AT1A mRNA Research Articles

Resveratrol Suppresses Cardiac Renin Angiotensin System in the Late Phase of Left Ventricular Hypertrophy

Background and objectives: Resveratrol(3,5,4′-trihydroxy-trans-stilbene) is a natural polyphenole phytoalexin which exerts potential cardioprotective effects, but the cellular and molecular mechanisms responsible for these effects are still unknown. Cardiac renin angiotensin system (RAS) over-activation plays an important role in pathogenesis of left ventricular hypertrophy (LVH) progression. The aim of the study was to investigate the effects of resveratrol on the main components of RAS during early and late phase of myocardial hypertrophy. Methods: To consider the early and late phase of LVH, the rats were studied two and sixteen weeks after abdominal aorta banding without treatment (H2w and H16w groups, respectively) or with resveratrol (R) treatment. Intact animals served as control (Ctl). Arterial blood pressure was recorded by carotid cannulation. Angiotensin II (Ang II) level was measured using ELISA kit. Gene expression was evaluated by Real time RT-PCR technique. Cardiomyocyte size and fibrosis were assessed using haematoxylin/eosin and Masson trichrome staining, respectively Results: Results of this study showed that in H2w group AT1a mRNA level was increased significantly (pConclusion: Progression of LVH is accompanied by dynamic changes in RAS components expression in myocardial tissue. Resveratrol protects the heart against pressure overload-induced hypertrophy in part via RAS suppression.

Interaction between angiotensin II and glucose sensing at the subfornical organ.

The subfornical organ (SFO) lacks the normal blood-brain barrier and senses the concentrations of many different circulating signals, including glucose and angiotensin II (ANG II). ANG II has recently been implicated in the control of food intake and body weight gain. The present study assessed whether single SFO neurones sense changes in glucose and ANG II, and also whether changes in glucose concentration alter the responsiveness of these neurones to ANG II. SFO neurones dissociated from male Sprague-Dawley rats (100-175g) were used. We first examined whether glucose concentration modulates AT1 receptor expression. Similar AT1a mRNA expression levels were found at glucose concentrations of 1, 5 and 10mmolL-1 in dissociated SFO neurones. Glucose responsiveness of SFO neurones was assessed using perforated current-clamp recordings and switching between 5 and 10mmolL-1 glucose artificial cerebrospinal fluid to classify single neurones as nonresponsive (nGS), glucose-excited (GE) or glucose-inhibited (GI). In total, 26.7% of the SFO neurones were GI (n=24 of 90), 21.1% were GE (n=19 of 90) and 52.2% were nGS (n=47 of 90). Once classified, the effects of 10nmolL-1 ANG II on the excitability of these neurones were tested, with 52% of GE (n=10 of 19), 71% of GI (n=17 of 24) and 43% of nGS (n=20 of 47) neurones being ANG II sensitive. Finally, we tested whether acute changes in glucose concentration modified the response to ANG II and showed that some neurones (4/17) only respond to ANG II at 10mmolL-1 glucose. Our data demonstrate that the same SFO neurone can sense glucose and ANG II and that acute changes in glucose concentration may change ANG II responsiveness.

Mechanisms of Cross‐Talk between Leptin and Angiotensin within the Arcuate Nucleus for the Control of Metabolic Rate

Recent studies from our laboratory have demonstrated that within the arcuate nucleus of the hypothalamus (ARC), angiotensin II (ANG) type 1A receptors (AT1A) are expressed by the subset of neurons which express both the leptin receptor (LepR) and agouti‐related peptide (AgRP). AT1A expression in this subpopulation of AgRP neurons is critically involved in the integrative control of thermogenic adipose sympathetic nerve activity (SNA) and thereby resting metabolic rate (RMR) by various stimuli including leptin. It remains unclear, however, how LepR signaling results in AT1A activation within AgRP neurons. We hypothesize that activation of LepR (i) increases de novo synthesis of AGT and thus autocrine and/or paracrine ANG signaling within the ARC, and (ii) increases AT1A expression and trafficking to the cell membrane, increasing AgRP neuron sensitivity to ANG. Consistent with our first hypothesis, AGT is co‐localized within the ARC of C57BL/6J mice with cells expressing AgRP and LepR, as determined by fluorescent in situ hybridization and confocal microscopy. Studies of an immortalized mouse hypothalamic cell line (N43/5 cells, which exhibit a gene signature typical of AgRP cells) demonstrate that AGT mRNA is increased in response to leptin treatment. Selective genetic disruption of the AGT gene in LepR or AgRP neurons (AGTLepR‐KO and AGTAgRP‐KO mice) has no effect on energy flux under baseline chow‐fed conditions, though ongoing studies are focused on examining metabolic responses of these animals to high fat diet and leptin. Consistent with our second hypothesis, leptin application to N43/5 cells results in increased AT1A mRNA, and leptin injection in wildtype C57BL/6J mice increases AT1A mRNA in the ARC. Ongoing studies utilizing the new HiBiT‐tagging system (Promega) support cell surface localization of AT1A in the N43/5 cell line, which is increased by leptin treatment and reduced by ANG application. Ongoing studies are aimed at (i) examining the roles of pSTAT3, PI3K and ERK in the stimulation of AGT and AT1A expression by leptin, (ii) further characterizing AGTLepR‐KO and AGTAgRP‐KO mice by examining SNA and RMR in response to high fat diet and leptin, and (iii) using the HiBiT extracellular detection system to clarify the mechanism(s) by which leptin modulates cell surface localization of AT1A.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

