AT1 Receptor Antagonist Losartan Research Articles

Angiotensin II (ANG II) does not utilize the Janus Kinase 2/Signal Transducers of Activated Transcription (JAK/STAT) pathway in the molecular mechanism of blood pressure response to a low salt diet

We recently reported that the JAK2 inhibitor AG490 prevented JAK/STAT pathway activation and hypertension during chronic iv. ANG II infusion. It is unknown, however, whether activation of JAK2 is required for ANG II to prevent blood pressure from decreasing in the physiologic response to low salt intake. This was tested by blocking JAK2 activation chronically in rats that were switched from normal‐ to low‐salt intake. Male SD rats with instrumented with artery and vein catheters for 24 hr/day mean arterial pressure (MAP) measurement and iv. infusions. Rats were maintained on a normal‐salt (NS) intake (~ 3 mEq/day), and after baseline measurements chronic iv. infusions of either: vehicle (Veh; saline), the AT1 receptor antagonist Losartan (LOS; 10 mg/kg/day) or AG490 (AG; 10 ng/kg/min) were begun. After a 4‐day control period, rats were switched to low‐salt intake (LS; 0.4 mEq/day), while Veh, LOS and AG infusions continued. There were no differences in MAP between any group under baseline conditions. During the control period, MAP averaged 93+4, 97+3 and 80+2 mm Hg in Veh, AG, and LOS groups, respectively. Switching to LS intake did not change MAP in Veh and AG rats (102+4 and 101+3 mm Hg, respectively); but MAP decreased significantly in LOS rats to 72+4 mm Hg by day 5. These data suggest that JAK2 activation is not involved in ANG II's role in the blood pressure response to LS intake. (HL74167 MWB; HL91177 AB)

Angiotensin II inhibits GABAergic synaptic transmission in dorsolateral periaqueductal gray neurons

The purpose of this study was to determine the role of angiotensin II (Ang II) in modulating inhibitory and excitatory synaptic inputs to the dorsolateral periaqueductal gray (dl-PAG). The whole cell voltage-clamp recording was performed to examine inhibitory and excitatory postsynaptic currents (IPSCs and EPSCs) of the dl-PAG neurons. Ang II, at the concentration of 2 μM, decreased the frequency of miniature IPSCs from 0.83 ± 0.02 to 0.45 ± 0.03 Hz ( P < 0.05) in 10 tested neurons. This did not significantly affect the amplitude and decay time constant. The effect of Ang II on miniature IPSCs was blocked by the prior application of Ang II AT1 receptor antagonist losartan, but not by AT2 receptor antagonist PD123319. Additionally, Ang II decreased the amplitude of evoked IPSCs from 148 ± 15 to 89 ± 7 pA ( P < 0.05), and increased the paired-pulse ratio from 96 ± 5% to 125 ± 7% ( P < 0.05) in eight tested neurons. In contrast, Ang II had no distinct effects on the EPSCs. Our data suggest that Ang II inhibits GABAergic synaptic inputs to the dl-PAG through activation of presynaptic AT1 receptors.

Angiotensin-(1–7) activates growth-stimulatory pathways in human mesangial cells

Angiotensin-(1-7) [Ang-(1-7)] is generated in part via ACE2-dependent degradation of angiotensin II (ANG II). In proximal tubular cells, Ang-(1-7) inhibits ANG II-stimulated phosphorylation of the mitogen-activated protein kinases (MAPKs) p38, extracellular signal-related kinase (ERK1/ERK2), and c-jun N-terminal kinase (JNK), suggesting that Ang-(1-7) protects against ANG II-mediated tubulointerstitial injury. We determined the effect of Ang-(1-7) on signaling and growth responses in cultured human mesangial cells. Ang-(1-7) increased phosphorylation of p38, ERK1/ERK2, and JNK MAPKs, which was blocked by the Ang-(1-7) antagonist A-779. Neither the AT(1) receptor antagonist losartan, nor the AT(2) antagonist PD123319 affected specific binding of [(125)I]Ang-(1-7) or Ang-(1-7)-stimulated p38 phosphorylation. Ang-(1-7) increased cell arachidonic acid release, an effect blocked by A-779. The p38 MAPK antagonist SB202190 completely prevented Ang-(1-7)-stimulated release of arachidonic acid, whereas inhibitors of ERK or JNK had no effect. Ang-(1-7) significantly enhanced DNA synthesis and increased production of transforming growth factor-beta1 (TGF-beta1), fibronectin, and collagen IV. Both A-779 and SB202190 blocked the Ang-(1-7)-stimulated increases in TGF-beta1, fibronectin, and collagen IV. These data indicate that Ang-(1-7) activates MAPK phosphorylation via binding to a specific receptor in human mesangial cells. Stimulation of p38 MAPK phosphorylation by Ang-(1-7) leads to release of arachidonic acid and production of TGF-beta1 and extracellular matrix proteins. We conclude that Ang-(1-7) exerts growth-stimulatory effects in human mesangial cells.

