Arteries Of SHR Research Articles

Elevated Vascular Sympathetic Neurotransmission and Remodelling Is a Common Feature in a Rat Model of Foetal Programming of Hypertension and SHR.

Hypertension is of unknown aetiology, with sympathetic nervous system hyperactivation being one of the possible contributors. Hypertension may have a developmental origin, owing to the exposure to adverse factors during the intrauterine period. Our hypothesis is that sympathetic hyperinnervation may be implicated in hypertension of developmental origins, being this is a common feature with essential hypertension. Two-animal models were used: spontaneously hypertensive rats (SHR-model of essential hypertension) and offspring from dams exposed to undernutrition (MUN-model of developmental hypertension), with their respective controls. In adult males, we assessed systolic blood pressure (SBP), diastolic blood pressure (DBP), heart rate (HR), sympathetic nerve function (3H-tritium release), sympathetic innervation (immunohistochemistry) and vascular remodelling (histology). MUN showed higher SBP/DBP, but not HR, while SHR exhibited higher SBP/DBP/HR. Regarding the mesenteric arteries, MUN and SHR showed reduced lumen, increased media and adventitial thickness and increased wall/lumen and connective tissue compared to respective controls. Regarding sympathetic nerve activation, MUN and SHR showed higher tritium release compared to controls. Total tritium tissue/tyrosine hydroxylase detection was higher in SHR and MUN adventitia arteries compared to respective controls. In conclusion, sympathetic hyperinnervation may be one of the contributors to vascular remodelling and hypertension in rats exposed to undernutrition during intrauterine life, which is a common feature with spontaneous hypertension.

19 - Apocynin reduces pressor and vasoconstrictor effects of Ang II in SHR

NAD(P)H oxidase (NOX) activity and expression can be activated by angiotensin (Ang) II. We have previously observed that apocynin, a NOX inhibitor, prevented endotelial dysfunction and reduced blood pressure by increasing nitric oxide (NO) and reducing ROS concentrations in endothelial cells. We evaluated the effect of apocynin on the contractile responses to Ang II in resistance arteries of SHR and the mechanisms involved on these effects. SHR were treated from the 4th to the 10th week of life with apocynin (30 mg/Kg). Wistar rats were used as normotensive control. Using mesenteric arteries, we performed concentration-response curves to Ang II and determined eNOS and NOX isoforms and subunits expressions, lucigenin chemiluminescence and nitrate/nitrite levels. Data were expressed as mean ± SEM. Apocynin increased endothelium modulation and/or NOS activity on Ang II vasoconstrictor responses in mesenteric arteries of SHR. Treatment with apocynin did not alter NO synthase activity, and eNOS, NOX1, NOXO1, and NOX4 expressions, however, it decreased NOX2 and p47phox expressions in mesenteric beds of SHR treated. Moreover, treatment apocynin was able to decrease ROS production in these vessels. The lower reactivity of Ang II in resistance arteries would lead to lower peripheral vascular resistance and consequently the reduction of mean arterial pressure and Ang II pressor effect in SHR treated with apocynin, as previously observed. All these results demonstrated that in vivo treatment with apocynin induces important alterations of several mechanisms that lead to the reduction of the pressor and vasoconstrictor effects of Ang II in SHR. Apocynin effect involves further mechanisms besides the modulation of vascular ROS, which improve NO availability in vascular cells of SHR. Ethics Comittee CEUA FOA 450/2015. Financial Support FAPESP 2016/22180-9, CAPES and CNPq.