AT1aR dependent GABAa inhibition in the MnPO

The median preoptic nucleus (MnPO), located along the anterior wall of the third ventricle, receives input from other circumventricular organs (e.g. SFO and the OVLT) sensitive to circulating Angiotensin II (Ang II) and plasma Na+ concentrations suggesting an involvement in hydromineral balance and blood pressure regulation. Additionally, evidence suggest that although the MnPO is not directly sensitive to circulating Ang II, the SFO synthesizes Ang II and releases it on the MnPO as a neurotransmitter suggesting the role of the MnPO in hydromineral balance and blood pressure regulation is mediated in part by Ang II. The Ang II activation of AT1aR has also been shown to influence the function of GABAaRs though the mechanisms are still unclear. Here we investigate the role of Ang II signaling via the AT1a receptor in the MnPO and its influence on excitatory/inhibitory balance. Male Sprague‐Dawley rats (250–350g) received microinfusions (0.4 μL) of recombinant AAV construct containing GFP reporter and shRNA against AT1aR (shAT1a) or an AAV construct containing the GFP reporter and a shRNA scramble (shScr) targeted to the MnPO. Two weeks following AAV injection, coronal slices (300 μm) containing the MnPO were cut using standard in vitro slice procedures. Loose patch recordings were obtained from GFP labeled neurons using borosilicate glass micropipettes containing aCSF as the internal solution (1–3 MΩ). Spontaneous action potential firing was recorded in response to focal application of Ang II (100 nM, 30s) or muscimol (100 uM, 30s). Western blot and RT‐qPCR analyses of punch samples containing the MnPO were used to investigate the effect of AT1a KD on GABAa receptor subunits and KCC2 protein and mRNA expression. Brief focal application of Ang II produced a time dependent increase in action potential frequency of MnPO neurons (n = 12). The Ang II dependent increase in activity was blocked by bath application of the AT1aR antagonist Losartan (n = 12). Additionally, Ang II failed to alter firing rate of MnPO neurons in slices prepared from shAT1a injected rats (n = 12). In animals injected with the control vector, the GABAa agonist muscimol decreased action potential activity (n = 10). In rats that received microinjections of the shAT1a, muscimol failed to decrease action potential frequency (n = 10). In AT1a KD rats, RT‐qPCR analysis shows a reduction in AT1a and KCC2 mRNA but no reduction in GABAa Beta subunit mRNA in the MnPO. The current findings demonstrate that Ang II dependent increases in the excitability of MnPO neurons are mediated by AT1aRs. Moreover, AT1aRs are also necessary for the inhibitory effects of GABAaR activation by muscimol. The current study suggests the reduction in GABAa dependent inhibition following AT1a KD could be mediated by a down regulation of KCC2 and subsequent disruption of chloride homeostasis. Taken together, AT1aR in the MnPO contribute to a balance of excitatory and inhibitory activity in MnPO neurons. This balance could be important for the MnPO's role in the regulation of blood pressure and hydromineral balance. However, the signaling mechanisms underlying theses contributions of AT1aRs to MnPO activity remain to be determined.Support or Funding InformationSupported by P01 HL088052

The effect of resveratrol on angiotensin II levels and the rate of transcription of its receptors in the rat cardiac hypertrophy model.

