AT1 Type Research Articles

Phosphorylation of Tyrosine 319 of the Angiotensin II Type 1 Receptor Mediates Angiotensin II-induced Trans-activation of the Epidermal Growth Factor Receptor

Although tyrosine kinases are critically involved in the angiotensin II (Ang II) type 1 (AT1) receptor signaling, how AT1 receptors activate tyrosine kinases is not fully understood. We examined the structural requirements of the AT1 receptor for transactivation of the epidermal growth factor (EGF) receptor (EGFR). Studies using carboxyl terminal-truncated AT1 receptors indicated that the amino acid sequence between 312 and 337 is required for activation of EGFR. The role of the conserved YIPP motif in this sequence in transactivation of EGFR was investigated by mutating tyrosine 319. Ang II failed to activate EGFR in cells expressing AT1-Y319F, whereas EGFR was activated even without Ang II in cells expressing AT1-Y319E, which mimics the AT1 receptor phosphorylated at Tyr-319. Immunoblot analyses using anti-phospho Tyr-319-specific antibody showed that Ang II increased phosphorylation of Tyr-319. EGFR interacted with the AT1 receptor but not with AT1-Y319F in response to Ang II stimulation, whereas the EGFR-AT1 receptor interaction was inhibited in the presence of dominant negative SHP-2. The requirement of Tyr-319 seems specific for EGFR because Ang II-induced activation of other tyrosine kinases, including Src and JAK2, was preserved in cells expressing AT1-Y319F. Extracellular signal-regulated kinase activation was also maintained in AT1-Y319F through activation of Src. Overexpression of wild type AT1 receptor in cardiac fibroblasts enhanced Ang II-induced proliferation. By contrast, overexpression of AT1-Y319F failed to enhance cell proliferation. In summary, Tyr-319 of the AT1 receptor is phosphorylated in response to Ang II and plays a key role in mediating Ang II-induced transactivation of EGFR and cell proliferation, possibly through its interaction with SHP-2 and EGFR.

Peripheral and central interactions between the renin-angiotensin system and the renal sympathetic nerves in control of renal function.

Increases in renal sympathetic nerve activity (RSNA) regulate the functions of the nephron, the vasculature, and the renin-containing juxtaglomerular granular cells. As increased activity of the renin-angiotensin system can also influence nephron and vascular function, it is important to understand the interactions between RSNA and the renin-angiotensin system in the control of renal function. These interactions can be intrarenal, that is, the direct (via specific innervation) and indirect (via angiotensin II) contributions of increased RSNA to the regulation of renal function. The effects of increased RSNA on renal function are attenuated when the activity of the renin-angiotensin system is suppressed or antagonized with angiotensin-converting enzyme inhibitors or angiotensin II-type AT1 receptor antagonists. The effects of intrarenal administration of angiotensin II are attenuated following renal denervation. These interactions can also be extrarenal, that is, in the central nervous system, wherein RSNA and its arterial baroreflex control are modulated by changes in activity of the renin-angiotensin system. In addition to the circumventricular organs, the permeable blood-brain barrier of which permits interactions with circulating angiotensin II, there are interactions at sites behind the blood-brain barrier that depend on the influence of local angiotensin II. The responses to central administration of angiotensin II type AT1 receptor antagonists, into the ventricular system or microinjected into the rostral ventrolateral medulla, are modulated by changes in activity of the renin-angiotensin system produced by physiological changes in dietary sodium intake. Similar modulation is observed in pathophysiological models wherein activity of both the renin-angiotensin and sympathetic nervous systems is increased (e.g., congestive heart failure). Thus, both renal and extrarenal sites of interaction between the renin-angiotensin system and RSNA are involved in influencing the neural control of renal function.

Angiotensin II AT(1) receptor antagonists and platelet activation.