The Role of Melatonin In Exercise Enhanced Endothelium-dependent Vasorelaxation In Mesenteric Arteries of SHR

PURPOSE: To determine if melatonin plays an important role in the hypotensive effects of exercise training. Further, to study the role of melatonin binding to melatonergic receptors in the vasorelaxation of small MAs in SHR. METHODS: Twelve-week-old male normotensive Wistar-Kyoto rats (WKY, n=18) and SHRs (n = 48) were used. SHRs were randomly divided into four groups: SHR sedentary group (SHR-SED, n = 18), SHR sedentary with melatonin receptor antagonist luzindole (Luz) injection group (SHR-SED + Luz, n = 6), SHR exercise group (SHR- EX, n = 18) , and SHR exercise with luzindole injection group (SHR-EX + Luz, n = 6). Luzindole injection groups were injected intraperitoneally (i.p.) with luzindole (1 mg/ kg/day in sterile saline) each day. Rats in the exercise groups were subjected to aerobic exercise. Blood pressure and heart rate were measured after exercise. Serum melatonin levels were examined by ELISA. The mechanical properties of small mesenteric arteries were studied with myograph. Western blot and immunofluorescence colocalization were performed to examine the protein expression and distribution of MT1, MT2 receptors and eNOS. RESULTS: Exercise training produced a significant reduction in blood pressure and heart rate in SHR, which was significantly attenuated by intraperitoneal administration of luzindole, a nonselective melatonin receptor (MT1/MT2) antagonist. Serum melatonin levels in the SHR group were significantly lower than those in the WKY group at 8:00-9:00 and 21:00-22:00, and exercise training reduced this difference. Endothelium-dependent vessel relaxation induced by acetylcholine was significantly blunted in SHR compared with age-matched WKY. Both exercise training and luzindole ameliorated this endothelium-dependent impairment of relaxation in hypertension. Immunohistochemistry and Western blotting showed that protein expression of the MT2 receptor and eNOS, as well as their colocalization in the endothelial cell layer in SHR, was significantly decreased; exercise training suppressed this reduction. CONCLUSIONS: These results provide evidence that regular exercise has a beneficial effect on improving endothelium-dependent vasorelaxation in MAs, in which melatonin plays a critical role by acting on MT2 receptor to increase NO production and/or NO bioavailability.

Altered contractile responses of arteries from spontaneously hypertensive rat: The role of endogenous mediators and membrane depolarization

AimsThe goal of our study was to reveal the important mechanism(s) responsible for the enhanced contractility of isolated arteries from animals suffering genetic hypertension. Main methodsContractile force of endothelium-denuded arteries, modulated by various interventions, was measured by wire myography. Key findingsSpontaneously hypertensive rat (SHR) and Wistar-Kyoto rat (WKY) arteries were stimulated by norepinephrine, increased extracellular K+ or tyramine. Strain difference was not observed in the contraction elicited by exogenous norepinephrine but SHR arteries responded more to tyramine (causing endogenous norepinephrine release from neuronal varicosities). K+-induced contraction was enhanced in SHR arteries, with no involvement of endogenous catecholamines. The α-adrenoceptor blockade lowered tyramine-induced contraction more in SHR arteries; similar effect was achieved by guanethidine-induced sympathectomy. Partial depolarization of WKY arteries by 20mM K+ enhanced its contraction to SHR level. The blockade of β-adrenoceptors by propranolol or selective β2-antagonist ICI-118,551 induced contraction of SHR endothelium-denuded arteries but was without significant effects on WKY arteries unless they were stimulated with K+. Both tyramine-induced and propranolol-induced contractions were attenuated by flupirtine and abolished by nifedipine. SignificanceThe differences of SHR and WKY arteries were not related to vascular expression of α- and β-adrenoceptors or G-proteins. Enhanced contractility of SHR arteries is related to both increased presence of endogenous norepinephrine in vascular wall and also to altered vascular smooth muscle membrane potential.