Letter by Csont and Ferdinandy Regarding Article, “Biglycan Is Required for Adaptive Remodeling After Myocardial Infarction”

To the Editor: We read with great interest the article by Westermann et al1 in which the authors showed that biglycan expression was strongly increased in the myocardium after permanent coronary occlusion in wild-type mice. In the hearts of bgn −/ 0 mice 21 days after coronary occlusion, however, collagen matrix organization was impaired, and several parameters of left ventricular function were deteriorated. Furthermore, these animals showed increased mortality rates after infarction. The authors conclude that the complete absence of biglycan is deleterious and prevents the establishment of a stable infarct scar and hemodynamic adaptation, suggesting that biglycan is required for proper infarct healing. Although the observations of the authors are most valuable, the major conclusion of the authors1 is …

The role of nuclear factor-κB in the effect of angiotensin II in the paraventricular nucleus in protecting the gastric mucosa from ischemia-reperfusion injury in rats

The purpose of this study was to elucidate the role of nuclear factor kappaB (NF-kappaB) in the development of gastric ischemia-reperfusion (GI-R) injury and in mediating the effects of angiotensin II (Ang II) in the paraventricular nucleus (PVN) on GI-R injury. GI-R injury was induced in rats by clamping the celiac artery for 30 min and then reperfusing for 1 h. A cannula was inserted into the unilateral PVN for microinjection of Ang II. The expressions and levels of NF-kappaB (p65), IkappaB-alpha, and phosphorylated IkappaB-alpha in rat gastric mucosa were examined by Western blotting and immunohistochemistry. A laser Doppler flowmeter was used to assess gastric blood flow (GBF). Malondialdehyde (MDA) was determined using the thiobarbituric acid (TBA) method, and superoxide dismutase (SOD) activity was determined by the xanthine/xanthine oxidase method. Microinjection of Ang II (3, 30, and 300 ng) into the PVN dose-dependently inhibited GI-R injury. The levels and expressions of NF-kappaB (p65) and phosphospecific IkappaB-alpha protein increased 1 h after GI-R and were markedly reduced by microinjection of Ang II into the PVN. In contrast, the level and expression of IkappaB-alpha protein decreased 1 h after ischemia-reperfusion and recovered to the normal level by microinjection of Ang II into the PVN. The effects of Ang II were prevented by pretreatment with the Ang II AT1 receptor antagonist losartan (5 microg) microinjected into the lateral cerebral ventricle. Inhibition of NF-kappaB activity by pyrrolidine dithiocarbamate (PDTC, 200 mg/kg) produced similar effects in rats subjected to ischemia-reperfusion with or without microinjection of Ang II into the PVN. Administration of PDTC attenuated gastric mucosal injury and suppressed the activation of NF-kappaB (p65). Ang II microinjection into the PVN increased GBF and decreased the MDA content but did not alter SOD activity in the gastric mucosa following ischemia-reperfusion. NF-kappaB plays a role in PVN Ang II-mediated protection against GI-R injury. These central effects of Ang II are mediated by AT1 receptors.

