Biochemical and pharmacological properties of SR 49059, a new, potent, nonpeptide antagonist of rat and human vasopressin V1a receptors.

SR 49059, a new potent and selective orally active, nonpeptide vasopressin (AVP) antagonist has been characterized in several in vitro and in vivo models. SR 49059 showed high affinity for V1a receptors from rat liver (Ki = 1.6 +/- 0.2) and human platelets, adrenals, and myometrium (Ki ranging from 1.1 to 6.3 nM). The previously described nonpeptide V1 antagonist, OPC-21268, was almost inactive in human tissues at concentrations up to 100 microM. SR 49059 exhibited much lower affinity (two orders of magnitude or more) for AVP V2 (bovine and human), V1b (human), and oxytocin (rat and human) receptors and had no measurable affinity for a great number of other receptors. In vitro, AVP-induced contraction of rat caudal artery was competitively antagonized by SR 49059 (pA2 = 9.42). Furthermore, SR 49059 inhibited AVP-induced human platelet aggregation with an IC50 value of 3.7 +/- 0.4 nM, while OPC-21268 was inactive up to 20 microM. In vivo, SR 49059 inhibited the pressor response to exogenous AVP in pithed rats (intravenous) and in conscious normotensive rats (intravenous and per os) with a long duration of action (> 8 h at 10 mg/kg p.o). In all the biological assays used, SR 49059 was devoid of any intrinsic agonistic activity. Thus, SR 49059 is the most potent and selective nonpeptide AVP V1a antagonist described so far, with marked affinity, selectivity, and efficacy toward both animal and human receptors. With this original profile, SR 49059 constitutes a powerful tool for exploring the therapeutical usefulness of a selective V1a antagonist.

Characterization of SR 121463A, a highly potent and selective, orally active vasopressin V2 receptor antagonist.

SR 121463A, a potent and selective, orally active, nonpeptide vasopressin V2 receptor antagonist, has been characterized in several in vitro and in vivo models. This compound displayed highly competitive and selective affinity for V2 receptors in rat, bovine and human kidney (0.6 < or = Ki [nM] < or = 4.1). In this latter preparation, SR 121463A potently antagonized arginine vasopressin (AVP)-stimulated adenylyl cyclase activity (Ki = 0.26+/-0.04 nM) without any intrinsic agonistic effect. In autoradiographic experiments performed in rat kidney sections, SR 121463A displaced [3H]AVP labeling especially in the medullo-papillary region and confirmed that it is a suitable tool for mapping V2 receptors. In comparison, the nonpeptide V2 antagonist, OPC-31260, showed much lower affinity for animal and human renal V2 receptors and lower efficacy to inhibit vasopressin-stimulated adenylyl cyclase (Ki in the 10 nanomolar range). Moreover, OPC-31260 exhibited a poor V2 selectivity profile and can be considered as a V2/V1a ligand. In normally hydrated conscious rats, SR 121463A induced powerful aquaresis after intravenous (0.003-0.3 mg/kg) or oral (0.03-10 mg/kg) administration. The effect was dose-dependent and lasted about 6 hours at the dose of 3 mg/kg p.o. OPC-31260 had a similar aquaretic profile but with markedly lower oral efficacy. The action of SR 121463A was purely aquaretic with no changes in urine Na+ and K+ excretions unlike that of known diuretic agents such as furosemide or hydrochlorothiazide. In addition, no antidiuretic properties have been detected with SR 121463A in vasopressin-deficient Brattleboro rats. Thus, SR 121463A is the most potent and selective, orally active V2 antagonist yet described and could be a powerful tool for exploring V2 receptors and the therapeutical usefulness of V2 blocker aquaretic agents in water-retaining diseases.

Abilities of some tryptophan and phenylalanine derivatives to inhibit gastric acid secretion.