Intracellular renin increases the inward calcium current in smooth muscle cells of mesenteric artery of SHR. Implications for hypertension and vascular remodeling

The influence of intracellular renin on the inward calcium current in isolated smooth muscle cells from SHR mesenteric arteries was investigated. Measurements of calcium current were performed using the whole cell configuration of pCLAMP. The results indicated that: 1) renin (100nM) dialyzed into smooth muscle cells, increased the inward calcium current; 2) verapamil (10–9M) administered to the bath inhibited the effect of renin on the inward calcium current; 3) concurrently with the increase of calcium current a depolarization of 6.8+/−2.1mV (n=16)(P<0.05) was found in cells dialyzed with renin; 4) intracellular dialysis of renin (100nM) into smooth muscle cells isolated from mesenteric arteries of normal Wystar Kyoto rats showed no significant change on calcium current; 5) aliskiren (10–9M) dialyzed into the cell together with renin (100nM) abolished the effect of the enzyme on the calcium current in SHR; 6) Ang II (100nM) dialyzed into the smooth muscle cell from mesenteric artery of SHR in absence of renin, decreased the calcium current-an effect greatly reduced by valsartan (10–9M) added to the cytosol; 7) administration of renin (100nM) plus angiotensinogen (100nM) into the cytosol of muscles cells from SHR rats reduced the inward calcium current; 8) extracellular administration of Ang II (100nM) increased the inward calcium current in mesenteric arteries of SHR. Conclusions: intracellular renin in vascular resistance vessels from SHR due to internalization or expression, contributes to the regulation of vascular tone and control of peripheral resistance-an effect independently of Ang II. Implications for hypertension and vascular remodeling are discussed.

Demethoxycurcumin Preserves Renovascular Function by Downregulating COX-2 Expression in Hypertension.

Hypertension-associated endothelial dysfunction is largely due to the exaggerated vasoconstrictor generation by cyclooxygenase-2 (COX-2). COX-2 is induced under inflammatory condition. Demethoxycurcumin (DMC) is a major component of Curcuma longa L, which possesses anti-inflammatory action. This study aimed to examine whether DMC protects endothelial function in hypertension by modulating COX-2. Changes in isometric tension showed that in vivo and ex vivo treatment with DMC rescued the attenuated endothelium-dependent relaxations (EDRs) and elevated endothelium-dependent contractions (EDCs) in the renal arteries of SHR, which were also corrected by acute usage of the COX-2 inhibitor celecoxib. The restoration of renovascular activity by DMC was accompanied by the normalization of COX-2 expression. The enhanced COX-2 expression observed in the renal arteries of hypertensive patients was suppressed by incubation of excised arteries with DMC for 12 hrs. In the renal arteries of Wistar-Kyoto rats (WKY), DMC prevented the endothelial dysfunction caused by angiotensin II. The reduction in the generation of nitric oxide (NO) and expression of eNOS phosphorylation (Ser1177) in human umbilical vein endothelial cells caused by angiotensin II (Ang II) were restored by DMC or celecoxib. Our findings suggest that DMC may decrease COX-2 expression and improve endothelial function in hypertension.

Expression of Calcium Channel Subunit Variants in Small Mesenteric Arteries of WKY and SHR.

Enhanced function of dihydropyridine-sensitive Ca2+ channels (CaV) in hypertensive arterial myocytes (HAM) is well accepted. Increased protein expression of pore forming α1-subunits contributes to this effect, but cannot explain all of the differences in CaV properties in HAM. We hypothesized that differences in expression of CaV subunits and/or their splice variants also contribute. RNA, protein, and myocytes were isolated from small mesenteric arteries (SMA) of 20-week-old male WKY and SHR and analyzed by polymerase chain reaction (PCR), sequencing, immunoblotting, and patch clamp methods. Cav1.2 α1, β2c, and α2δ1d were the dominant subunits expressed in both WKY and SHR with a smaller amount of β3a. Real-time PCR indicated that the mRNA abundance of β3a and α2δ1 but not total Cav1.2 α1 or β2c were significantly larger in SHR. Analysis of alternative splicing of Cav1.2 α1 showed no differences in abundance of mutually exclusive exons1b, 8, 21 and 32 or alternative exons33 and 45. However, inclusion of exon9* was higher and a 73 nucleotide (nt) deletion in exon15 (exon15Δ73) was lower in SHR. Immunoblot analysis showed higher protein levels of Cav1.2 α1 (1.61±0.05), β3 (1.80±0.32), and α2δ1 (1.80±0.24) but not β2 in SHR. The lower abundance of exon15Δ73 transcripts in SHR results in a larger fraction of total Cav1.2 mRNA coding for full-length CaV protein, and the higher abundance of exon9* transcripts and CaVβ3a protein likely contribute to differences in gating and kinetics of CaV currents in SHR. Functional studies of Ca2+ currents in native SMA myocytes and HEK cells transiently transfected with CaV subunits support these conclusions.