Administration of angiotensin II in the paraventricular nucleus protects gastric mucosa from ischemia-reperfusion injury

Our previous study demonstrated that electrical stimulation of the hypothalamic paraventricular nucleus (PVN) protects against gastric ischemia-reperfusion (GI-R) injury, but it is still unknown whether angiotensin II (Ang II) in the PVN plays a role in the development of GI-R. The purpose of this study was to investigate the effect of Ang II in the PVN on GI-R injury. GI-R injury was induced in rats by clamping the celiac artery for 30 min, and then reperfusing for 30 min, 1 h, 3 h, 6 h or 24 h, respectively. A cannula was inserted into the unilateral PVN for microinjection of Ang II. The extent of gastric mucosal damage was determined by gross and histological methods. We found that microinjection of pharmacological doses of Ang II (3, 30, and 300 ng) into the PVN dose-dependently inhibited GI-R injury, and that Ang II (30 ng) markedly attenuated GI-R injury at 1 h and 3 h after reperfusion. The effect of Ang II was prevented by pretreatment with the Ang II AT1 receptor antagonist losartan (5 µg) into the lateral cerebral ventricle. Furthermore, the protective effect of Ang II on GI-R injury was abolished by propranolol (1 mg/kg, i.v.) or disconnection of the nerves innervating the adrenal glands, was augmented by sympathectomy or phentolamine (1 mg/kg, i.v.), and was not affected by subdiaphragmatic vagotomy or atropine (1 mg/kg, i.v.). These results indicate that the PVN is a responsive site for central Ang II-induced protection against GI-R injury. The central effects of Ang II are mediated by AT1 receptors in the PVN, and the peripheral effects by a sympathetic-adrenal gland/β-adrenoceptor pathway.

Pathophysiological role of intrarenal Angiotensin II (AngII) in Dahl salt‐sensitive (SS) hypertensive rats

Experiments investigated the role of intrarenal AngII in normotensive and salt‐sensitive hypertensive rats. Increasing dietary NaCl from 0.4% to 4.0% in normal Sprague Dawley rats (n=4–7/group) significantly suppressed plasma renin concentration (5.3±1.3 to 2.9±0.7 ng AngI/ml/hr), plasma AngII (11.7±2.2 to 6.2±0.8 pg/ml), and renal tissue AngII (84±12 to 37±6 pg/gram tissue). The ratio of kidney to plasma AngII was unaltered from the 0.4% NaCl value of 7.2±1.0 when sodium intake was increased. In contrast, increasing dietary NaCl from 0.4 to 4.0% in Dahl SS significantly decreased kidney renin concentration (from 1343±247 to 527±142 ng AngI/μg protein/hr) but the ratio of kidney to plasma AngII significantly increased from 5.9±1.1 to 10.8±2.9. Experiments were then performed to evaluate the functional role of the elevated intrarenal AngII in Dahl SS rats fed high salt. Compared to vehicle‐treated SS rats fed 4.0% NaCl, administration of the AT1 receptor antagonist losartan (30 mg/kg/day, oral) to Dahl SS rats led to a significant attenuation of salt‐sensitive hypertension from 163±6 to 137±1 mmHg and albuminuria from 62±12 to 30±5 mg/day. Administration of captopril (15 mg/kg/day, oral) had similar effects on hypertension and albuminuria in Dahl SS fed high salt. These studies indicate that inappropriately elevated intrarenal AngII may participate in the development of Dahl SS hypertension.

AT1, NMDA, and GABAA receptors in the median preoptic nucleus mediate greater vascular responses in Brown‐Norway/Mcw rats compared to Dahl salt‐sensitive/Mcw rats

We have shown that GABAA, NMDA, and AT1 receptors in the hypothalamic median preoptic nucleus (MnPO) contribute to hemodynamic response variability to cocaine in Sprague‐Dawley rats. These receptors may also contribute to differences in response pattern in two unrelated strains of rats. We named Dahl salt‐sensitive/Mcw (DS) rats cardiac responders since they exhibit an increase in arterial pressure (AP) following cocaine that was due entirely to an increase in cardiac output (CO). We called Brown‐Norway/Mcw (BN) rats vascular responders since they exhibit a similar, sustained increase in AP following cocaine that was due entirely to an increase in systemic vascular resistance (SVR). Microinjection of either the GABAA agonist muscimol (80 pmol/100 nl) or the NMDA receptor antagonist MK‐801 (dizocilpine, 20 nmol) into the MnPO attenuated the decrease in CO and the increase in SVR in BN after cocaine without affecting the responses in DS. Injection of the AT1 receptor antagonist losartan (20 ng) into the MnPO prevented hemodynamic response differences to cocaine in DS and BN. Microinjection of l‐NAME (4 nmol) into the MnPO did not alter responses to cocaine in DS or BN. These results suggest that MnPO GABAA, NMDA, and AT1 receptors participate in producing the vascular response to cocaine in BN. Supported by USPHS DA017371, GM008306, and a predoctoral fellowship from the American Heart Association, Heartland Affiliate.

