Angiotensin II type 1 receptor-mediated inhibition of K+ channel subunit kv2.2 in brain stem and hypothalamic neurons.
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Angiotensin II (Ang II) has powerful modulatory actions on cardiovascular function that are mediated by specific receptors located on neurons within the hypothalamus and brain stem. Incubation of neuronal cocultures of rat hypothalamus and brain stem with Ang II elicits an Ang II type 1 (AT1) receptor-mediated inhibition of total outward K+ current that contributes to an increase in neuronal firing rate. However, the exact K+ conductance(s) that is inhibited by Ang II are not established. Pharmacological manipulation of total neuronal outward K+ current revealed a component of K+ current sensitive to quinine, tetraethylammonium, and 4-aminopyridine, with IC50 values of 21.7 micromol/L, 1.49 mmol/L, and 890 micromol/L, respectively, and insensitive to alpha-dendrotoxin (100 to 500 nmol/L), charybdotoxin (100 to 500 nmol/L), and mast cell degranulating peptide (1 micromol/L). Collectively, these data suggest the presence of Kv2.2 and Kv3.1b. Biophysical examination of the quinine-sensitive neuronal K+ current demonstrated a macroscopic conductance with similar biophysical properties to those of Kv2.2 and Kv3.1b. Ang II (100 nmol/L), in the presence of the AT2 receptor blocker PD123,319, elicited an inhibition of neuronal K+ current that was abolished by quinine (50 micromol/L). Reverse transcriptase-polymerase chain reaction analysis confirmed the presence of Kv2.2 and Kv3.1b mRNA in these neurons. However, Western blot analyses demonstrated that only Kv2.2 protein was present. Coexpression of Kv2.2 and the AT1 receptor in Xenopus oocytes demonstrated an Ang II-induced inhibition of Kv2.2 current. Therefore, these data suggest that inhibition of Kv2.2 contributes to the AT1 receptor-mediated reduction of neuronal K+ current and subsequently to the modulation of cardiovascular function. (+info)
Recent progress in angiotensin II type 2 receptor research in the cardiovascular system.
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Angiotensin II (Ang II) plays an important role in regulating cardiovascular hemodynamics and structure. Multiple lines of evidence have suggested the existence of Ang II receptor subtypes, and at least 2 distinct receptor subtypes have been defined on the basis of their differential pharmacological and biochemical properties and designated as type 1 (AT1) and type 2 (AT2) receptors. To date, most of the known effects of Ang II in adult tissues are attributable to the AT1 receptor. Recent cloning of the AT2 receptor contributes to reveal its physiological functions, but many functions of the AT2 receptor are still an enigma. AT1 and AT2 receptors belong to the 7-transmembrane, G protein-coupled receptor family. However, accumulating evidence demonstrates that the function and signaling mechanisms of these receptor subtypes are quite different, and these receptors may exert opposite effects in terms of cell growth and blood pressure regulation. We will review the role of the AT2 receptor in the cardiovascular system and the molecular and cellular mechanisms of AT2 receptor action. (+info)
Intracellular sodium modulates the expression of angiotensin II subtype 2 receptor in PC12W cells.
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Although the angiotensin II subtype 2 receptor (AT2-R) is expressed abundantly in the adrenal medulla, its physiological significance has not yet been determined. To obtain fundamental knowledge of the regulation of AT2-R expression in the adrenal medulla, we investigated the effects of modulating several ion channels on AT2-R expression in PC12W cells. Experiments were performed after 24 hours of serum depletion under subconfluent conditions. After 48 hours of treatment with various agonists or antagonists, the receptor density and mRNA level of AT2-Rs were quantified by 125I-[Sar1, Ile8]angiotensin II binding and Northern blot analysis. Ouabain (10 to 100 nmol/L) and insulin (10 to 100 nmol/L) dose-dependently increased receptor density and mRNA level. Analysis of the binding characteristics revealed that the ouabain-dependent increase in AT2-R levels was due to an increase in binding capacity without a change in the Kd value. These increases were blocked by lowering the Na+ concentration in the medium. A low concentration of the sodium ionophore monensin (10 nmol/L), the K+-channel blocker quinidine (10 micromol/L), and the ATP-sensitive K+-channel blockers tolbutamide (100 micromol/L) and glybenclamide (10 micromol/L) also significantly increased receptor density, but the ATP-sensitive K+-channel agonist cromakalim (100 micromol/L) decreased receptor density significantly (P<0.01). Nifedipine (10 micromol/L) decreased basal receptor density and completely blocked the increase in receptor density caused by these agents. The increase in receptor density caused by an increase in intracellular Na+ was accompanied by an increase in mRNA level, whereas the ATP-sensitive K+-channel blockers did not change mRNA level. Nifedipine slightly decreased mRNA level. These results suggest that AT2-R expression is sensitively regulated by intracellular cation levels. The change in intracellular Na+ level transcriptionally regulates AT2-R expression, whereas the K+-channel blocker-dependent upregulation appears to be at least in part posttranslational. (+info)
Irbesartan reduces QT dispersion in hypertensive individuals.