C-Src, ERK1/2 and Rho kinase mediate hydrogen peroxide-induced vascular contraction in hypertension: role of TXA2, NAD(P)H oxidase and mitochondria.

: The aim of this study was to analyse the signalling pathways involved in H2O2 vascular responses in hypertension. Vascular function, thromboxane A2 (TXA2) production, oxidative stress and protein expression were determined in mesenteric resistance arteries (MRAs) from hypertensive (spontaneously hypertensive rats, SHR) and normotensive Wistar Kyoto (WKY) rats. H2O2 and the TP agonist U46619 induced greater contractile responses in MRA from SHR than WKY. Moreover, H2O2 increased TXA2 production more in SHR than in WKY. The c-Src inhibitor PP1 reduced H2O2 and U46619-induced contraction and TXA2 release in both strains. The ERK1/2 inhibitor PD98059 reduced H2O2 but not U46619-induced contraction only in SHR arteries. The Rho kinase inhibitor Y26372 reduced H2O2 and U46619-induced contractions only in SHR arteries. Basal c-Src, ERK1/2 and Rho kinase expression were greater in MRA from SHR than WKY. In SHR, the combination of PD98059 with the TP antagonist SQ29548 but not with Y27632 inhibited the H2O2 contraction more than each inhibitor alone. H2O2 and U46619 increased NAD(P)H oxidase activity and O2 production and decreased mitochondrial membrane potential in vessels from SHR. The effects induced by H2O2 were abolished by inhibitors of TXA2 synthase, ERK1/2 and c-Src. The mitochondrial antioxidant mitoTEMPO reduced H2O2-induced contraction and NAD(P)H oxidase activation. In arteries from WKY, c-Src mediates H2O2 contractile responses by modulating TXA2 release and TXA2 effect. In SHR, H2O2 induces c-Src dependent TXA2 release that provokes vascular contractile responses through Rho kinase, c-Src and O2 from NAD(P)H Oxidase and mitochondria. Moreover, ERK1/2 activation contributes to H2O2 contraction in SHR through effects on mitochondria/NAD(P)H Oxidase.

High salt intake does not produce additional impairment in the coronary artery relaxation of spontaneously hypertensive aged rats

The effect of a salt-based diet on the coronary responsiveness in aged hypertensive rats (SHR) still is unclear. We investigated the effects of high salt intake on the relaxation properties of coronary arteries of aged SHRs. Male SHR (32 week-old) received drinking water (SHR) or 1% NaCl solution (SHR-Salt) for 8 weeks. Isolated coronary segments were subjected to concentration-response curves to acetylcholine (ACh) in the presence or absence of L-NAME (100 μM), enalaprilate (10 μM), losartan (10 μM), and spironolactone (100 μM). Salt intake did not increase blood pressure in old SHRs, but caused ventricular hypertrophy. The endothelium-dependent relaxation in SHRs was lower than in Wistar rats. However, salt intake did not add further impairment. Both enalaprilate and losartan reduced the vasodilator response in coronary arteries from Wistar, but did not affect SHR-salt rats. Conversely, losartan attenuated the impaired ACh relaxation observed in SHR. Spironolactone reduced the relaxation induced by ACh in coronary arteries from Wistar rats but not in SHR. The renin-angiotensin-aldosterone system participates in the impaired coronary relaxation in aged SHR, but does not partake in this deleterious effect under increased salt intake, indicating that age could differentiate the effects of high sodium intake in coronary arteries of SHR.