Angiotensin‐(1–7) has a dual role on growth‐promoting signalling pathways in rat heart in vivo by stimulating STAT3 and STAT5a/b phosphorylation and inhibiting angiotensin II‐stimulated ERK1/2 and Rho kinase activity

Angiotensin (ANG) II contributes to cardiac remodelling by inducing the activation of several signalling molecules, including ERK1/2, Rho kinase and members of the STAT family of proteins. Angiotensin‐(1–7) is produced in the heart and inhibits the proliferative actions of ANG II, although the mechanisms of this inhibition are poorly understood. Accordingly, in the present study we examined whether ANG‐(1–7) affects the ANG II‐mediated activation of ERK1/2 and Rho kinase, STAT3 and STAT5a/b in rat heart in vivo. We hypothesized that ANG‐(1–7) inhibits these growth‐promoting pathways, counterbalancing the trophic action of ANG II. Solutions of normal saline (0.9% NaCl) containing ANG II (8 pmol kg−1) plus ANG‐(1–7) in increasing doses (from 0.08 to 800 pmol kg−1) were administered via the inferior vena cava to anaesthetized male Sprague–Dawley rats. After 5 min, hearts were removed and ERK1/2, Rho kinase, STAT3 and STAT5a/b phosphorylation was determined by Western blotting using phosphospecific antibodies. Angiotensin II stimulated ERK1/2 and Rho kinase phosphorylation (2.3 ± 0.2‐ and 2.1 ± 0.2‐fold increase over basal values, respectively), while ANG‐(1–7) was without effect. The ANG II‐mediated phosphorylation of ERK1/2 and Rho kinase was prevented in a dose‐dependent manner by ANG‐(1–7) and disappeared in the presence of the Mas receptor antagonist d‐Ala7‐ANG‐(1–7). Both ANG II and ANG‐(1–7) increased STAT3 and STAT5a/b phosphorylation to a similar extent (130–140% increase). The ANG‐(1–7)‐stimulated STAT phosphorylation was blocked by the AT1 receptor antagonist losartan and not by d‐Ala7‐ANG‐(1–7). Our results show a dual action of ANG‐(1–7), that is, a stimulatory effect on STAT3 and 5a/b phosphorylation through AT1 receptors and a blocking action on ANG II‐stimulated ERK1/2 and Rho kinase phosphorylation through Mas receptor activation. The latter effect could be representative of a mechanism for a protective role of ANG‐(1–7) in the heart by counteracting the effects of locally generated ANG II.

Angiotensin II protects fibroblast-like synoviocytes from apoptosis via the AT1-NF- B pathway

To evaluate the effects of angiotensin II (Ang II) treatment on apoptosis of fibroblast-like synoviocytes (FLS) from patients with osteoarthritis (OA) and rheumatoid arthritis (RA). AT1 receptor expression was detected by western blotting and flow cytometry. Apoptosis induction was quantified by nucleosome ELISA and by TUNEL; cell proliferation was determined by a bromodeoxyuridine (BrdU) incorporation assay. Silencing of p65 NF-kappaB was obtained by using a specific siRNA. Caspase 3 activation was evaluated by a colorimetric assay and its cleavage by western blotting. AT1 expression resulted comparable in FLS from OA and RA patients. Ang II pre-treatment reduced FLS apoptotic response to serum starvation and nitric oxide (NO) exposure. This protective effect was reverted in the presence of the AT1 receptor antagonist losartan as well as after silencing the expression of NF-kappaB. Moreover, FLS treatment with the caspase inhibitor z-VAD-fmk cancelled this Ang II effect on apoptosis. Caspase 3 activation was reduced in presence of Ang II. Ang II could represent an important mediator involved in FLS expansion, reducing their capacity to undergo apoptosis, through the activation of NF-kappaB and the blockage of caspase cascade.