Benzotript (N-p-chlorobenzoyl-L-tryptophan) has been shown to be a receptor-antagonist in vivo and in vitro for peptides from the gastrin family. In the present study, we examine tryptophan, and some of its N- and C-acylated derivatives, as well as some phenylalanine derivatives, to show their ability to inhibit gastrin-induced acid secretion in the rat in vivo and to compete for the binding of [125I]-(Leu-15)-HG-17 to its cellular receptor on rabbit isolated gastric mucosal cells. N- and C- derivatives of tryptophan and phenylalanine were found to inhibit gastrin-induced acid secretion and binding of [125I]-(Leu-15)-HG-17 to its mucosal cell receptors. By either criterion, the relative antagonistic potencies of the compounds tested were: tert-butyloxycarbonyl-L-tryphophan-p-nitrophenyl ester approximately equal to tert-butyloxycarbonyl-L-tryptophan-carbamoylmethyl ester greater than tert-butyloxycarbonyl-L-tryptophyl-L-methionyl-carbamoylmethyl ester approximately equal to tert-butyloxycarbonyl-L-phenylalanine-carbamoylmethyl ester approximately equal to tert-butyloxycarbonyl-L-tryptophyl-L-methionyl-amide greater than tert-butyloxycarbonyl-L-tryptophan greater than tert-butyloxycarbonyl-L-phenylalanine greater than benzyloxycarbonyl-L-tryptophan approximately equal to benzotript, with minor differences between the in vivo and the in vitro experiments. These results demonstrate that both the nature of the amino acid residue and the N- and C-substitutions are important in determining antagonist activity and affinity for gastrin receptors.

Effects of long-term angiotensin II AT1 receptor blockade on survival, hemodynamics and cardiac remodeling in chronic heart failure in rats.

OBJECTIVE: The beneficial effect of chronic angiotensin I converting enzyme (ACE) inhibition on survival has for long been established in the rat post-infarction model of chronic heart failure (CHF) and has subsequently been confirmed in humans. This study investigates in rats whether chronic angiotensin II AT1 receptor blockade shares with ACE inhibition the same beneficial effect. METHODS: Rats we subjected to coronary artery ligation and, from 7 days later, orally treated for 7.5 months with placebo or irbesartan (5 or 50 mg/kg/day). RESULTS: Irbesartan dose-dependently increased survival (placebo: 27%, low dose: 52%, high dose: 82%, sham-ligated: 100%; high dose vs placebo: P < 0.001 and vs low dose: P < 0.05; low dose vs placebo: P = 0.11). Irbesartan also dose-dependently decreased urinary cyclic GMP excretion throughout the study. At 7.5 months, it dose-dependently decreased left ventricular (LV) end diastolic pressure. normalized LV pressure maximal rate of rise (dP/dt) and cardiac index values and improved LV and right ventricular regional blood flows (radioactive microspheres) and resistances. At 7.5 months, irbesartan markedly decreased myocardial hypertrophy but had almost no effect on LV dilatation and subendocardial fibrosis. CONCLUSIONS: Long-term angiotensin II AT1 receptor blockade with irbesartan strongly and dose-dependently increases survival in the rat model of coronary ligation-induced CHF. This effect is due to the combination of the beneficial effects that the drug exerts on systemic and coronary hemodynamics, on cardiac pump function and vs cardiac hypertrophy development. Long-term AT1 receptor blockade might thus prove useful and prolong survival in human CHF.

Aldehydic peptides inhibiting renin.

Aldehydic peptides in which the C terminal residue is leucinal or phenylalaninal were synthesized. These compounds exhibited potent inhibition of renin activity and appeared to be precursors of transition state analogues for renin-catalysed amide bond hydrolysis.

Pharmacological study of SR 47436, a non-peptide angiotensin II AT1-receptor antagonist, in conscious monkeys.