This study investigated the effect of resveratrol on serum and cardiac levels of angiotensin II and transcription of its main receptors following pressure overload induced-hypertrophy. Rats were divided into untreated (Hyp) and resveratrol treated hypertrophied groups (H+R). Intact animals served as the control (Ctl). Cardiac hypertrophy was induced by abdominal aortic banding. Blood pressure (BP) was recorded via left carotid artery cannula. Fibrosis was confirmed by Masson trichrome staining. Angiotensin II level was measured using an ELIZA test. Gene expression was assessed by a real time PCR (RT-PCR) technique. We observed that in the H+R group BP and heart weight/body weight were decreased significantly (p<0.001, p<0.05, respectively vs Hyp). The cardiac levels of angiotensin II and AT1a mRNA were increased in the Hyp group (p<0.01 vs Ctl). In the H+R group the AT1a mRNA level was decreased significantly (p<0.05 vs Hyp). It could be concluded that resveratrol protects the heart against hypertrophy progression in part by affecting cardiac AT1a transcription.

Effect of Chronic Mild Stress on AT1 Receptor Messenger RNA Expression in the Brain and Kidney of Rats.

The purpose of the study was to determine whether exposure to chronic mild stress (CMS) affects expression of angiotensin II Type 1a receptor (AT1aR) messenger RNA (mRNA) in the brain and kidney. Male Sprague-Dawley rats were divided into an unchallenged control group, which remained at rest, and an experimental group, exposed to CMS produced by a series of unexpected, disturbing stimuli applied at random over a period of 4 weeks. After sacrificing the animals, samples of the septal/accumbal and hypothalamic/thalamic diencephalon, brain medulla, cerebellum, and the renal medulla were harvested for determination of AT1aR mRNA. Expression of AT1a receptor mRNA was significantly greater in the rats in the CMS condition than in the controls (septal/accumbal diencephalon: 1.689 [0.205] versus 0.027 [0.004], hypothalamic/thalamic diencephalon: 1.239 [0.101] versus 0.003 [0.001], brain medulla: 2.694 [0.295] versus 0.028 [0.003], cerebellum: 0.013 [0.002] versus 0.005 [0.001; p < .001 for all comparisons], and renal medulla: 409.92 [46.92] versus 208.06 [30.56; p < .01]). There was a significant positive correlation between AT1a mRNA expression in the septal/accumbal diencephalon and brain medulla (p < .025). The results provide evidence that CMS significantly enhances expression of the AT1aR gene in the brain and kidney and indicate that changes in expression of AT1aR mRNA in different brain regions during CMS may be causally related. It is suggested that the up-regulation of AT1a receptors by chronic stress may potentiate negative effects of angiotensin II in pathologies associated with activation of the renin-angiotensin system.

Abstract 458: Loss of Smooth Muscle α-actin in Mice Results in Thoracic Aortic Aneurysms via Increased Reactive Oxygen Species, Increased Nox4,and Increased Angiotensin II type 1 Receptor-Mediated Signaling

Objective: ACTA2 mutations cause 10-14% of familial thoracic aortic aneurysms and dissections. Mice deficient in smooth muscle α-actin (Acta2-/-) develop root and ascending thoracic aortic enlargement associated with thickening of the aortic media and fragmentation and disarray of elastic fibers. We hypothesized that blocking AT1 activation would block the aortic pathology and prevent aortic enlargement in Acta2-/- mice. Methods and Results: Beginning at 4 weeks of age, Acta2-/- mice were treated with losartan or placebo (n≥10) for 6 months and echocardiograms were performed at baseline and every other month. The aortic root in Acta2-/- mice was found to undergo progressive dilatation. After 6 months of treatment, there was no difference in the diameter of the aortic root between wild-type (WT) mice and the losartan treated mice (p=0.44). Histologic analysis of Acta2-/- aortas demonstrated medial thickening and fragmentation of elastic fibers which was normalized by treatment with losartan. Gene expression of matrix metalloproteinase-2 and -9 (Mmp2, Mmp9), along with lumican and decorin, interleukin-6 (Il6) and phosphorylation of RelA (a subunit of nuclear factor κB, NF-κB) was increased in Acta2-/- aortas, and was corrected by treatment with losartan. NADPH oxidase 4 and AT1a mRNA was increased in Acta2-/- aortic smooth muscle cells (SMCs) and aortas. Increase of AT1a was blocked by lowering reactive oxygen species (ROS) with N-actetyl cysteine (NAC). Angiotensin II (AngII) dose response studies suggested Acta2-/- aortic SMCs had increased sensitivity to Ang II. Additionally, Acta2-/- SMCs had increased ROS compared to WT by flow cytometry (P&lt;0.05). Increased ROS was also observed in aortic samples derived from Acta2-/- mice. Blocking ROS using NAC attenuated aneurysm formation by reducing the expression of Mmps and IL6 and reducing signaling through AT1 in Acta2-/- mice. Genetic deletion of AT1a attenuated ROS and expression of Mmps and IL6 levels in the Acta2-/-aortas but not to WT levels. Conclusions: Our results demonstrate that complete loss of α-SMA leads to aortic dilation and pathologic changes by increasing SMC ROS levels, thus increasing sensitivity to AngII, which results in NF-κB activation and increased expression of Mmps and IL6.