Modulation of a Delayed-Rectifier K+ Current by Angiotensin II in Rat Sympathetic Neurons

It is well known that angiotensin II (Angio II) mimics most of the muscarinic-mediated excitatory actions of acetylcholine on superior cervical ganglion neurons. For instance, in addition to depolarization and stimulation of norepinephrine release, muscarinic agonists and Angio II modulate the M-type K(+) current and the N-type Ca(2+) current. We recently found that muscarinic receptors modulate the delayed rectifier current I(KV) as well. Therefore a whole cell patch-clamp experiment was carried out in rat cultured sympathetic neurons to assess whether Angio II modulates I(KV). We found that Angio II increased I(KV) by about 30% with a time constant of approximately 30 s. In comparison, inhibition of M-current was faster (tau approximately 8 s) and stronger ( approximately 61%). Modulation of I(KV) was disrupted by the AT(1) receptor-antagonist losartan but not by the AT(2)-antagonist PD123319. I(KV) enhancement was reduced by the G-protein inhibitor GDP-beta-S, whereas current modulation remained unaltered after cell treatment with pertussis toxin. The peptidergic modulation of I(KV) was severely disrupted when internal ATP was replaced by its nonhydrolyzable analogue AMP-PNP. Angio II enhanced I(KV) and further reduced the stimulatory action of a muscarinic agonist on I(KV). Likewise, the muscarinc agonist enhanced I(KV) and occluded the effect of Angio II on I(KV). We have also found that the protein kinase C activator PMA enhanced I(KV), thereby mimicking and further attenuating the action of Angio II on I(KV). These results suggest that AT(1) receptors by coupling to pertussis toxin-insensitive G proteins, stimulate an ATP-dependent and PKC-mediated pathway to modulate I(KV).

Intracerebroventricular injection of losartan inhibits angiotensin II-sensitive neurons via GABA inputs in the anterior hypothalamic area of rats

Previously, we have demonstrated that pressure-ejected application of angiotensin II and losartan, an angiotensin AT1 receptor antagonist, onto some neurons in the anterior hypothalamic area (AHA) of the rat increases and decreases, respectively, the basal firing rate of the neurons. To investigate possible participation of these AHA neurons in the brain angiotensin system, we examined whether intracerebroventricular injection of the angiotensin AT1 receptor antagonist losartan inhibits the neuronal activity of angiotensin II-sensitive neurons via GABA inputs in the AHA of rats. Intracerebroventricular injection of losartan decreased the firing rate of AHA angiotensin II-sensitive neurons. However, the intracerebroventricular injection of losartan did not affect the increase in firing rate of AHA angiotensin II-sensitive neurons induced by pressure application of angiotensin II onto the same neurons, although losartan similarly injected abolished the increase in firing rate of AHA angiotensin II-sensitive neurons induced by intracerebroventricular injection of angiotensin II. The losartan-induced decrease of unit firing in AHA neurons was abolished by pressure application of the GABAA receptor antagonist bicuculline onto the same neurons. Bicuculline itself did not affect the basal firing rate of AHA neurons. These findings suggest that intracerebroventricular injection of losartan inhibits AHA angiotensin II-sensitive neurons via GABA inputs to the neurons.

Angiotensin II type 1 receptor signaling contributes to platelet-leukocyte-endothelial cell interactions in the cerebral microvasculature