OBJECTIVE: The hypotensive and hormonal responses of an AT1-subtype angiotensin II receptor antagonist, SR 47436, were investigated and compared with those of DuP 753 (losartan), the leading AT1-receptor antagonist, and captopril and enalapril, two major angiotensin converting enzyme (ACE) inhibitors, in conscious, sodium-replete and sodium-depleted non-human primates. DESIGN AND METHODS: Blood pressure and heart rate were measured in conscious, chronically instrumented sodium-replete (n = 3-5) and sodium-depleted (n = 4) cynomolgus monkeys (Macaca fascicularis). Plasma renin activity (PRA), active renin and angiotensin II plasma concentrations were determined. RESULTS: SR 47436 induced a dose- and time-related fall in blood pressure in sodium-depleted monkeys; the blood pressure-lowering effect was obtained at a range of doses from one-third to one-tenth the equihypotensive dose of DuP 753 after intravenous and oral administrations. The hypotensive effect obtained with SR 47436 was similar to that of captopril and was sustained in sodium-replete monkeys, although it was weaker and less long-lasting than that of enalaprilat. In both sodium-depleted and sodium-replete monkeys the AT1 antagonist and ACE inhibitors caused similar increases in PRA and active renin. However, although angiotensin II levels increased after SR 47436 or DuP 753 treatment, they decreased after treatment with enalaprilat. Modest decreases in the heart rate sometimes accompanied the hypotension, irrespective of the compound tested. CONCLUSION: These data demonstrate that the AT1 antagonist SR 47436 is an effective hypotensive agent in both sodium-replete and sodium-depleted monkeys, with an intrinsic potency three to 10 times that of DuP 753 and similar to that of ACE inhibitors.

Early and late haemodynamic and morphological effects of angiotensin II subtype 1 receptor blockade during genetic hypertension development.

OBJECTIVE: To investigate the haemodynamic and morphological effects resulting from the chronic administration to young spontaneously hypertensive rats (SHR) of a new selective angiotensin II subtype 1 (AT1) receptor antagonist, SR 47436/BMS-186295 (SR/BMS) both during and after the treatment period. METHODS: SR/BMS (60 mg/kg per day, orally) or distilled water was chronically (from 4 to 20 weeks of age) administered to SHR. At age 8, 14, 20 and 28 weeks the effects of SR/BMS on the systemic and regional haemodynamic (radioactive microsphere technique) and the cardiac and vascular morphological parameters (automatic image analysis) were investigated. RESULTS: SR/BMS limited genetic hypertension development and opposed the age-related rises in total peripheral and regional vascular resistances. Simultaneously, it limited the age-related increases in heart weight, left ventricular cross-sectional area and collagen content. Age-related increases in aortic media thickness and amount of collagen were also significantly reduced, whereas aortic compliance was increased. Eight weeks after withdrawal of treatment the antihypertensive effect of SR/BMS, although attenuated, and the limitation of cardiac and vascular remodelling, persisted. CONCLUSIONS: Early AT1 receptor blockade in SHR opposes genetic hypertension development during the treatment period and persistently after its interruption. Prevention of genetic hypertension development during the treatment period can be accounted for by the limitation of the age-related development of the haemodynamic and morphological abnormalities, whereas the persistence of the antihypertensive effect observed after drug withdrawal is due mainly to a maintained prevention of the development of the cardiovascular morphological alterations.

A novel class of calcium-entry blockers: the 1[[4-(aminoalkoxy)phenyl]sulfonyl]indolizines.

The synthesis and initial biological evaluation of a series of 1-sulfonylindolizines is described. These compounds have been shown to be representatives of a novel class of potent, slow-channel calcium antagonists. All compounds were found to be at least as active as the reference calcium antagonists verapamil and cis-(+)-diltiazem. Structure-activity relationship studies have shown that all compounds possessing an aralkyl group in the amine moiety and an isopropyl or cyclopropyl group at the 2 position of the indolizine are among the most potent calcium antagonists known outside the 1,4-dihydropyridine series. The IC50 values for the inhibition of [3H]nitrendipine binding vary between 0.19 and 4.5 nM whereas the IC50 value for nifedipine is 2.5 nM. One of the compounds in this group (9ab) has now been selected for clinical development.