A Subpressor Dose of Angiotensin II Elevates Blood Pressure in a Normotensive Rat Model by Oxidative Stress

Oxidative stress is an imbalance between free radicals and antioxidants, and is an important etiological factor in the development of hypertension. Recent experimental evidence suggests that subpressor doses of angiotensin II elevate oxidative stress and blood pressure. We aimed to investigate the oxidative stress related mechanism by which a subpressor dose of angiotensin II induces hypertension in a normotensive rat model. Normotensive male Wistar rats were infused with a subpressor dose of angiotensin II for 28 days. The control group was sham operated and infused with saline only. Plasma angiotensin II and H2O2 levels, whole-blood glutathione peroxidase, and AT-1a, Cu/Zn SOD, and p22phox mRNA expression in the aorta was assessed. Systolic and diastolic blood pressures were elevated in the experimental group. There was no change in angiotensin II levels, but a significant increase in AT-1a mRNA expression was found in the experimental group. mRNA expression of p22phox was increased significantly and Cu/Zn SOD decreased significantly in the experimental group. There was no significant change to the H2O2 and GPx levels. Angiotensin II manipulates the free radical-antioxidant balance in the vasculature by selectively increasing O2(-) production and decreasing SOD activity and causes an oxidative stress induced elevation in blood pressure in the Wistar rat.

Role of the renin-angiotensin system, renal sympathetic nerve system, and oxidative stress in chronic foot shock-induced hypertension in rats.

Objective: The renin-angiotensin system (RAS) and renal sympathetic nerve system (RSNS) are involved in the development of hypertension. The present study is designed to explore the possible roles of the RAS and the RSNS in foot shock-induced hypertension.Methods: Male Sprague-Dawley rats were divided into six groups: control, foot shock, RSNS denervation, denervation plus foot shock, Captopril (angiotensin I converting enzyme inhibitor, ACE inhibitor) plus foot shock, and Tempol (superoxide dismutase mimetic) plus foot shock. Rats received foot shock for 14 days. We measured the quantity of thiobarbituric acid reactive substances (TBARS), corticosterone, renin, and angiotensin II (Ang II) in plasma, the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), and renal noradrenaline content. RAS component mRNA and protein levels were quantified in the cerebral cortex and hypothalamus.Results: The two week foot shock treatment significantly increased systolic blood pressure, which was accompanied by an increase in angiotensinogen, renin, ACE1, and AT1a mRNA and protein expression in the cerebral cortex and hypothalamus, an increase of the plasma concentrations of renin, Ang II, corticosterone, and TBARS, as well as a decrease in plasma SOD and GSH-Px activities. Systolic blood pressure increase was suppressed by denervation of the RSNS or treatment with Captopril or Tempol. Interestingly, denervation or Tempol treatment both decreased main RAS components not only in the circulatory system, but also in the central nervous system. In addition, decreased antioxidant levels and increased TBARS and corticosterone levels were also partially restored by denervation or treatment with Tempol or Captopril.Conclusions: RAS, RSNS and oxidative stress reciprocally potentiate to play important roles in the development of foot shock-induced hypertension.

Telmisartan protects central neurons against nutrient deprivation-induced apoptosis in vitro through activation of PPARγ and the Akt/GSK-3β pathway