Angiotensin II type 1 (AT(1)) receptor signaling has been implicated in cerebral microvascular alterations associated with ischemia, diabetes mellitus, hypercholesterolemia, and atherosclerosis. Platelets, which express AT(1) receptors, also appear to contribute to the thrombogenic and inflammatory responses that are elicited by these pathological conditions. This study assesses the role of AT(1) receptor activation on platelet-leukocyte-endothelial cell interactions elicited in cerebral microvasculature by ischemia and reperfusion. Intravital microscopy was used to monitor the adhesion of platelets and leukocytes that were labeled with different fluorochromes, whereas dihydrorhodamine-123 was used to quantify oxygen radical production in cerebral surface of mice that were either treated with the AT(1) receptor agonist Val-angiotensin II (ANG II) or subjected to bilateral common carotid artery occlusion (BCCAO) followed by reperfusion. ANG II elicited a dose- and time- dependent increase in platelet-leukocyte-endothelial cell interactions in cerebral venules that included rolling platelets, adherent platelets on the leukocytes and the endothelial cells, rolling leukocytes, and adherent leukocytes. All of these interactions were attenuated by treatment with either P-selectin or P-selectin glycoprotein ligand 1 (PSGL-1) antibody. The AT(1) receptor antagonist candesartan and losartan as well as diphenyleneiodonium, an inhibitor of flavoproteins including NAD(P)H oxidase, significantly reduced the platelet-leukocyte-endothelial cell interactions elicited by either ANG II administration or BCCAO/reperfusion. The increased oxygen radical generation elicited by BCCAO/reperfusion was also attenuated by candesartan. These findings are consistent with an AT(1) receptor signaling mechanism, which involves oxygen radical production and ultimately results in P-selectin- and PSGL-1-mediated platelet-leukocyte-endothelial cell interactions in the cerebral microcirculation.

Stimulation of cerebral and dermal microvascular endothelial cell proliferation by AT1 receptor antagonist losartan

Aim: We have shown that cerebral angiogenesis is stimulated by chronic treatment of rats with the angiotensin II (ANG II) type 1 receptor (AT1R) antagonist, losartan (LOS). Because the trophic actions of ANG II via AT1R are well known, the finding that an AT1R antagonist increases angiogenesis is paradoxical. The aim is to determine whether cerebral (CMVEC) and dermal (DMVEC) microvascular endothelial cells differ in their growth responses to LOS treatment. Methods: Primary human CMVEC and DMVEC, passage 3–8, were seeded into 96-well plates (5000 cells/well) and allowed to adhere for 24 hrs. Cells were exposed to serum-free media (SFM) alone and in the presence of LOS (10 μM), ANG II (1 μM) or both for 48 hours. Cell density was quantified by a fluorescent dye proliferation assay at 480/520 nm. Results: Density of CMVEC increased with either ANG II stimulation (27.5%) or treatment with LOS, with (33.7%) or without ANG II (27.1%) compared to SFM alone. A similar but less robust pattern was seen with DMVEC with ANG II (8.9%), LOS + ANG II (15.2%) and LOS alone (19.0%). Conclusion: This suggests that CMVEC and DMVEC have similar proliferative responses so they do not appear to be tissue-specific. However, LOS alone has a similar effect to treatment with both the agonist and antagonist suggesting that LOS directly stimulates cell proliferation and that the effect is not due to receptor blockade per se. Supported by NIH HL29587 and HV28182

Cardiac sympathetic afferent stimulation inhibits the arterial baroreflex in heart failure at the level of the nucleus tractus solitarius by an angiotensin II mechanism

The enhanced cardiac sympathetic afferent reflex (CSAR) has been suggested to contribute to increased sympathetic outflow and blunted baroreflex via a central angiotensin II (Ang II) mechanism in chronic heart failure (CHF). This work was designed to determine the Ang II mechanism in the nucleus tractus solitarius (NTS) underlying the interaction between the baroreflex and the CSAR using extracellular recording in rats with CHF. CHF was produced by coronary artery ligation. Of 38 NTS cells tested for barosensitivity by elevation of arterial pressure, 28 were recorded in CHF rats and 10 were tested in sham rats. CSAR activation by epicardial application of capsaicin (0.4 μg) attenuated the barosensitivity of 12 NTS neurons by 86.2 ± 9.3% and this was significantly greater in CHF rats than in sham rats (65.7 ± 6.3%). In 8 cells from CHF rats, tonic inhibition of CSAR by epicardial lidocaine (6 μl) improved the barosensitivity of NTS neurons by 42.8 ± 5.9 %. In the remaining 8 cells from CHF rats, intravenous injection of the Ang II AT1 receptor antagonist losartan (2 mg/kg) not only sensitized the barosensitivity by 83.1 ± 7.2% but also reversed the inhibitory effect of CSAR activation on barosensitivity (−84.7 ± 9.6 vs. −12.5 ± 4.8 %). In conclusion, these data suggest that the NTS plays an important role in mediating the blunting of baroreflex by CSAR activation via an AT1 receptor dependent mechanism in CHF.