Multiple factors are involved in thrombus formation and require complex and highly therapeutic strategies. Platelet activation plays a critical role in the genesis of acute coronary syndromes involving not only platelets but also endothelial cells, leucocytes and erythrocytes. Angiotensin II (Ang II) is a vasoconstrictor that could participate in the thrombotic process. Platelets also express Ang II AT1 type receptors on their surface. Losartan is a non-peptidic inhibitor of AT1 receptors. It has been demonstrated that losartan reduced platelet aggregation induced by the thromboxane A2 (TXA2) analogue U46619. This effect was not observed with the losartan metabolite EXP 3174. The effect of losartan was assessed in binding studies in which losartan competitively inhibited the binding of [3H]U46619 to platelets in a dose-dependent manner. Irbesartan also inhibits the TXA2 receptor in platelets, an effect that was not obtained with the active form of candesartan, CV11974, and with valsartan. These results suggest that the structural requirements necessary to antagonize the TXA2/PGH2 platelet receptor may be different from those involved in AT1 receptor antagonism. The in vivo relevance of the in vitro findings has been confirmed by the fact that in vivo administration of losartan decreases P-selectin expression in platelets obtained from stroke-prone spontaneously hypertensive rats.

The Renin‐Angiotensin System in the Bovine Utero‐Placental Unit

ContentsRenin, angiotensin converting enzyme, and angiotensin II receptors are expressed in the bovine utero‐placental unit, indicating the presence of a local renin‐angiotensin system (RAS). Angiotensin II receptors of type AT1 and AT2 as well as non‐AT1/non‐AT2 binding sites were identified. The expression of both renin and the angiotensin II receptor types in the bovine utero‐placental unit differs from that in humans. This fact is probably related to differences in placental architecture. The RAS in the bovine utero‐placental unit might be important for growth processes in the endometrium and for placentation. It might also participate in the regulation of the utero‐placental blood flow and secretion of hormones. Disturbances in the utero‐placental RAS might cause a reduced utero‐placental blood flow, intrauterine growth retarda_tion and intrauterine death.

Efecto del losartán sobre la activación de plaquetas humanas por tromboxano A2

Previous studies have demonstrated that losartan, an AT-1 receptor antagonist of angiotensin II (Ang II) could block the receptor of thromboxane A2 (TXA2) in the vascular wall. The aim of the present study was to assess the effect of losartan on human platelet activation. Platelets were obtained from 15 healthy men between the age 26 and 40. Platelet activation was measured by changes in the light transmission of platelet-rich plasma stimulated by a synthetic TXA2 analogue, U46619 (5 x 10(-6) mol/l). The U46619-stimulated platelet aggregation was significantly inhibited by losartan in a dose-response manner. Only a high dose of EXP 3174 (5 10-5 mol/l), the in vivo active metabolite of losartan, was able to attenuate U46619-induced platelet activation. Captopril, an angiotensin I-converting inhibitor failed to modify U46619-induced platelet aggregation. Despite the platelets expressing AT-1 type receptors, of Ang II exogenous Ang II did not modify platelet aggregation induced by U46619. The binding of U46619 to platelets was competitively inhibited by losartan in dose-dependent manner. However, only a high dose of EXP 3174 reduced the binding of U46619. Captopril failed to modify the binding of U46619 to platelets. Losartan decreased platelet aggregation by a TXA2-dependent mechanism. EXP 3174 showed a lesser potency than losartan to reduce TXA2-platelet activation. Captopril and exogenous angiotensin II had no effect on human platelet activation. These results suggest that losartan reduced TXA2-dependent platelet activation independently of the blockade of AT-1 receptors.

Candesartan cilexetil: an angiotensin II receptor blocker.