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Angiotensin type 1 receptor antagonists have direct effects on the autonomic nervous system and myocardium. Because of this, we hypothesized that irbesartan would reduce QT dispersion to a greater degree than amlodipine, a highly selective vasodilator. To test this, we gathered electrocardiographic (ECG) data from a multinational, multicenter, randomized, double-blind parallel group study that compared the antihypertensive efficacy of irbesartan and amlodipine in elderly subjects with mild to moderate hypertension. Subjects were treated for 6 months with either drug. Hydrochlorothiazide and atenolol were added after 12 weeks if blood pressure (BP) remained uncontrolled. ECGs were obtained before randomization and at 6 months. A total of 188 subjects (118 with baseline ECGs) were randomized. We analyzed 104 subjects who had complete ECGs at baseline and after 6 months of treatment. Baseline characteristics between treatments were similar, apart from a slight imbalance in diastolic BP (irbesartan [n=53] versus amlodipine [n=51], 99.2 [SD 3. 6] versus 100.8 [3.8] mm Hg; P=0.03). There were no significant differences in BP normalization (diastolic BP <90 mm Hg) between treatments at 6 months (irbesartan versus amlodipine, 80% versus 88%; P=0.378). We found a significant reduction in QT indexes in the irbesartan group (QTc dispersion mean, -11.4 [34.5] milliseconds, P=0.02; QTc max, -12.8 [35.5] milliseconds, P=0.01), and QTc dispersion did not correlate with the change in BP. The reduction in QT indexes with amlodipine (QTc dispersion, -9.7 [35.4] milliseconds, P=0.06; QTc max, -8.6 [33.2] milliseconds, P=0.07) did not quite reach statistical significance, but there was a correlation between the change in QT indexes and changes in systolic BP. In conclusion, irbesartan improved QT dispersion, and this effect may be important in preventing sudden cardiac death in at-risk hypertensive subjects. (+info)
Role of the angiotensin type 2 receptor gene in congenital anomalies of the kidney and urinary tract, CAKUT, of mice and men.
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Angiotensin type 2 receptor gene null mutant mice display congenital anomalies of the kidney and urinary tract (CAKUT). Various features of mouse CAKUT impressively mimic human CAKUT. Studies of the human type 2 receptor (AGTR2) gene in two independent cohorts found that a significant association exists between CAKUT and a nucleotide transition within the lariat branchpoint motif of intron 1, which perturbs AGTR2 mRNA splicing efficiency. AGTR2, therefore, has a significant ontogenic role for the kidney and urinary tract system. Studies revealed that the establishment of CAKUT is preceded by delayed apoptosis of undifferentiated mesenchymal cells surrounding the urinary tract during key ontogenic events, from the ureteral budding to the expansive growth of the kidney and ureter. (+info)
Angiotensin receptor subtype 1 mediates angiotensin II enhancement of isoproterenol-induced cyclic AMP production in preglomerular microvascular smooth muscle cells.