Structure-activity relationships of C-terminal tri- and tetrapeptide fragments that inhibit gastrin activity.

A series of tri- and tetrapeptide derivatives, analogues of the gastrin C-terminal region with no phenylalanine residue, were synthesized. These peptides were tested for their ability to inhibit gastrin-stimulated acid secretion in vivo as well as binding of [125I]-(Nle11)-HG-13 to gastric mucosal cell receptors in vitro. Most of the peptides tested exhibited gastrin antagonist activity in vivo and in vitro. Most active derivatives were 20-30 times more potent than the well-known gastrin antagonist derivatives proglumide and benzotript and had 20-200 times more binding affinity. The smallest fragment exhibiting antagonist activity was the tripeptide Boc-L-tryptophyl-L-methionyl-L-aspartic acid amide.

Renin inhibitors. Free-Wilson and correlation analysis of the inhibitory potency of a series of pepstatin analogues on plasma renin.

Free-Wilson and correlation analysis were combined to study a series of 34 pepstatin analogues in which mainly position 2 was varied. A statistically highly significant correlation was found between the inhibitory activity of the analogues on an enriched plasma renin preparation and structural parameters of the amino acid side chain in position 2. The crucial parameters were found to be the NMR chemical shift of the alpha-carbon, the localized electrical (inductive) effect, and the van der Waals radius related steric parameter, which demonstrated the dominating influence of electronic inductive effects compared to steric bulk. The model gives insight into the structural requirements for effective inhibition and suggests the histidine-2 derivative, a positive outlier in this series, as a lead compound for further structure-activity studies.

Study of SR 142801, a new potent non-peptide NK3 receptor antagonist on cardiovascular responses in conscious guinea-pig.

1. The cardiovascular responses to intravenous (i.v.) injection of natural tachykinins, substance P (SP), neurokinin A (NKA), neurokinin B (NKB) and selective tachykinin (NK) receptor agonists, [Sar9, Met(O2)11]SP, [beta Ala8]NKA(4-10), [MePhe7]NKB and senktide were assessed in conscious, freely moving, guinea-pigs. 2. SP and [Sar9, Met(O2)11]SP (1-1000 pmol kg-1) induced dose-dependent decreases in mean arterial blood pressure (MAP) accompanied by increases in heart rate (HR). NKA evoked only weak hypotensive effects at high doses (3000 pmol kg-1) whereas [beta Ala8]NKA(4-10) (1-3000 pmol kg-1) had no effects. By contrast, NKB [MePhe7]NKB (1-10,000 pmol kg-1) and senktide (1-1000 pmol kg-1), produced dose-related hypertensive effects with the following rank order of potency: senktide > [MePhe7]NKB > NKB. Bradycardia occurred simultaneously with the increases in arterial pressure. 3. The pressor response to intravenous injection of senktide (300 pmol kg-1) was partially reduced by pretreatment with prazosin (0.71 mumol kg-1), or clonidine (0.38 mumol kg-1) and was completely inhibited by the combination of the two compounds. Atropine (1.5 mumol kg-1) suppressed the decrease in HR induced by senktide without altering the blood pressure response. These findings suggest that the blood pressure response to senktide is an indirect effect mediated by noradrenaline released from sympathetic nerve endings, whereas the bradycardia is of vagal reflex origin. 4. SR 142801, ((S)-(N)-(1-(3-(1-benzoyl-3-(3,4-dichlorophenyl) piperidin-3-yl) propyl)-4-phenyl-piperidin-4-yl)-N-methylacetamide), a potent and specific non-peptide NK3 receptor antagonist dose-dependently (0.46-4.6 mumol kg-1, i.v.; 4.6-46 mumol kg-1, p.o.) inhibited the cardiovascular effects of senktide and displayed a long-lasting inhibitory effect after oral administration. By contrast, SR 142806 (4.6 mumol kg-1, i.v.), the (R)-enantiomer of SR 142801 had no effect on the responses to senktide. SR 142801 at a high dose (15 mumol kg-1, i.v.) was inactive toward the [Sar9, Met(O2)11]SP-induced hypotension. 5. SR 142801 did not modify MAP in conscious guinea-pigs both after i.v. (4.6 and 15 mumol kg-1) and oral (46 and 150 mumol kg-1) administration, showing a lack of agonistic properties. However, a slight reduction in HR was observed only after i.v. injection. 6. In conclusion, these results show evident differences in the functional role of tachykinin receptors in the peripheral control of the cardiovascular system. Furthermore, a clear pressor effect of senktide, which was selectively blocked by SR 142801, was observed in conscious guinea-pigs. Hence, this antagonist appears suitable for investigating the functional role of NK3 receptors.