To summarize and critique the medical literature on candesartan cilexetil, an angiotensin II receptor blocker (ARB). MEDLINE searches (January 1966-January 1999) and manufacturer prescribing literature were used to identify articles on candesartan cilexetil. Bibliographies were also reviewed for germane articles. Study and review articles describing the chemistry, human pharmacology, pharmacodynamics, pharmacokinetics, placebo-controlled trials, comparative trials, and clinical application of candesartan cilexetil based on the published literature and premarketing clinical trials were reviewed. All literature on the use of candesartan cilexetil for treating hypertension and congestive heart failure were included. ARBs are a new class of drugs with increasing use in treating hypertension. Studies are ongoing to determine the role of these agents in preventing remodeling after myocardial infarction and in patients with congestive heart failure. Candesartan cilexetil is among the newest drugs in the class that includes losartan, irbesartan, and valsartan. Candesartan cilexetil has more than 1000 times more affinity for the angiotensin II, type AT1 receptor ARBs, and the binding affinity and competitive angiotensin II receptor antagonism is stronger than that of losartan. Clinical studies in patients with hypertension have demonstrated that candesartan cilexetil, in doses of 4-16 mg, is more effective in reducing sitting diastolic blood pressure than are placebo and losartan 50 mg. Candesartan cilexetil has demonstrated reductions in blood pressure comparable to those of enalapril, with the rate of adverse events greater in the enalapril group. Dosage adjustments are not necessary in elderly patients or in patients with mild hepatic or renal dysfunction. In diabetic patients, blood glucose, hemoglobinA1c, and serum lipids are not affected. The clinical studies demonstrated that the adverse effect profile of candesartan cilexetil was similar to that of placebo and there were no dose-dependent adverse effects. Candesartan cilexetil provides an alternative antihypertensive therapy that is well tolerated and effective in reducing blood pressure in a wide range of patients. Due to its greater binding affinity to the angiotensin II receptor, candesartan cilexetil appears to have a longer antihypertensive effect than losartan. This may be advantageous in decreasing morbidity and mortality associated with hypertension, although further studies are required to validate this potential advantage.

Endogenous angiotensin II produced by endothelium regulates interleukin-1beta-stimulated nitric oxide generation in rat isolated vessels.

The endothelium is a source of several factors that regulate vascular functions. Angiotensin II is one of the main active factors released by the endothelium. The aim of the present work was to analyze the role of angiotensin II released by the endothelium in the regulation of the inducible nitric oxide synthase expression in rat isolated aortic vessels. Interleukin-1beta (0.03 U/L) stimulated nitrite release by the aortic vessels. The nitrite released was less in vessels with endothelium than in deendothelialized aortic segments. This effect was accompanied by a reduced expression of the inducible nitric oxide synthase in the aortic rings with endothelium. Exogenous angiotensin II inhibited IL-1beta-stimulated inducible nitric oxide synthase protein expression in both deendothelialized vessels and those with endothelium, although with reduced ability on the aortic segments with endothelium by a nitric oxide-independent mechanism. In the aortic rings with endothelium, either inhibition of the AT-1 receptor with losartan or blocking of angiotensin II generation with fosinopril enhanced interleukin-1beta-stimulated inducible nitric oxide synthase protein expression. In conclusion, the endothelium decreases inducible nitric oxide synthase expression in the vascular wall. Angiotensin II released from endothelial cells is a main mediator responsible for this inhibition through an AT-1-type receptor-dependent mechanism.

Intracerebroventricular losartan inhibits postprandial drinking in sheep.

We investigated the contribution of brain angiotensinergic mechanisms to postprandial drinking in sheep. Sheep in fluid balance were given 0.8 kg chaff for 30 min, and water intake was measured for the next hour. Intracerebroventricular infusion of the AT1 type angiotensin II (ANG II) receptor blocker losartan (1 mg/h) reduced postprandial drinking by approximately 70% (n = 7, P < 0.01) but did not affect food intake. The same losartan dose given intravenously had little or no effect on prandial drinking. Feeding increased Na+ concentrations in plasma and cerebrospinal fluid (CSF; n = 5, P < 0.05). Intracerebroventricular losartan (1 mg/h) inhibited the drinking responses to intracarotid infusion of ANG II (0.8 microg/min for 30 min, n = 4, P < 0.01) and to intracerebroventricular infusion of 0.5 M NaCl (1 ml/h for 1 h, n = 5, P < 0.05) but had no effect on drinking responses to intravenous infusion of 4 M NaCl (1.3 ml/min for 30 min). These findings indicate that a brain ANG II-dependent mechanism is involved in postprandial drinking in sheep. They suggest also that the mechanism by which increasing CSF Na+ causes thirst involves brain ANG II and is different from the mechanism subserving the drinking response to changes in blood Na+.

Cloning, expression and regulation of angiotensin II receptors.