To determine whether angiotensin II receptor blockers (ARBs) could protect central neurons against nutrient deprivation-induced apoptosis in vitro and to elucidate the underlying mechanisms. Primary rat cerebellar granule cells (CGCs) underwent B27 (a serum substitute) deprivation for 24 h to induce neurotoxicity, and cell viability was analyzed using LDH assay and WST-1 assay. DNA laddering assay and TUNEL assay were used to detect cell apoptosis. The expression of caspase-3 and Bcl-2, and the phosphorylation of Akt and GSK-3β were detected using Western blot analysis. AT1a mRNA expression was determined using RT-PCR analysis. B27 deprivation significantly increased the apoptosis of CGCs, as demonstrated by LDH release, DNA laddering, caspase-3 activation and positive TUNEL staining. Pretreatment with 10 μmol/L ARBs (telmisartan, candesartan or losartan) partially blocked B27 deprivation-induced apoptosis of CGCs with telmisartan being the most effective one. B27 deprivation markedly increased the expression of AT1a receptor in CGCs, inhibited Akt and GSK-3β activation, decreased Bcl-2 level, and activated caspase-3, which were reversed by pretreatment with 1 μmol/L telmisartan. In addition, pretreatment with 10 μmol/L PPARγ agonist pioglitazone was more effective in protecting CGCs against B27 deprivation-induced apoptosis, whereas pretreatment with 20 μmol/L PPARγ antagonist GW9662 abolished all the effects of telmisartan in CGCs deprived of B27. ARBs, in particular telmisartan, can protect the nutrient deprivation-induced apoptosis of CGCs in vitro through activation of PPARγ and the Akt/GSK-3β pathway.

ROCK/NF-κB axis-dependent augmentation of angiotensinogen by angiotensin II in primary-cultured preglomerular vascular smooth muscle cells

In angiotensin II (ANG II)-dependent hypertension, the augmented intrarenal ANG II constricts the renal microvasculature and stimulates Rho kinase (ROCK), which modulates vascular contractile responses. Rho may also stimulate angiotensinogen (AGT) expression in preglomerular vascular smooth muscle cells (VSMCs), but this has not been established. Therefore, the aims of this study were to determine the direct interactions between Rho and ANG II in regulating AGT and other renin-angiotensin system (RAS) components and to elucidate the roles of the ROCK/NF-κB axis in the ANG II-induced AGT augmentation in primary cultures of preglomerular VSMCs. We first demonstrated that these preglomerular VSMCs express renin, AGT, angiotensin-converting enzyme, and ANG II type 1 (AT1) receptors. Furthermore, incubation with ANG II (100 pmol/l for 24 h) increased AGT mRNA (1.42 ± 0.03, ratio to control) and protein (1.68 ± 0.05, ratio to control) expression levels, intracellular ANG II levels, and NF-κB activity. In contrast, the ANG II treatment did not alter AT1a and AT1b mRNA levels in the cells. Treatment with H-1152 (ROCK inhibitor, 10 nmol/l) and ROCK1 small interfering (si) RNA suppressed the ANG II-induced AGT augmentation and the upregulation and translocalization of p65 into nuclei. Functional studies showed that ROCK exerted a greater influence on afferent arteriole responses to ANG II in rats subjected to chronic ANG II infusions. These results indicate that ROCK is involved in NF-κB activation and the ROCK/NF-κB axis contributes to ANG II-induced AGT upregulation, leading to intracellular ANG II augmentation.

Angiotensin II contractile effects in mouse colon: role for pre‐ and post‐junctional AT1A receptors

This study investigates whether a local renin-angiotensin system (RAS) exists in mouse colon and whether angiotensin II (Ang II) may play a role in the regulation of the contractile activity. Isometric recordings were performed in vitro on the longitudinal muscle of mouse proximal and distal colon. Transcripts encoding for RAS components were investigated by RT-PCR. Ang II caused, in both preparations, a concentration-dependent contractile effect, antagonized by losartan, AT(1) receptor antagonist, but not by PD123319, AT(2) receptor antagonist. The combination of losartan plus PD123319 caused no change on the Ang II-induced contraction than losartan alone. Tetrodotoxin, neural blocker, reduced the contractile response to Ang II in the proximal colon, whilst the response was abolished in the distal colon. In both preparations, atropine, muscarinic receptor antagonist, or SR140333, NK(1) receptor antagonist, reduced the Ang II responses. Ondansetron, 5-HT(3) receptor antagonist, SR48968, NK(2) receptor antagonist, or hexamethonium, nicotinic receptor antagonist, were ineffective. The joint application of atropine and SR140333 produced no additive effect. Atropine reduced NK(1) -induced contraction. Transcripts encoding RAS components were detected in the colon samples. However, just AT(1A) mRNA was expressed in both preparations, and AT(2) mRNA was expressed only in the distal colon. In the murine colon, local RAS may play a significant role in the control of contractile activity. Ang II positively modulates the spontaneous contractile activity via activation of post-junctional and pre-junctional AT(1A) receptors, the latter located on the enteric neurones, modulating the release of tachykinins and acetylcholine.