A pharmacodynamic study of SR 47436, a selective AT1 receptor antagonist, on blood pressure in conscious cynomolgus monkeys.

1. Conscious normotensive cynomolgus monkeys were chronically instrumented for the measurement of arterial blood pressure and heart rate to investigate the relationships between the plasma concentration, suppression of the pressor response to angiotensin II (AII), compensatory increase in plasma AII, and hypotensive effect obtained after a single oral dose of SR 47436, a potent and specific nonpeptide AT1 receptor antagonist. As blood sampling could influence the hypotensive effect of SR 47436 through activation of the renin angiotensin system (RAS), drug effects were studied in groups of animals with or without blood samplings. 2. SR 47436 at 10 mg kg-1 induced a hypotensive effect which was not greater following a second dose of 30 mg kg-1, indicating that a maximal hypotensive effect had already been obtained. 3. A single oral dose of SR 47436 (10 mg kg-1) caused a sustained hypotension and a marked inhibition of the AII-induced pressor response, lasting for up to 28 h. These effects of SR 47436 are consistent with good oral bioavailability and a slow elimination of the drug (t 1/2 approximately 20 h), and were accompanied by a sustained increase in plasma AII concentration. Taken together, both the hypotensive response and the compensatory increase in AII indicated that vascular and juxtaglomerular AII receptors were blocked. 4. Although a fair correlation between individual plasma drug concentrations and inhibition of AII-induced pressor response was observed, neither the hypotensive effect nor the compensatory increase in AII correlated with the plasma drug levels. 5. Basal arterial pressure and AII-induced pressor response were not affected by blood samplings. 6. These results suggest that SR 47436 is an effective and long lasting AT1 receptor antagonist with a potent hypotensive action in normotensive cynomolgus monkeys. It may be an efficacious blocker of the RAS in man and suitable for once-a-day dosing.

Electrophysiological effects of dronedarone (SR 33589), a noniodinated amiodarone derivative in the canine heart: comparison with amiodarone.

The electrophysiological effects of dronedarone, a new nonionidated analogue of amiodarone were studied after chronic and acute administration in dog Purkinje fibres, papillary muscle and isolated ventricular myocytes, and compared with those of amiodarone by applying conventional microelectrode and patch-clamp techniques. Chronic treatment with dronedarone (2x25 mg(-1) kg(-1) day p.o. for 4 weeks), unlike chronic administration of amiodarone (50 mg(-1) kg(-1) day p.o. for 4 weeks), did not lengthen significantly the QTc interval of the electrocardiogram or the action potential duration (APD) in papillary muscle. After chronic oral treatment with dronedarone a small, but significant use-dependent V(max) block was noticed, while after chronic amiodarone administration a strong use-dependent V(max) depression was observed. Acute superfusion of dronedarone (10 microM), similar to that of amiodarone (10 microM), moderately lengthened APD in papillary muscle (at 1 Hz from 239.6+/-5.3 to 248.6+/-5.3 ms, n=13, P<0.05), but shortened it in Purkinje fibres (at 1 Hz from 309.6+/-11.8 to 287.1+/-10.8 ms, n=7, P<0.05). Both dronedarone (10 microM) and amiodarone (10 microM) superfusion reduced the incidence of early and delayed afterdepolarizations evoked by 1 microM dofetilide and 0.2 microM strophantidine in Purkinje fibres. In patch-clamp experiments 10 microM dronedarone markedly reduced the L-type calcium current (76.5+/-0.7 %, n=6, P<0.05) and the rapid component of the delayed rectifier potassium current (97+/-1.2 %, n=5, P<0.05) in ventricular myocytes. It is concluded that after acute administration dronedarone exhibits effects on cardiac electrical activity similar to those of amiodarone, but it lacks the 'amiodarone like' chronic electrophysiological characteristics.