Complementary DNAs for angiotensin II type 1 receptor isoforms AT1A and AT1B were cloned by expression cloning from bovine adrenal and rat vascular smooth muscles. Human AT1 receptor was also cloned. Seven transmembrane structures emerged. The AT1 type receptor interacted with more than one type of G-proteins. The ligand binding site of AT1 involving Arg167, Lys199, and Asp263 has been identified by site directed mutagenesis. The regulation of the receptors occur at many stages. The isoform, AT2, was also expression cloned from rat pheochromocytoma cells. Although its ligand binding is not affected by stable GTP analogs, it is a seven transmembrane domain receptor. It mediates the modulations of phosphotyrosine phosphatase by angiotensin II and AT2 specific CGP42112A. The modulation was abolished by pertussis toxin. Thus, AT2 belongs to a new class of angiotensin receptors with unique signalling and regulatory mechanisms. AT1 mediates cellular growth. Interestingly, AT2 expression is inversely related to the mitogenic activity of cells.

Angiotensin II Effects on Second Messengers Involved in Prolactin Secretion Are Mediated by AT1 Receptor in Anterior Pituitary Cells

The two forms of angiotensin II (Ang II) receptors, AT1 and AT2 subtypes, have been demonstrated in many other cells beside the anterior pituitary cells. Attempting to investigate the subtype(s) of Ang II receptors implicated in the multiple transduction mechanisms involved in Ang II stimulation of prolactin (PRL) release by lactotropes, we studied the effect of selective nonpeptidergic Ang II antagonists on the PRL release, adenylate cyclase (AC), and phospholipase C activities. In intact cells, the AT1 antagonist DuP753 blocked Ang II-induced PRL release, reversed in a dose dependent manner Ang II-evoked inositol phosphates production, and inhibited completely the PLC and protein kinase C (PKC) dependent cAMP accumulation induced by Ang II. In membrane preparations, the Ang II receptors were negatively coupled to AC. The AT1 antagonist blocked in a dose dependent manner the inhibitory effect of Ang II on cAMP production. In intact cells, the negative coupling of Ang II receptor with AC was observed only when PKC was down regulated by long term 12-O-tetradecanolylphorbol-13-acetate pretreatment. Ang II was able to inhibit vasoactive intestinal peptide-induced cAMP accumulation, a response which was also prevented by DuP753. The different coupling of Ang II receptor described above implicated only the AT1 type receptor since the AT2 antagonists (PD123177 and PD123319) were ineffective at any doses tested (10(-8) to 10(-5) M). The obtained results indicate that the regulation of PRL secretion involves the AT1 receptor subtype and that this receptor might be coupled to multiple effectors.

Dual effects of angiotensin II on calcium currents in neonatal rat nodose neurons

Angiotensin II (AII) reversibly modulates calcium current in isolated neonatal rat nodose ganglion cells by two different pathways. A maximum inhibitory effect of 43 +/- 6% (n = 25) of the peak calcium current at -10 mV was observed at 10 nM AII. The IC50 of the inhibitory response was 100 pM. Losartan, a specific antagonist for the AT1 type of AII receptor, abolished the AII-induced inhibition, as did preincubation with pertussis toxin (PTX). When omega-conotoxin GVIA (CTX) was added to the bath solution, AII produced no inhibition of the remaining calcium current, indicating that the AII inhibition was mediated through CTX-sensitive calcium channels. Reversible facilitation of calcium current was seen more rarely. The AII-induced facilitation was unaffected by losartan and PTX, indicating that the effect is mediated by a non-AT1 receptor and does not depend upon a PTX-sensitive G-protein. The facilitation is present when the CTX-sensitive current has been blocked and involves activation of a reserve pool of dihydropyridine (DHP)-sensitive channels. In general, a particular neuron exhibited either inhibition or facilitation. However, in some neurons both inhibition and facilitation could be demonstrated in the presence of the appropriate blocking agents.

Fetal angiotensin II levels and vascular (type I) angiotensin receptors in pregnancies complicated by intrauterine growth retardation