Cardiovascular effects of Tulbaghia violacea Harv. (Alliaceae) root methanolic extract in Dahl salt-sensitive (DSS) rats

Ethnopharmacological relevanceIn African folk medicine, Tulbaghia violacea Harv.(Alliaceae), a herbaceous plant also known by other common local names such as ‘wild garlic’, ‘society garlic’ and ‘sweet garlic’, has been used for the management, control and/or treatment of several human ailments; including hypertension.The aim of the present study was to investigate the cardiovascular effects of methanolic extract of Tulbaghia violacea Harv. in Dahl salt-sensitive (DSS) rats. Materials and methodsDSS rats were treated intraperitoneally (i.p) as follows: distilled water, control (CON) (3ml/kg/b.w.); captopril (CAP) (25mg/kg/b.w.); or methanolic extract of Tulbaghia violacea (TVL) (50mg/kg/b.w.) for 7 weeks. Systolic blood pressure (SBP) was measured bi-weekly whilst 24-hour urine volumes and sodium concentrations were assessed weekly. Animals were sacrificed on day 49 by halothane overdose. Blood was removed for the determination of aldosterone concentration and angiotensin converting enzyme (ACE) activity. Left kidney was harvested for angiotensin Ia (AT1a) receptor gene expression. ResultsCON showed a significant percentage increase in SBP (mmHg) on day 49 compared to day 0 (baseline values) (9.75±1.7% vs. 2.33±0.77%; p<0.05). CAP and TVL showed significant percentage decreases in SBP at week 7 when compared to baseline (8.13±1.23% and12.88±3.12% p<0.01). Urine volumes (ml) increased significantly in CAP from week 1 to week 4 (9.63±2.22 vs.10.97±1.75 p<0.05) and in TVL from week 2 to 7 (9.23±0.91 vs. 11.90±1.17 p<0.05). Urinary sodium concentration (mmol/L/24h urine samples) was significantly higher in CAP (0.72±0.07 vs.1.31±0.03 p<0.05) and in TVL (0.71±0.09 vs. 1.68±0.16; p<0.05) at week 7 compared to baseline levels. There was no significant difference in the plasma aldosterone levels (pmol/L) of all the three groups. Serum ACE (U/L) activity was significantly higher in CAP compared to CON and TVL (277.0±11.6 vs. 121.5±3.93; and 147.0±6.63; p<0.05 respectively). CAP showed a significant 6.4 fold increase in AT1a mRNA expression vs. CON (p<0.05) and a 4.5 fold increase vs. TVL (p<0.05). There was no significant difference between CON and TVL treated rats. CAP and TVL showed no significant change in the expression of AT2. ConclusionThis study showed that TVL possesses antihypertensive effect in Dahl salt-sensitive rats when administered over a long-term.

Renin-angiotensin system gene expression in the kidney and in the heart in hypertensive ISIAH rats

The content of mRNA of renin-angiotensin system (RAS) genes in the kidney and heart of hypertensive ISIAH and normotensive WAG rats was measured by the real-time PCR. Statistically significant decrease of RAS gene mRNA was registered in the kidney of ISIAH rats, including Ren (by 45%), Ace (43%), AT1A (34%), COX-2 (50%). In the myocardium AT1A mRNA expression decreased by 28% while Ace mRNA expression increased by 80%. These results demonstrate the reduction of renal RAS basal activity in the hypertensive ISIAH rats, and this allows us to consider the ISIAH rat, as a low-renin hypertensive strain. In support of this viewpoint, in the ISIAH rats, a two-fold increase in the connective tissue sodium concentration as well as statistically significant plasma sodium increase (from 136 +/- 0,25 micromol/l in WAG to 139 +/- 0,3 micromol/l in the ISIAH rats) were found. Our conclusion backed by a tendency of the ISIAH plasma aldosterone level decrease giving in sum a classical picture of a low-renin hypertensive state in the ISIAH rats. It was suggested that the formation of low-renin arterial hypertension in the ISIAH rats may depend on changes in kidney ion channels function. In addition, renal NO system alterations could be also involved in the pathogenesis of arterial hypertension in the ISIAH rats.