Selective AT1 Receptor Antagonism Enhances Sympathetically Mediated Vasoconstriction in Man

The vasoconstrictive action of angiotensin II (AII) is partly, sympathetically mediated and angiotensin-converting enzyme (ACE) inhibitors appear to exert a sympatholytic effect. We examine the effect of an orally administered, selective AT(1) receptor antagonist (losartan 50 mg) on sympathetically mediated vasoconstriction in healthy volunteers in an observer blind crossover study. Seven healthy, normotensive volunteers (21-32 years), were studied on two occasions at the end of each 6-week treatment period (losartan or placebo). Forearm blood flow (FABF) (ml/dl forearm/min) was measured by venous occlusion plethysmography during the application of lower body negative pressure (LBNP) (-20 cm H(2)O) and at the end of each incremental infusion of norepinephrine (60, 120, and 240 pmol/min). Comparison of blood flow changes was by repeated measures analysis of variance; P<0.05 was taken as statistically significant. Losartan did not alter blood pressure compared to placebo. It did significantly enhance LBNP-induced vasoconstriction in both the left arm compared to placebo (-36.6+/-3.4 vs -23.5+/-3.3%; P=0.017) and the right arm compared to placebo (-39.5+/-3.8 vs -21.0+/-3.6%; P=0.005). The FABF response to all doses of infused norepinephrine (60, 120, and 240 pmol/min) was also enhanced by losartan compared to placebo (-35.0+/-2.7 vs -18.2+/-6.0%; -43.6+/-4.3 vs -28.6+/-5.8%, and -53.9+/-3.2 vs -42.5+/-6.8%; P=0.057, respectively. Losartan enhances locally mediated sympathetic vasoconstriction in the forearm circulation of man, probably through its effect on circulating AII concentrations and we postulate that the adrenergic sympathetic constrictor action of AII is not mediated by the AT(1) receptor or is surmountable at this receptor.

Angiotensin II activates two cation conductances with distinct TRPC1 and TRPC6 channel properties in rabbit mesenteric artery myocytes.

Angiotensin II (Ang II) is a potent vasoconstrictor with an important role in controlling blood pressure; however, there is little information on cellular mechanisms underlying Ang II-evoked vasoconstrictor responses. The aim of the present study is to investigate the effect of Ang II on cation conductances in freshly dispersed rabbit mesenteric artery myocytes at the single-channel level using patch-clamp techniques. In cell-attached patches, bath application of low concentrations of Ang II (1 nM) activated cation channel currents (Icat1) with conductances states of about 15, 30 and 45 pS. At relatively high concentrations, Ang II (100 nM) inhibited Icat1 but evoked another cation channel (Icat2) with a conductance of approximately 2 pS. Ang II-evoked Icat1 and Icat2 were inhibited by the AT1 receptor antagonist losartan and the phospholipase C (PLC) inhibitor U73122. The diacylglycerol (DAG) lipase inhibitor RHC80267 initially induced Icat1 which was subsequently inhibited to reveal Icat2. The DAG analogue 1-oleoyl-2-acetyl-sn-glycerol (1 microM) activated Icat1 and Icat2 but inositol 1,4,5-trisphosphate did not evoke either conductance. The protein kinase C (PKC) inhibitor chelerythrine (3 microM) potentiated Ang II-evoked Icat1 and inhibited Icat2 whereas the PKC activator phorbol-12,13-dibutyrate (1 microM) reduced Ang II-induced Icat1 but activated Icat2. Moreover in cell-attached patches pretreated with chelerythrine, application of 100 nM Ang II activated Icat1. These data indicate that PKC inhibits Icat1 but stimulates Icat2. Agents that deplete intracellular Ca2+ stores also activated cation channel currents with similar properties to Icat2. Bath application of anti-TRPC6 and anti-TRPC1 antibodies to inside-out patches inhibited Icat1 and Icat2, respectively. Also flufenamic acid and zero external Ca2+ concentration, respectively, potentiated and reduced Ang II-evoked Icat1. Immunocytochemical studies showed TRPC6 and TRPC1 expression with TRPC6 preferentially distributed in the plasma membrane and TRPC1 expression located throughout the myocyte. These results indicate that Ang II activates two distinct cation conductances in mesenteric artery myocytes by stimulation of AT1 receptors linked to PLC. Icat1 is activated by DAG via a PKC-independent mechanism whereas Icat2 involves DAG acting via a PKC-dependent pathway. Higher concentrations of Ang II inhibit Icat1 by activating an inhibitory effect of PKC. It is proposed that TRPC6 and TRPC1 channel proteins are important components of Ang II-induced Icat1 and Icat2, respectively.