Corrigendum: Ca2+ Channel Inactivation in Small Mesenteric Arteries of WKY and SHR.

Corrigendum: Ca2+ Channel Inactivation in Small Mesenteric Arteries of WKY and SHR.

Transcriptional upregulation of α2δ‐1 subunits elevates CaV1.2 currents in cerebral artery smooth muscle cells of hypertensive rats

Voltage‐dependent L‐type (CaV1.2) channels contain pore‐forming α1 and auxiliary α2δ subunits. CaV1.2 channels are the principal Ca2+ entry pathway in arterial myocytes and regulate physiological functions, including contractility. CaV1.2 currents are larger in arterial myocytes of hypertensive subjects, leading to elevated arterial contractility, but molecular mechanisms that underlie this pathological alteration are unclear. Here, we investigated the molecular identity and physiological functions of α2δ isoforms in resistance‐size cerebral arteries of normotensive (WKY) and hypertensive (SHR) rats. RT‐PCR indicated that α2δ‐1 was the only α2δ isoform expressed in WKY and SHR arterial myocytes. CaV1.2α1 and α2δ‐1 protein were 2‐ and 2.5‐fold higher in SHR than WKY arteries, respectively. Surface biotinylation indicated that both subunits were present primarily in the plasma membrane of SHR and WKY arteries. Arteries from SHR developed more myogenic tone than WKY. Chronic pregabalin (24h), an α2δ‐1 ligand, decreased surface expression of both subunits, reduced CaV1.2 current density, and dilated pressurized arteries more in SHR than WKY. In summary, our data indicate that transcriptional upregulation of α2δ‐1 elevates surface expression of CaV1.2α1 in myocytes, leading to vasoconstriction. We identify α2δ‐1 as a novel molecular target for inducing vasodilation in hypertensive arteries.

Antihypertensive Properties of Angiotensin-Converting Enzyme Inhibitors (ACEI) Independent of the Renin-Angiotensin System

Angiotensin converting enzyme inhibitors (ACEi) lead to inhibition of angiotensin II formation and bradykinin inactivation. However, ACEi also have antihypertensive activity that is independent of the renin-angiotensin system. For example, the enhanced peripheral resistance due to smooth muscle cell hypertrophy and hyperplasia (related to lower apoptosis) increases vascular tonus in established hypertension. ACEi treatment restores susceptibility to apoptosis in these cells through stimulation of the Bax protein. Also, an altered function of the Na+/K+ ATPase leads to elevated levels of intracellular sodium concentration, and depolarization of smooth muscle membranes increasing their excitability. ACEi treatment normalizes this pump function, and the arterial tonus by release of endothelial nitric oxide and prostacyclin. Endothelium- dependent hyperpolarization (to acetylcholine) in mesenteric arteries mediated by release of the endotheliumderived hyperpolarizing factor, is impaired in hypertensive humans and animals. In this instance, ACEi restored this impaired response, since this treatment causes vascular remodeling in resistance SHR arteries through induction of apoptosis of vascular smooth muscle cells reducing the subendothelial thickening, which improves the EDHF circulation. The purpose of this review is to analyze the performance of ACEi in normalizing the vascular tonus during the hypertensive state, which may render new perspectives in basic and clinical research for the next years.