The midgestational maternal blood pressure decline is absent in mice lacking expression of the angiotensin II AT2 receptor

The midgestational maternal blood pressure (BP) decrease is absent in mice treated with an angiotensin II AT2 receptor blocker. We tested the hypotheses that there would be 1) no midgestational decrease in maternal systolic BP (SBP) in AT2-/- mice, and 2) a pattern of increased AT2 and/or decreased AT1a mRNA expression in tissues from normal (wild-type, WT) mice, corresponding with SBP changes. Heart, aorta, placenta and kidney tissue were obtained from WT and AT2-/- mice before pregnancy and on gestational days (Gd) 5-6, 12-13 and 18-19. AT1a and AT2 mRNA expression was quantified. SBP was measured. SBP was significantly decreased in WT Gd12-13 mice, but did not change during pregnancy in AT2-/- mice. In WT mice, aortic AT1a mRNA expression levels were significantly higher at Gd12-13 and Gd18-19 compared with before pregnancy. AT1a kidney and heart mRNA did not change during pregnancy. There were no changes in AT2 mRNA expression. There was no distinct pattern of change in AT1a expression in AT2-/mice. Placental AT1a and AT2 expression levels increased markedly between Gd12-13 and Gd18-19 in WT mice. We conclude that the AT2 receptor is essential for the midgestational SBP decline in WT mice. There is no consistent relationship between changes in tissue angiotensin II receptor mRNA expression and SBP in WT mice.

High-salt diet during pregnancy and angiotensin-related cardiac changes

High-salt intake has been demonstrated in link to hypertension, and cardiovascular diseases could be programmed in fetal origins. We determined the influence of high-salt diet during pregnancy on the development of the heart. Fetal cardiac structures, cell cycle, renin-angiotensin system (RAS), and epigenetic alternations in the heart following maternal high salt intake during pregnancy were examined. Following exposure to high salt, disorganized myofibrillae and mitochondria cristae loss were found in the fetus, S-phase for cardiac cells was enhanced, plasma angiotensin II decreased, and cardiac angiotensin II increased in the fetus. Angiotensin II-increased S-phase in the fetal cardiac cells was primarily via AT1 receptor mechanisms. AT2 receptor mRNA and protein in the fetal heart were not affected, whereas AT1 receptor protein, AT1a, and AT1b mRNA were increased. DNA methylation was found at the CpG sites that were related to AT1b receptors in the fetal heart. Cardiac AT1 receptor protein in the adult offspring was also higher following exposure to prenatal high salt. The results suggest a relationship between high-salt diet in pregnancy and developmental changes of the cardiac cells and renin-angiotensin system.

Angiotensin II-induced reduction in body mass is Ang II receptor mediated in association with elevated corticosterone

The mechanisms by which elevated glucocorticoids contribute to decreased body mass via angiotensin II (Ang II) infusion are not completely described. This study addressed the hypothesis that chronic Ang II infusion suppresses hepatic growth hormone receptor (GHr) and IGF1 expressions via an Ang II receptor (AT1)-mediated pathway associated with elevated glucocorticoids. Sprague–Dawley rats were assigned to three groups: 1) Control, 2) Ang II-infused (80 ng/min x 28d) and 3) Ang II + angiotensin receptor blocker (ARB; 10 mg losartan/kg/d × 21d). After 28 d, Ang II decreased body mass by 14% (407 ± 8 vs 350 ± 17 g) and hepatic AT1a, GHr, and IGF1 mRNA expressions by 45%, 44%, and 44%, respectively. ARB treatment completely prevented the loss in body mass (409 ± 9 g) and AT1a and GHr expressions and partially recovered the loss of hepatic IGF1. Ang II increased plasma corticosterone (B) 3-fold (173 ± 28 vs 555 ± 42 ng/mL) and ARB treatment prevented the response (150 ± 47 ng/mL). Food consumption did not change suggesting that the decrease in body mass resulted from the catabolic actions of the Ang II-induced increase in systemic B and not from reduced caloric intake. The prevention by ARB treatment of the Ang II-induced decrease in body mass and downregulation of AT1a, GHr and IGF1 coinciding with suppression of plasma B suggests that the Ang II-induced decrease in body mass is AT1 receptor mediated in conjunction with elevated B. These data suggest that alleviating the Ang II-induced cachexia requires targeting AT1 and suppressing glucocorticoid secretion.

