damage. The effect of Ang II on EPCs and the involved molecular mechanisms are incompletely understood. Stimulation with Ang II decreased the number of cultured human early outgrowth EPCs, which express both AT(1)-R and Ang II type 2 receptor, mediated through AT(1)-R activation and induction of oxidative stress. Ang II redox-dependently induced EPC apoptosis through increased apoptosis signal-regulating kinase 1, c-Jun N-terminal kinase, and p38 mitogen-activated protein kinase phosphorylation; decreased Bcl-2 and increased Bax expression; and activation of caspase 3 but had no effect on the low cell proliferation. In addition, Ang II impaired colony-forming and migratory capacities of early outgrowth EPCs. Ang II infusion diminished numbers and functional capacities of EPCs in wild-type (WT) but not AT(1)a-R knockout mice (AT(1)a(-/-)). Reendothelialization after focal carotid endothelial injury was decreased during Ang II infusion. Salvage of reendothelialization by intravenous application of spleen-derived progenitor cells into Ang II-treated WT mice was pronounced with AT(1)a(-/-) cells compared with WT cells, and transfusion of Ang II-pretreated WT cells into WT mice without Ang II infusion was associated with less reendothelialization. Transplantation of AT(1)a(-/-) bone marrow reduced atherosclerosis development in cholesterol-fed apolipoprotein E-deficient mice compared with transplantation of apolipoprotein E-deficient or WT bone marrow. Randomized treatment of patients with stable coronary artery disease with the AT(1)-R blocker telmisartan significantly increased the number of circulating CD34/KDR-positive EPCs. Ang II through AT(1)-R activation, oxidative stress, and redox-sensitive apoptosis signal-regulating kinase 1-dependent proapoptotic pathways impairs EPCs in vitro and in vivo, resulting in diminished vascular regeneration.

Angiotensin II induces RhoA activation through SHP2-dependent dephosphorylation of the RhoGAP p190A in vascular smooth muscle cells

Angiotensin II (ANG II) is a major regulator of blood pressure that essentially acts through activation of ANG II type 1 receptor (AT1R) of vascular smooth muscle cells (VSMC). AT1R activates numerous intracellular signaling pathways, including the small G protein RhoA known to control several VSMC functions. Nevertheless, the mechanisms leading to RhoA activation by AT1R are unknown. RhoA activation can result from activation of RhoA exchange factor and/or inhibition of Rho GTPase-activating protein (GAP). Here we hypothesize that a RhoGAP could participate to RhoA activation induced by ANG II in rat aortic VSMC. The knockdown of the RhoGAP p190A by small interfering RNA (siRNA) abolishes the activation of RhoA-Rho kinase pathway induced after 5 min of ANG II (0.1 microM) stimulation in rat aortic VSMC. We then show that AT1R activation induces p190A dephosphorylation and inactivation. In addition, expression of catalytically inactive or phosphoresistant p190A mutants increases the basal activity of RhoA-Rho kinase pathway, whereas phosphomimetic mutant inhibits early RhoA activation by ANG II. Using siRNA and mutant overexpression, we then demonstrate that the tyrosine phosphatase SHP2 is necessary for 1) maintaining p190A basally phosphorylated and activated by the tyrosine kinase c-Abl, and 2) inducing p190A dephosphorylation and RhoA activation in response to AT1R activation. Our work then defines p190A as a new mediator of RhoA activation by ANG II in VSMC.

Targeting Brain AT 1 Receptors By RNA Interference

The dipsogenic and pressor actions of angiotensin II (Ang II) in the central nervous system (CNS) have been well documented in many species and are now accepted as dogma. The major central cardiovascular effects of Ang II are elicited by a complex receptor-dependent signaling cascade initiated by the detection of circulating Ang II in regions of the brain lacking a blood brain barrier (BBB) through local production of Ang II in regions outside and inside the BBB or through the neurotransmitter actions of Ang II. It is generally accepted that most of the central neurocardiovascular actions of Ang II occur through the activation of intracellular signaling cascades that originate at the Ang II type 1 (AT1) receptor. A second Ang II receptor, termed AT2, has been implicated in some cardiovascular and behavioral responses. However, the AT2 receptor is distantly related to AT1 and can be effectively distinguished from AT1 pharmacologically. In humans, a single gene encodes the AT1 Ang II receptor, whereas in rodents a duplication resulted in 2 AT1 receptor genes, located on different chromosomes, each known to encode 1 of 2 distinct subtypes of the receptor, AT1a and AT1b. Both receptors are highly homologous, use the same signaling mechanisms, and cannot be distinguished using currently available pharmacological reagents or antisera. Despite their high similarity at the protein level, sufficient divergence is present in the sequence of the mRNA that allows them to be distinguished by RT-PCR and in situ hybridization. Through

Angiotensin II and Epidermal Growth Factor Induce Cyclooxygenase-2 Expression in Intestinal Epithelial Cells through Small GTPases Using Distinct Signaling Pathways