Centrally injected angiotensin II trans-synaptically activates angiotensin II-sensitive neurons in the anterior hypothalamic area of rats

Previously, we have demonstrated that pressure-ejected application of angiotensin II onto some neurons in the anterior hypothalamic area (AHA) of rats increases their firing rate. In contrast, pressure application of the angiotensin AT1 receptor antagonist losartan onto AHA neurons blocked the basal firing of the neurons. To investigate possible participation of these AHA neurons in the brain angiotensin system, we examined whether intracerebroventricular injection of angiotensin II results in an activation of angiotensin II-sensitive neurons in the AHA of rats. Intracerebroventricular injection of angiotensin II increased the firing rate of AHA angiotensin II-sensitive neurons. The angiotensin II-induced increase of unit firing in AHA neurons was abolished by pressure application of losartan onto the same neurons. In addition, the angiotensin II-induced increase of firing in AHA neurons was abolished by pressure application of N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride (W7), a calmodulin inhibitor, onto the same neurons. Pressure application of W7 onto AHA neurons affected neither the basal firing rate nor the increase in unit firing induced by pressure application of angiotensin II onto the same neurons. Intracerebroventricular injection of the cholinergic agonist carbachol did not affect the firing rate of angiotensin II-sensitive neurons in the AHA. These findings suggest that intracerebroventricular injection of angiotensin II activates AHA angiotensin II-sensitive neurons via angiotensinergic inputs to the neurons.

Blood pressure is the major driving force for plaque formation in aortic-constricted ApoE−/− mice

Using an aortic constriction model in mice, we studied whether the increase in pressure or the activation of the renin-angiotensin system (RAS) and its main receptors is the main driving force for plaque progression. Male ApoE mice underwent sham surgery or placement of a suprarenal silver clip around the aorta (AoC). Half the group was treated with the selective AT1 receptor antagonist losartan (30 mg/kg per day) for 4 weeks. Anesthetized mean arterial pressure (MAP) was increased in AoC mice compared to sham (106 +/- 3 versus 90 +/- 1 mmHg, P < 0.001). Losartan reduced MAP in sham mice (78 +/- 2 mmHg, P < 0.01) but not in AoC (AoC losartan 104 +/- 2 mmHg). Plasma renin concentration (PRC) was increased in AoC mice compared to sham [1.6 +/- 0.3 versus 0.8 +/- 0.2 milliGoldblatt units (mGU)/ml, P < 0.001]. Losartan treatment augmented this difference (18.7 +/- 3.7 versus 4.6 +/- 1.7 mGU/ml, P < 0.01). AT2 receptor mRNA expression was increased 5.8-fold by aortic constriction in thoracic aorta (P < 0.05) and the major site for expression of the AT2 receptor protein was within the plaques. The plaque area was increased in AoC mice compared to sham (0.61 +/- 0.09 versus 0.07 +/- 0.01%, P < 0.001); however, losartan did not alter plaque area. Our data do not support a role for the AT1 receptor in the progression of atherosclerosis in this model, since blockade with losartan did not alter plaque distribution. Furthermore, we found no support for the counteraction of atherogenesis by increased activity of the RAS acting on the AT2 receptor. Our data suggest that increased pressure is the main driving force for atherosclerosis in this model.

A “love triangle” elicited by electrochemistry: complex interactions among cardiac sympathetic afferent, chemo-, and baroreflexes

there are three major cardiovascular reflexes: the baroreflex, chemoreflex, and cardiac sympathetic afferent reflex (CSAR). This commentary provides a brief overview of these three reflexes and also discusses the interplay among them. In addition, it outlines the potential centrally integrative