Objective To investigate the status of the fetal renin-angiotensin system (RAS) in pregnancies complicated by severe intrauterine growth retardation (IUGR), and its possible relationship to elevated fetoplacental vascular resistance as indicated by abnormal umbilical artery Doppler flow velocity waveforms (FVW). Design Prospective survey of pregnancies falling into predefined categories and presenting at the Queen Mothers Hospital, Glasgow, over the study period. Subjects Effects of mode of delivery and gestational age were investigated using uncomplicated term pregnancies delivered vaginally (SVD group, n= 15) or by elective caesarean section (ECS group, n= 9), and normal pregnancies with spontaneous preterm onset of labour (PREM group, n= 6; normal birthweight for gestational age (31 weeks)). These groups were used as controls for the 13 IUGR cases delivered preterm (31 weeks) by caesarean section in the fetal interest. Main outcomes measures Umbilical artery FVW, birthweight, cord venous angiotensin II concentration ([cv ANG II]), fetoplacental vascular ANG II receptor concentration. Results Cord venous angiotensin II concentration was similar to maternal values in the ECS group (31–101 pmol/1, 95% CI), but was elevated (81–288 pmol/l, P= 0.03) after vaginal delivery. The concentration of ANG II receptors (type AT1, dissociation equilibrium constant, 1.27 nmol/l) in placental primary/secondary stem vascular tissue was lower in the SVD group (18–44 fmol/mg membrane protein, 95% CI), compared with the ECS group (29–122 fmol/mg, P= 0.03) consistent with acute receptor down-regulation by the elevated ANG II levels. No effect of gestational age on receptor number was demonstrable (P= 0.13, PREM (premature delivery) vs ECS group). In the IUGR group, [cv ANG II] (94–378 pmol/l) was markedly elevated compared with the ECS controls (P= 0.001) but receptor concentration (28–84 fmol/mg) was not significantly altered (P= 0.13). No relationships between [cv ANG II] or receptor number and umbilical artery FVW could be identified. No changes in receptor affinity were observed. Conclusion These results indicate activation of the fetal RAS in IUGR and suggest that responsiveness of the fetoplacental vasculature to the peptide is not diminished as would be expected from the elevated plasma ANG II levels. ANG II may contribute to the increased fetoplacental vascular resistance observed in this disorder, but does not apparently account for the abnormal umbilical artery FVW that is observed in a proportion of IUGR cases.

Angiotensin II contractions in coronary artery. Nature of receptors and calcium pools.

Pig coronary artery rings denuded of endothelium contract to the vasoactive hormone angiotensin II (Ang II). The nature of Ang II receptors and their Ca(2+)-pool utilization were examined for contraction of the artery rings and for increase in ultracellular [Ca2+] ([Ca2+]i) in smooth muscle cells cultured from them. Ang II contracted the arteries (EC50 = 7 +/- 4 nM) but with a lower maximal force (1.4 +/- 0.25 N/g tissue) than the contraction with 60 mM K+ (6.11 +/- 0.63 N/g tissue). In the cultured cells it caused a transient increase in [Ca2+]i with an EC50 value of 11 +/- 4 nM. The cells bound Ang II with a dissociation constant (Kd) of 7 +/- 2 nM. Based on the effects of the Ang II antagonists saralasin, DuPont 753, dithiothreitol and PD123319, the Ang II receptors responsible for contraction, increase in [Ca2+]i and Ang II binding to coronary artery smooth muscle were of type AT1. The contraction to Ang II was abolished by EGTA but not by nitrendipine. The sarcoplasmic Ca2+ pump inhibitors cyclopiazonic acid (10 microM CPA) and thapsigargin (1 microM) produced contractions of 4.35 +/- 0.73 and 2.07 +/- 0.54 N/g, respectively. Ang II contractions in the control arteries were nearly abolished upon pretreatment with CPA and thapsigargin. CPA and thapsigargin induced contractions were abolished by exposure to EGTA for 1 h but short exposure of the cells to EGTA only modulated the CPA or thapsigargin induced increase in [Ca2+]i; Ang II induced increase in [Ca2+]i was not inhibited by 1 microM nitrendipine but was reduced significantly by a 30-60 sec exposure to EGTA.(ABSTRACT TRUNCATED AT 250 WORDS)

Rat angiotensin II (type 1A) receptor mRNA regulation and subtype expression in myocardial growth and hypertrophy.