Angiotensin II receptors in a rat model of hypobaric hypoxic pulmonary hypertension

Hypoxic pulmonary hypertension is characterized by profound remodeling of the walls of small pulmonary arteries: endothelial swelling and proliferation of media and adventitia to a degree that may narrow the lumen. The molecular mechanism responsible for these changes is unknown. Recent studies have demonstrated the existence of a vascular and cardiac renin-angiotensin system. This system can induce hypertrophy and proliferation of aortic smooth muscle cells and cardiac muscle cells in animal models of systemic hypertension. In the present experiments, Northern blot analysis and a receptor binding assay specific to angiotensin II (Ang II) receptors was used to study the pulmonary renin angiotensin system in a rat model of hypobaric hypoxic pulmonary hypertension. mRNA transcripts for the Ang II receptor AT(1A) were constitutively expressed, and specific binding of Ang II to the receptors occurred in lung tissue of control rats that breathed ambient air. After 3 days of hypobaric hypoxia (380 Torr), the AT(1A) mRNA level and the index of Ang II receptors had increased, but right ventricular hypertrophy was not still evident. Ang II receptors were significantly increased after 14 days of hypoxia, when histological evidence of pulmonary hypertension was observed. In addition, the Ang II level in pulmonary tissue was higher after 14 days of hypoxia than in normal controls. These findings suggest that the renin-angiotensin system in pulmonary tissue was activated in these rats, and that it contributes to the morphological changes observed in hypoxic pulmonary hypertension.

Angiotensin II-induced hypertension regulates AT1 receptor subtypes and extracellular matrix turnover in mouse retinal pigment epithelium

Accumulation of specific deposits and extracellular molecules under the retinal pigment epithelium (RPE) has been previously observed in eyes with age-related macular degeneration (AMD) and may play a role in the pathogenesis of AMD. Even though age is the major determinant for developing AMD, clinical studies have revealed hypertension (HTN) as another systemic risk factor. Angiotensin II (Ang II) is considered the most important hormone associated with HTN. To evaluate the relationship of Ang II to AMD, we studied whether mouse RPE expresses functional Ang II receptor subtypes and whether HTN-induced Ang II regulates expression of these receptors as well as critical ECM molecules (MMP-2 and type IV collagen) involved in ECM turnover in RPE. We used 9-month-old C57BL/6 male mice infused with Ang II alone or Ang II in combination with the AT1 receptor antagonist candesartan or the AT2 receptor antagonist PD123319 for 4 weeks to determine whether HTN-associated Ang II was important for ECM regulation in RPE. We found that mouse RPE expressed both Ang II receptor subtypes at the mRNA and protein levels. Infusion with Ang II induced HTN and elevated plasma and ocular Ang II levels. Ang II also regulated AT1a and AT1b receptor mRNA expression, the intracellular concentration of calcium [Ca 2+] i, MMP-2 activity, and type IV collagen accumulation. Concurrent administration of Ang II with the AT1 receptor blocker prevented the increase in blood pressure and rise in ocular Ang II levels, as well as the calcium and MMP-2 responses. In contrast, the type IV collagen response to Ang II was prevented by blockade of AT2 receptors, but not AT1 receptors. Plasma Ang II levels were not modified by the AT1 or AT2 receptor blockade. Since the effects of Ang II on MMP-2 and type IV collagen require inhibition of both Ang II receptor subtypes, these receptors may play a role as a potential therapeutic targets to prevent ECM turnover dysregulation in the RPE basement membrane, suggesting a pathogenic mechanism to explain the link between HTN and AMD.

Expression and transport of Angiotensin II AT 1 receptors in spinal cord, dorsal root ganglia and sciatic nerve of the rat

To clarify the role of Angiotensin II in the regulation of peripheral sensory and motor systems, we initiated a study of the expression, localization and transport of Angiotensin II receptor types in the rat sciatic nerve pathway, including L 4–L 5 spinal cord segments, the corresponding dorsal root ganglia (DRGs) and the sciatic nerve. We used quantitative autoradiography for AT 1 and AT 2 receptors, and in situ hybridization to detect AT 1A, AT 1B and AT 2 mRNAs. We found substantial expression and discrete localization of Angiotensin II AT 1 receptors, with much higher numbers in the grey than in the white matter. A very high AT 1 receptor expression was detected in the superficial dorsal horns and in neuronal clusters of the DRGs. Expression of AT 1A mRNA was significantly higher than that of AT 1B. AT 1 receptor binding and AT 1A and AT 1B mRNAs were especially prominent in ventral horn motor neurons, and in the DRG neuronal cells. Unilateral dorsal rhizotomy significantly reduced AT 1 receptor binding in the ipsilateral side of the superficial dorsal horn, indicating that a substantial number of dorsal horn AT 1 receptors have their origin in the DRGs. After ligation of the sciatic nerve, there was a high accumulation of AT 1 receptors proximal to the ligature, a demonstration of anterograde receptor transport. We found inconsistent levels of AT 2 receptor binding and mRNA. Our results suggest multiple roles of Angiotensin II AT 1 receptors in the regulation of sensory and motor functions.