Effect of D3 receptor on α-adrenergic receptor mediated proliferation of vascular smooth muscle cells

results with quantitative real time RT-PCR and Northern Blot. Blood pressure was measured to determine the time of development of hypertension. Results: Among the 10,000 genes examined, we found that expression of the human gene homolog FXYD domain containing ion transport regulator 5 (FXYD5)was downregulated in thekidneys and second-order mesenteric arteries of SHR. FXYD5 expression was 14.8fold lower in SHR than in WKY rats (P<0.01). Northern blot results indicated that expression of FXYD5 occurred in a timeand tissuedependent manner. FXYD5 expression did not differ between SHR and WKY rats at 4 and 8 weeks of age, but FXYD5 mRNA levels were significantly lower in SHR by 13 weeks of age (P<0.01). Peak FXYD5 expression was seen at 8 weeks in SHR and at 13 weeks in WKY rats. Conclusion: The FXYD5gene,which is involved in the induction ofNa, KATPase activity, was significantly suppressed in SHR. This downregulation may have a profound effect on blood pressure regulation. Thus, FXYD5 may serve as a potential target for antihypertensive therapy.

Upregulated TRPC3 expression and function in vascular smooth muscle during hypertension

TRPC3 has been demonstrated to play a role in regulating arterial tone through smooth muscle cell (SMC) depolarization. Recent studies indicate that TRPC3 expression is upregulated in pulmonary artery SMC in idiopathic pulmonary hypertension. We sought to determine if TRPC3 was upregulated in the systemic circulation during hypertension, and if so, whether this upregulation renders arteries more contractile to physiological agonists.Protein was extracted from carotid arteries (CA) of male SHR and WKY rats. Western blot analysis demonstrated 3.1 ± 1.2 times greater TRPC3 expression in SHR versus WKY. Next, TRPC3 function was evaluated in freshly isolated CA SMC by whole cell patch clamp; TRPC3 currents were elicited by extracellular application of uridine triphosphate (UTP; 60 µM). UTP‐stimulated currents were greater in SHR compared with WKY or previous studies with normotensive Long‐Evans rats. Peak UTP‐stimulated inward currents (at ‐110 mV) were ‐36, ‐18, and ‐23 pA, respectively.In summary, we have demonstrated upregulated TRPC3 expression and function in CA during hypertension. Future planned studies will evaluate CA responses to contractile agonists. Supported by: AHA 0665100Y and NIH HL088435 (SPM)

Effect of protein kinase C and protein kinase A inhibitors on contraction of isolated femoral arteries of SHR and Wistar rats

Alterations of calcium handling and other second messenger cascades including protein kinase C (PKC) and A (PKA) were suggested to be responsible for abnormal vascular function in spontaneously hypertensive rats (SHR). However, the relative contribution of these pathways to vasoconstriction is still not completely understood. We investigated the effect of Ro 31-8220 (PKC inhibitor) and H89 (PKA inhibitor) on vasoconstriction induced by 120 mM KCl or by addition of 10 microM noradrenaline (NA) in isolated femoral arteries of control Wistar rats and SHR. Moreover, we investigated these responses in the presence and absence of Ca(2+) ions in the incubation medium in order to assess the role of calcium influx in these contractions. We observed that while the vasoconstriction in the presence of calcium was not different between Wistar and SHR, the difference between constriction elicited by NA addition in the absence and presence of external calcium was larger in SHR. The inhibition of PKC had no effect on constrictions in SHR, but diminished constrictions in Wistar rats. PKA inhibition slightly enhanced constrictions in Wistar rats, but reduced them in the presence of calcium in SHR. We conclude that vasoconstriction elicited by adrenergic stimulation is more dependent on extracellular calcium influx in SHR compared to Wistar rats. Moreover, the activation of PKA contributes to this calcium-dependent vasoconstriction in SHR but not in Wistar. On the other hand, PKC activation seems to play a less important role in vasoconstriction in SHR than in Wistar rats.