Two subtypes of angiotensin II (Ang II) receptors (AT1 and AT2) are distinguished by using the respective specific antagonists. In the present study, we report the regulation of cardiac AT1 type A (AT1A) receptor mRNA levels and the expression pattern of AT1 and AT2 receptors in the growth of the heart and the development and regression of cardiac hypertrophy. The ventricular AT1A mRNA level and the density of Ang II receptors at the neonatal period were significantly increased (3.5-fold and 2.5-fold, respectively) and then downregulated with maturation. The cardiac hypertrophy established in spontaneously hypertensive rats or two-kidney one-clip renovascular hypertensive rats resulted in substantial increases in ventricular AT1A mRNA levels (threefold) and Ang II receptor densities (twofold) as compared with those in respective control rats, whereas the receptor affinity was similar. The proportion of AT1 and AT2 subtypes in the specific Ang II binding in ventricular membranes prepared from normal adult rats was nearly equal. This proportion did not change significantly in the development of myocardial hypertrophy. The regression of cardiac hypertrophy by the normalization of elevated blood pressure completely reversed the increased levels of AT1A mRNA and the receptor density to the control level. Thus, AT1 and AT2 receptors are present in rat ventricular myocardium, and their expression is developmentally regulated and upregulated in response to hypertrophic change. Ang II action exerted through the increased number of Ang II receptors may contribute to the growth of the heart and thus to the maintenance of established hypertrophy as one of the hormones involved in hypertrophy development.

Fetal angiotensin II levels and vascular (type I) angiotensin receptors in pregnancies complicated by intrauterine growth retardation.

To investigate the status of the fetal renin-angiotensin system (RAS) in pregnancies complicated by severe intrauterine growth retardation (IUGR), and its possible relationship to elevated fetoplacental vascular resistance as indicated by abnormal umbilical artery Doppler flow velocity waveforms (FVW). Prospective survey of pregnancies falling into predefined categories and presenting at the Queen Mothers Hospital, Glasgow, over the study period. Effects of mode of delivery and gestational age were investigated using uncomplicated term pregnancies delivered vaginally (SVD group, n = 15) or by elective caesarean section (ECS group, n = 9), and normal pregnancies with spontaneous preterm onset of labour (PREM group, n = 6; normal birthweight for gestational age (31 weeks)). These groups were used as controls for the 13 IUGR cases delivered preterm (31 weeks) by caesarean section in the fetal interest. Umbilical artery FVW, birthweight, cord venous angiotensin II concentration ([cv ANG II]), fetoplacental vascular ANG II receptor concentration. Cord venous angiotensin II concentration was similar to maternal values in the ECS group (31-101 pmol/l, 95% CI), but was elevated (81-288 pmol/l, P = 0.03) after vaginal delivery. The concentration of ANG II receptors (type AT1, dissociation equilibrium constant, 1.27 nmol/l) in placental primary/secondary stem vascular tissue was lower in the SVD group (18-44 fmol/mg membrane protein, 95% CI), compared with the ECS group (29-122 fmol/mg, P = 0.03) consistent with acute receptor down-regulation by the elevated ANG II levels. No effect of gestational age on receptor number was demonstrable (P = 0.13, PREM (premature delivery) vs ECS group). In the IUGR group, [cv ANG II] (94-378 pmol/l) was markedly elevated compared with the ECS controls (P = 0.001) but receptor concentration (28-84 fmol/mg) was not significantly altered (P = 0.13). No relationships between [cv ANG II] or receptor number and umbilical artery FVW could be identified. No changes in receptor affinity were observed. These results indicate activation of the fetal RAS in IUGR and suggest that responsiveness of the fetoplacental vasculature to the peptide is not diminished as would be expected from the elevated plasma ANG II levels. ANG II may contribute to the increased fetoplacental vascular resistance observed in this disorder, but does not apparently account for the abnormal umbilical artery FVW that is observed in a proportion of IUGR cases.

Angiotensin-II inhibits Na+/K+ pump in rat adrenal glomerulosa cells: possible contribution to stimulation of aldosterone production.