Colorectal carcinogenesis is a multistep process involving genetic mutations and alterations in rigorously controlled signaling pathways and gene expression that control intestinal epithelial cell proliferation, differentiation, and apoptosis. Cyclooxygenase-2 (COX-2) is aberrantly expressed in premalignant adenomatous polyps and colorectal carcinomas and is associated with increased epithelial cell proliferation, decreased apoptosis, and increased cell invasiveness. Currently, knowledge of the regulation of expression of COX-2 by endogenous cell-surface receptors is inadequate. Recently, in a non-transformed rat intestinal epithelial cell line (IEC-18), we showed induction of cell proliferation and DNA synthesis by angiotensin II (Ang II) via the endogenous Ang II type 1 receptor (Chiu, T., Santiskulvong, C., and Rozengurt, E. (2003) Am. J. Physiol. 285, G1-G11). We report that Ang II potently stimulated expression of COX-2 mRNA and protein as an immediate-early gene response through the Ang II type 1 receptor, correlating with an increase in prostaglandin I2 production. Ang II induced Cdc42 activation and filopodial formation. COX-2 expression was induced by epidermal growth factor (EGF), which activated Rac with lamellipodial formation. Inhibition of small GTPases by Clostridium difficile toxin B blocked COX-2 expression by Ang II and EGF. Inhibition of ERK activation by U0126 or PD98059 significantly decreased EGF-dependent COX-2 expression, but did not affect Ang II-dependent COX-2 expression. Conversely, inhibition of p38MAPK by SB202190 or PD169316 inhibited COX-2 expression by Ang II, but did not block COX-2 induction by EGF. Ang II caused Ca2+ mobilization. Inhibition of Ca2+ signaling by 2-aminobiphenyl borate blocked Ang II-dependent COX-2 expression. EGF did not induce Ca2+ mobilization, and 2-aminobiphenyl borate did not inhibit EGF-dependent COX-2 expression. Inhibition of COX-2 expression correlated with inhibition of prostaglandin I2 production. Luciferase promoter assays showed that Ang II-dependent transcriptional activation of the COX-2 promoter was dependent on activation of small GTPases and p38(MAPK) and on Ca2+ signaling via the cAMP-responsive element/activating transcription factor cis-acting element.

Proapoptotic effects of ANG II in human coronary artery endothelial cells: role of AT1 receptor and PKC activation.

Anoxia-reoxygenation, tumor necrosis factor-alpha (TNF-alpha), and angiotensin II (ANG II) have been shown to induce apoptosis in myocytes. However, the role of these mediators in causing apoptosis of human coronary artery endothelial cells (HCAEC) is not known. This study was designed to examine the interaction of these mediators in induction of apoptosis in HCAEC. Cultured HCAEC were exposed to anoxia-reoxygenation, TNF-alpha, and ANG II. TNF-alpha enhanced apoptosis of HCAEC (determined by DNA nick-end labeling in situ and DNA laddering) caused by anoxia-reoxygenation. ANG II increased apoptosis beyond that caused by anoxia-reoxygenation and TNF-alpha. Apoptosis caused by ANG II was reduced by losartan, a specific ANG II type 1 receptor (AT1R) blocker, whereas PD-123,177, a specific ANG II type 2 receptor blocker, under identical conditions had minimal effect. The proapoptotic effects of ANG II were associated with the activation of protein kinase C (PKC). The importance of PKC activation as a signal transduction mechanism became evident in experiments wherein treatment of HCAEC with a specific inhibitor of PKC activation decreased ANG II-mediated apoptosis. Thus AT1R activation appears to be responsible for apoptosis caused by ANG II in HCAEC, and AT1R activation-mediated apoptosis involves activation of PKC.

Modulation of constitutive nitric oxide synthase, bcl-2 and Fas expression in cultured human coronary endothelial cells exposed to anoxia-reoxygenation and angiotensin II: role of AT1 receptor activation.

Angiotensin II (Ang II) plays a critical role in the pathophysiology of myocardial ischemia-reperfusion injury. We have recently shown that reoxygenation following a period of anoxia causes apoptosis of cultured human coronary artery endothelial cells (HCAECs). Ang II further enhances apoptosis of HCAECs via Ang II type 1 receptor (AT1R) activation. Recent studies suggest an important role of constitutive nitric oxide synthase (cNOS), Fas and bcl-2 proteins in apoptosis. This study was designed to examine the modulation of cNOS, and Fas and bcl-2 expression in HCAECs during exposure to anoxia-reoxygenation and Ang II. HCAECs were exposed to anoxia-reoxygenation and Ang II. Anoxia-reoxygenation significantly decreased cNOS mRNA, protein and activity in cultured HCAECs (P < 0.05 vs. control). Anoxia-reoxygenation also caused an increase in Fas and a decrease in bcl-2 protein expression in cultured HCAECs (both P < 0.05 vs. control). The presence of Ang II (0.3 microM) further enhanced these effects of anoxia-reoxygenation on cNOS, Fas and bcl-2 expression. The effects of Ang II were significantly attenuated by the AT1R inhibitor losartan (10 microM). During exposure of HCAECs to anoxia-reoxygenation and Ang II, AT1R activation induces important changes in cNOS mRNA, protein expression and activity, as well as bcl-2 and Fas protein expression which may have a bearing on the development of apoptosis.

Modulation of K+ and Ca2+ currents in cultured neurons by an angiotensin II type 1a receptor peptide.

Angiotensin II (ANG II) inhibits delayed rectifier K+ current (IK) and stimulates total Ca2+ current (ICa) in neurons cocultured from newborn rat hypothalamus and brain stem, effects mediated via ANG II type 1 (AT1) receptors. Here, we identify potential G protein activator regions of the AT1 receptor responsible for initiating the intracellular changes that lead to alterations in these currents. Intracellular application into cultured neurons of a peptide corresponding to the third cytoplasmic loop of the AT1 receptor (AT1a/i3) mimicked the actions of ANG II on IK and ICa, whereas application of a peptide corresponding to the second cytoplasmic loop (AT1a/i2) did not alter these currents. This modulation of IK and ICa by AT1a/i3 involves intracellular messengers (G alpha q, protein kinase C, and intracellular Ca2+) that are identical to those involved in the modulation of IK and ICa following ANG II activation of AT1 receptors. These data provide functional evidence for a role of the third cytoplasmic loop of the AT1 receptor in G protein coupling and subsequent modulation of ion channel effectors.