Long-term effect of losartan administration on blood pressure, heart and structure of coronary artery of young spontaneously hypertensive rats

Alterations in geometry and structure of coronary arteries have marked consequences on blood flow to the respective area. We evaluated long-term effect of losartan on blood pressure (BP), heart weight/body weight (HW/BW), geometry and structure of septal branch of coronary artery (RS) of young SHR and Wistar rats. Four-week-old Wistar rats and SHR were used. Losartan was administered (20 mg/kg/day) in drinking water by gavage for 5 weeks. BP was measured by plethysmographic method. Cardiovascular system was perfused with a fixative (120 mm Hg). RS was processed for electron microscopy. Wall thickness of intima + media (WT), inner diameter (ID), cross-sectional area of intima + media (CSA), volume densities (VD) of endothelial cells (EC), extracellular matrix (ECM) of intima, smooth muscle cells (SMC) and ECM of media were evaluated. BP of 4-week-old SHR did not differ from that of Wistar rats. BP, HW/BW, WT, CSA, WT/ID, CSAs of SMC, ECM of media were increased in 9-week-old SHR, whereas their VD and CSA of EC were decreased. Losartan administration decreased BP and HW/BW in both groups. Geometry of RS was affected only in SHR (reduction of WT, CSA, WT/ID and increased of ID, circumferential tension, VD and CSA of EC). Losartan administration reduced BP and myocardial mass in both groups and beneficially affected geometry and structure of coronary artery in SHR.

Enhanced Stretch-Induced Myogenic Tone in the Basilar Artery of Spontaneously Hypertensive Rats

We investigated if the magnitude of myogenic tone in the basilar artery of SHR differs from that in WKY and, if so, whether RhoA- or PKC-dependent mechanisms were involved. Myogenic tone was developed in response to stretch. Stretch-induced myogenic contraction was significantly greater in the SHR than WKY in the presence of external Ca²⁺. However, in the absence of external Ca²⁺, stretch did not evoke a myogenic tone. The [Ca²⁺]_i-induced contraction was larger in SHR than WKY and the [Ca²⁺]_i-force curve was significantly shifted to the left in SHR compared to WKY. Y-27632 significantly inhibited stretch-induced myogenic tone, but the inhibitory effect was larger in the SHR than WKY. However, PKC inhibitors had no significant effect on the myogenic tone. RhoA and PKCΕ were expressed at higher levels in the SHR compared to the WKY. RhoA and PKCα translocated from the cytosol to the cell membrane in response to stretch in both animals, but PKCΕ was translocated only in SHR. Our results strongly suggest that stretch-induced myogenic tone is enhanced in SHR, and the activation of RhoA/Rho kinase plays an important role in the enhanced myogenic tone in SHR.

Angiotensin II type 2 receptor activation induces RhoA inhibition through Phosphorylation of its Serine 188 in Vascular Smooth Muscle Cells

Angiotensin II (Ang II) plays important roles in the control of blood pressure. Evidences indicate that Ang II type 2 receptor (AT2R) antagonizes the effects of type 1 receptor (AT1R) and inactivation of RhoA signaling has been suggested to be involve in this action. As serine 188 phosphorylation of RhoA inhibits its activity, we analyzed the effect of Ang II on RhoA phosphorylation both in vitro and in vivo. In rat aortic smooth muscle cells (VSMC), Ang II (0.1 μM) induces serine-phosphorylation of RhoA. This effect is abolished by the AT2R antagonist PD 123319, mimicked by the AT2R agonist CGP42112A and does not involved NOS, PKA or PKG. S188A- or S188E-RhoA mutants expressed in VSMC are not phosphorylated by AT2R stimulation indicating that AT2R activation promotes phosphorylation of RhoA on Ser 188. AT2R-induced phosphorylation of RhoA is associated to an increased binding to the guanidine dissociation inhibitor GDI, leading to inactivation of RhoA/Rho kinase signaling. In vivo, inhibition of AT1R has been shown to up-regulate AT2R expression. We shown that in aorta and pulmonary artery of WKY and SHR, Ser 188 phosphorylation of RhoA is strongly increased in animals treated with the AT1R inhibitor candesartan (2wks, 2 mg/kg/d). Our results thus indicate that AT2R stimulation induces phosphorylation of RhoA on Ser 188, leading to RhoA inactivation through increased association to cytosolic GDI. In vivo experiments suggest that AT2R-mediated inhibition of RhoA by Ser 188 phosphorylation may contribute to the antihypertensive effect of AT1R inhibitors. American Physiological Society