The control of Na+/K+ pump activity was studied in rat adrenal glomerulosa cells. Ninety percent of K+/86Rb accumulation was blocked by ouabain, and the dose-response curve of inhibition by ouabain was monophasic (IC50, approximately 80 microM), suggesting the role of a single type of Na+/K+ pump (alpha-isoenzyme) in 86Rb accumulation by rat glomerulosa cells. The basal activity of the Na+/K+ pump was much higher in glomerulosa cells than in adrenal fasciculata cells or hepatocytes, as judged by the ouabain-sensitive uptake of 86Rb. In contrast to the two other cell types, increasing Na+ influx with the Na+ ionophore monensin failed to significantly affect ouabain-sensitive 86Rb uptake in glomerulosa cells, suggesting that in glomerulosa cells even the resting intracellular Na+ concentration is sufficient for maximal activity of the Na+/K+ pump. Angiotensin-II (AII) inhibited the ouabain-sensitive 86Rb uptake by glomerulosa cells. The effect of AII was abolished by the selective antagonist of the AT1 type of AII receptors (DuP 753), while PD 123177, an AT2 antagonist was ineffective. AT1 receptors of glomerulosa cells coupled to phospholipase-C activation and, thus, to Ca2+ signal. The inhibitory effect of AII was dependent on the extracellular Ca2+ concentration, but an elevation of cytoplasmic Ca2+ by Ca2+ ionophore ionomycin failed to mimic the effect of AII. These data suggest that Ca2+ is required for but does not mediate the inhibitory effect of AII on the Na+/K+pump. Pharmacological activation of protein kinase-C by phorbol ester did not modify 86Rb accumulation by the cells. Ouabain induced a nifedipine-sensitive elevation in the cytoplasmic Ca2+ concentration and exerted a stimulatory effect on aldosterone production, suggesting participation of the inhibition of the Na+/K+ pump in the aldosterone stimulatory action of AII.

Mapping of angiotensin II receptor subtype heterogeneity in rat brain.

Angiotensin II (Ang II) exerts a number of central actions on fluid and electrolyte homeostasis, autonomic activity, and neuroendocrine regulation. In order to evaluate likely sites where these actions are mediated, Ang II receptor binding was localized in rat brain by in vitro autoradiography with the aid of the antagonist analogue 125I-[Sar1, Ile8]Ang II. Two subtypes of Ang II receptor have been identified using recently developed peptide and nonpeptide antagonists. In the periphery, the receptor subtypes differ in distribution, second messenger coupling, and function. Brain Ang II receptor subtypes were therefore differentiated into AT-1 (type I) and AT-2 (type II) subtypes by using unlabelled nonpeptide antagonists specific for the two Ang II subtypes. AT-1 binding was determined to be that inhibited by Dup 753 (10 microM) and AT-2 binding to be that inhibited by PD 123177 (10 microM). The reducing agent dithiothreitol (DTT) decreased binding to AT-1 receptors and enhanced binding to AT-2 receptors. Many brain structures, such as the vascular organ of the lamina terminalis, subfornical organ, median preoptic nucleus, area postrema, nucleus of the solitary tract, and dorsal motor nucleus of the vagus, which are known to be related to the central actions of Ang II, contain exclusively AT-1 Ang II receptors. By contrast, the locus coeruleus, ventral and dorsal parts of lateral septum, superior colliculus and subthalamic nucleus, many nuclei of the thalamus, and nuclei of the inferior olive contain predominantly AT-2 Ang II receptors. The detailed binding characteristics of each subtype were determined by competition studies with a series of analogues of angiotensin and antagonists. The pharmacological specificity obtained in rat superior colliculus and the nucleus of the solitary tract agreed well with published data on AT-1 and AT-2 receptors, respectively. There was a high degree of correlation between the distribution of Ang II binding sites with published data on Ang II-immunoreactive fields and on the sites of Ang II-responsive neurons. The present study also reveals pharmacological heterogeneity of brain Ang II receptors. The subtype-specific receptor mapping described here is relevant to understanding the role of angiotensin peptides in the central nervous system and newly discovered central actions of nonpeptide Ang II receptor antagonists.

Estrogens regulate angiotensin-converting enzyme and angiotensin receptors in female rat anterior pituitary.