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In a previous study, we found that angiotensin (Ang) II enhances beta-adrenoceptor-induced cAMP production in cultured preglomerular microvascular smooth muscle cells (PMVSMCs) obtained from spontaneously hypertensive rats. The purpose of the present investigation was to identify the Ang receptor subtypes that mediate this effect. In our first study, we compared the ability of Ang II, Ang III, Ang (3-8), and Ang (1-7) to increase cAMP production in isoproterenol (1 microM)-treated PMVSMCs. Each peptide was tested at 0.1, 1, 10, 100, and 1000 nM. Both Ang II and Ang III increased intracellular (EC50s, 1 and 11 nM, respectively) and extracellular (EC50s, 2 and 14 nM, respectively) cAMP levels in a concentration-dependent fashion. In contrast, Ang (3-8) and Ang (1-7) did not enhance either intracellular or extracellular cAMP levels at any concentration tested. In our second study, we examined the ability of L 158809 [a selective Ang receptor subtype 1 (AT1) receptor antagonist] to inhibit Ang II (100 nM) and Ang III (100 nM) enhancement of isoproterenol (1 microM)-induced cAMP production in PMVSMCs. L 158809 (10 nM) abolished or nearly abolished (p <.001) Ang II and Ang III enhancement of isoproterenol-induced intracellular and extracellular cAMP levels. In contrast, PD 123319 (300 nM; a selective AT2 receptor antagonist) did not significantly alter Ang II enhancement of isoproterenol-induced intracellular or extracellular cAMP levels. We conclude that AT1 receptors, but not AT2, Ang (3-8), nor Ang (1-7) receptors mediate Ang II and Ang III enhancement of beta-adrenoceptor-induced cAMP production in cultured PMVSMCs. (+info)
Angiotensin II increases the release of endothelin-1 from human cultured endothelial cells but does not regulate its circulating levels.
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We investigated the effect of angiotensin II on endothelin-1 secretion in vitro and in vivo. In vivo, angiotensin II was given intravenously to 23 essential hypertensive and 8 control subjects according to different protocols: Study A, 1.0 ng x min-1 x kg-1 and 3.0 ng x min-1 x kg-1 angiotensin II for 30 min each; Study B, 1.0 ng x min-1 x kg-1 and 3.0 ng x min-1 x kg-1 angiotensin II for 120 min each; Study C, 3.0 ng x min-1 x kg-1 angiotensin II for 30 min followed by a dose increment of 3.0 ng x min-1 x kg-1 every 30 min until mean blood pressure levels increased by 25 mmHg; Study D, 1.0 ng x min-1 x kg-1 followed by 3.0 ng x min-1 x kg-1 angiotensin II for 60 min each on two different NaCl diets (either 20 mmol NaCl/day or 220 mmol NaCl/day, both for 1 week). In all in vivo studies neither plasma nor urine endothelin-1 levels changed with angiotensin II infusion. In contrast, angiotensin II (10(-9), 10(-8), 10(-7) mol/l) stimulated endothelin-1 secretion from cultured human vascular endothelial cells derived from umbilical cord veins in a time- and dose-dependent manner. The in vitro angiotensin II effects were abolished by candesartan cilexetil, an inhibitor of the membrane-bound AT1 receptor, and also by actinomycin D, an RNA synthesis inhibitor, and cycloheximide, a protein synthesis inhibitor, indicating that endothelin-1 release depended on AT1 receptor subtype and de novo protein synthesis. Our findings indicate that angiotensin II regulates endothelin-1 release by cultured endothelial cells through an AT1 receptor-dependent pathway, but does not influence circulating endothelin-1 levels in vivo. (+info)
Role of aromaticity of agonist switches of angiotensin II in the activation of the AT1 receptor.
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We have shown previously that the octapeptide angiotensin II (Ang II) activates the AT1 receptor through an induced-fit mechanism (Noda, K., Feng, Y. H., Liu, X. P., Saad, Y., Husain, A., and Karnik, S. S. (1996) Biochemistry 35, 16435-16442). In this activation process, interactions between Tyr4 and Phe8 of Ang II with Asn111 and His256 of the AT1 receptor, respectively, are essential for agonism. Here we show that aromaticity, primarily, and size, secondarily, of the Tyr4 side chain are important in activating the receptor. Activation analysis of AT1 receptor position 111 mutants by various Ang II position 4 analogues suggests that an amino-aromatic bonding interaction operates between the residue Asn111 of the AT1 receptor and Tyr4 of Ang II. Degree and potency of AT1 receptor activation by Ang II can be recreated by a reciprocal exchange of aromatic and amide groups between positions 4 and 111 of Ang II and the AT1 receptor, respectively. In several other bonding combinations, set up between Ang II position 4 analogues and receptor mutants, the gain of affinity is not accompanied by gain of function. Activation analysis of position 256 receptor mutants by Ang II position 8 analogues suggests that aromaticity of Phe8 and His256 side chains is crucial for receptor activation; however, a stacked rather than an amino-aromatic interaction appears to operate at this switch locus. Interaction between these residues, unlike the Tyr4:Asn111 interaction, plays an insignificant role in ligand docking. (+info)