Making Up and Breaking Up

The concept that blood pressure influences vessel wall composition and structure is well established. Any alterations in stretch (caused by changes in blood pressure) solicit counteracting radial and tangential forces, driving transformations in the vessel wall that aim to accommodate the new conditions and to ultimately restore basal levels of tensile stress.1–3 Thus, when tensile stress increases because of a rise in arterial pressure, smooth muscle cell hypertrophy and collagen and elastin contents augment. Inversely, when the circumferential stress falls, the wall undergoes atrophy.4,5 See page 957 Understanding how vessel wall remodeling occurs is valuable because it relies on the delicate balance between restructuralization and overcompensation. For instance, vascular accumulation of type I, III, and IV collagen in hypertensive patients6 and in animal models of hypertension7,8 effectively counteracts the distending force of blood pressure, but at length it also results in increased vascular stiffness. Clinically, vascular stiffness translates to greater pulse wave velocity, which is an independent predictor of mortality in patients with end-stage renal failure, hypertension, and diabetes, as well as in older individuals.9 In this issue of Atherosclerosis, Thrombosis and Vascular Biology , Jackson et al10 show that the opposite holds true for vessels in which axial tension is off-loaded. In rabbit carotid arteries elongated by interposition of the contralateral carotid artery, the authors found that tortuosity caused by loss of axial strain was not rectified in the months that followed the intervention. Smooth muscle cell proliferation was observed, but this was offset by equivalent cell apoptosis. Likewise, elastin and collagen accumulation was countered by matrix metalloproteinase (MMP) upregulation and activation. But on close observation of the extracellular matrix (ECM), it became apparent that the fenestrae of the internal elastic laminae were greatly enlarged, and indeed the authors found that …

Acute and Chronic Captopril, but Not Prazosin or Nifedipine, Normalize Alterations in Adrenergic Intracellular Ca2+ Handling Observed in the Mesenteric Arterial Tree of Spontaneously Hypertensive Rats

The effect of hypertension and acute (36-h) or chronic (from age 6 to 16 weeks) antihypertensive treatment with prazosin (2 mg kg(-1) per day), nifedipine (50 mg kg(-1) per day), or captopril (50 mg kg(-1) per day) on Ca2+ mobilization due to alpha1-adrenoceptor activation was analyzed in functional studies using arterial rings [four conductance/distributing vessels: aorta, main mesenteric, iliac, and tail arteries and two resistance vessels; first and second small mesenteric artery branches obtained from spontaneously hypertensive rats (SHR, 6 and 16 weeks old) and age-matched Wistar Kyoto rats (WKY)]. Maximal response to noradrenaline in the presence of extracellular Ca2+ is not affected by hypertension or by the antihypertensive treatment. The extracellular Ca2+-independent contractile responses increased with age in iliac, tail, and small mesenteric arteries (SMA) and were further increased in SHR in SMA from both young and adult animals and in the main mesenteric artery of adult SHR. In main mesenteric artery, this increased contraction in SHR was associated with a higher increase in cytosolic [Ca2+] mobilized by noradrenaline without changes in the total stored Ca2+. Acute or chronic treatment with captopril abolished the differences observed between WKY and SHR in the noradrenaline-induced contraction in mesenteric arteries loaded in Ca2+-free medium. In contrast, animals acutely treated with prazosin or chronically treated with either prazosin or nifedipine exhibit the same differences in Ca2+ handling than untreated rats. In conclusion, these differences are not a consequence of increased blood pressure but precede it and can only be normalized by inhibition of the rennin-angiotensin system.