We studied the effects of the estrous cycle, ovariectomy and estrogen replacement on angiotensin-converting enzyme (ACE) (kininase II, EC 3.4.15.1) and angiotensin II (AT) receptors in the pituitary gland of the female rat. Quantitative autoradiography, with the use of consecutive pituitary sections, allowed for simultaneous determination of changes in binding and in the potential AT synthetic ability of individual pituitaries, and for a correlation between these two phenomena. In the anterior pituitary, ACE activity and binding of the ACE inhibitor [125I]-351A were not changed during the estrous cycle. Ovariectomy produced a significant increase in ACE activity and binding, and both of these parameters returned to normal after estrogen replacement. There were no changes in ACE activity or binding in the posterior pituitary during the estrous cycle or after ovariectomy or hormone replacement. AT receptors were characterized as of the AT1 type, since they were displaced by the selective AT1 antagonist DuP 753 and not by the AT2 competitor PD 123177. There were marked changes in the concentration of AT1 receptors during the estrous cycle, with highest numbers in metestrus, lower in estrus and diestrus, and lowest during proestrus. Estrogen replacement in ovariectomized rats decreased AT1 receptor number in the anterior pituitary. Our results indicate a dual effect of estrogen on anterior pituitary AT, physiologically on AT receptor expression and pharmacologically on ACE activity.

Variation of the Albedo of the First Artificial Earth Satellite as a Result of the Influence of External Factors

T VARIATION of the albedo and of the character of the surface reflection of a satellite as the result of the influence of the atmosphere, meteoric matter and other physical factors, plays a considerable role in the observation of a satellite, and may also be of independent scientific interest. In connection with this, an attempt was made to establish the fact of the variation in the surface reflectivity according to data from the observation of the first artificial satellite which had a mirror surface and, consequently, at the initial period of its existence had a specular reflection. This circumstance makes it easier to analyze available observations. The method used to determine the variation of the albedo of the first satellite in flight is based on the following facts: 1 The known differences between the specular and diffuse reflection of a sphere. 2 The possible decrease in the amount of the reflected light if the surface condition were altered by the influence of external factors. In Fig. 1 is shown the dependence of the difference of stellar magnitudes (Am = m3 — md) of a sphere on the phase 6 for a specular and a diffused reflection of light (0 is the angle between the directions satellite-sun and satellite-observer). Evidently, with the increase of the angle 0, the difference Am increases, and for 6 > 135-140 deg attains 2.5-3.0 magnitudes. Consequently, it is impossible to observe the diffuse reflection of a sphere in the direction almost opposite the sun, whereas the brilliance of a specular reflection of a sphere does not depend on the phase. During the period from Oct. 4, 1957 to Dec. 6, 1957 more than 200 optical observations of the first Earth satellite were made with astronomical tubes of the AT-1 type. For all these observations the phase angles 6 and the distance of the sun below the horizon A have been computed. Next, the data containing 0, A, the time of observation, the elevation f3, the azimuths A, distances to the satellite — S, stellar magnitudes m (the average observed magnitude of the first satellite reduced to the distance of 500 km was approximately equal to 5-5.5) and differences Am have been separated into two groups corresponding to the two periods of time considerably different from each other because of the conditions of observations. During the first period (from Oct. 9-Oct. 31, 1957) 159 observations were made with 106 passes of the satellite in 23 days (4.6 passes every 24 hr); during the second period (from Nov. 1-Dec. 7,1957) 63 observations were made with 42 passes in 37 days (1.1 passes every 24 hr). At the same time, the number of ephemerides sent to stations during the first period was 904, i.e., 39 every 24 hr, and 1131 during the second period (30 every 24 hr). The insignificant relative decrease in the number of ephemerides does not explain the sharp drop in the number of observations in November. The main reason for the decrease in the number of observations was a large number of cloudy days during this period. On the basis of the processed data of the 200 observations, one can arrive at the following conclusions: 1 During the first period, numerous observations were made at small elevations (20 deg and less), that is, at large distances to the satellite. There are also observations with large phase angles (0 > 130). 2 During the second period, there were no observations with great distances and large phase angles. Table 1 shows the data concerning the number of observations with different fi and 6 during the two periods in question. Table 2 shows the data for @at, 6av, Sav and the altitude of the satellite's flight above the observation points on ascending and descending parts of the orbit in the two described periods. By ascending (direct) loop, we define that part of the orbit which corresponds to the motion of the satellite from southern latitudes to the northern ones, by the descending (retrograde) loop we mean the motion from the northern latitudes to the southern ones. From the data it is evident that in October the satellite did not yet lose its specular reflecting properties, since there were observations with phase angles 6 up to 140 deg. If the surface deteriorates, i.e., if a sharp decrease of the specular reflection coefficient occurs, the brilliance of the satellite with 6 = 140 deg must be two and a half magnitudes smaller than with the