Pharmacological profile of a new peptidic gastrin antagonist.

Boc-Trp-Met-Asp-NH2 was described as the smallest peptidic fragment which presented gastric antisecretory activity. Some pharmacological aspects of a peptide analogue, Boc-Trp-Leu-Asp-NH2 (Boc-WLD-NH2), were studied on the main biological functions of gastrin. This compound was found to inhibit the binding of gastrin to isolated gastric fundic mucosal cells (IC50 50 microM). On pentagastrin-induced gastric acid secretion in the rat, a dose-dependent inhibition was observed with an ID50 of 55 mumol/kg when pentagastrin (1 microgram/kg per h) was continuously infused and with an ID50 of 7.8 mumol/kg when pentagastrin (1 microgram/kg) was bolus i.v. injected. Similar inhibition was observed on acid secretion induced by pentagastrin in the isolated rat gastric mucosa (IC50 100 microM), whereas the tripeptide had no effect when acid output was triggered by histamine. A dose-dependent inhibition with the tripeptide was shown on pentagastrin induced guinea-pig ileum contractions (IC50 31 microM). The compound had no activity on histamine-stimulated guinea-pig atria (histamine H2-receptor). These results suggest some evidence for a selective antigastrin activity.

Evaluation of the ability of irbesartan to cross the blood-brain barrier following acute intragastric treatment.

The present study evaluated in functional tests the ability of the angiotensin AT1 receptor antagonist irbesartan, 2-n-butyl-3-[(2'-(1H-tetrazol-5-yl)-biphenyl-4-yl)methyl]-1,3-d iaza-spiro[4,4]non-1-en-4-one, in comparison to losartan, 2-n-butyl-4-chloro-5-hydroxymethyl-1-[(2'(1H-tetrazol-5-yl) bi-phenyl-4-yl)methyl]imidazole, to cross the blood-brain barrier following acute intragastric administration. Two tests were used: the dipsogenic response to intracerebroventricular injection of angiotensin II, and Na+ intake in response to adrenalectomy. In normotensive rats, irbesartan reduced the dipsogenic response to angiotensin II, 10 pmol per rat, at the dose of 90 mg/kg, but not at lower doses. Losartan significantly reduced angiotensin II-induced drinking at 30 mg/kg, but not at a lower dose. In spontaneously hypertensive rats, irbesartan reduced the response to angiotensin II at 50 mg/kg, but not at lower doses, while losartan significantly inhibited angiotensin II-induced drinking even at 10 mg/kg. In adrenalectomized rats, the intake of 2% NaCl was inhibited by the intragastric administration of losartan 30 or 50 mg/kg, while irbesartan did not reduce it in doses up to 50 mg/kg. The results of the present study consistently indicate that after acute intragastric administration, the ability of irbesartan to cross the blood-brain barrier is lower than that of losartan.

Effects of irbesartan (SR47436/BMS-186295) on angiotensin II-induced pressor responses in the pithed rat: potential mechanisms of action.

The effects of two new non-peptide angiotensin receptor antagonists, irbesartan (SR 47436/BMS-186295, (2-n-butyl-4-spirocyclopentane-1-[((2'-tetrazol-5-yl)bipheny l-4-yl)methyl]2 - imidazolin-5-one) and SR 47155A (2-n-butyl-4-spirocyclopentane-1-[((2'-carboxy)biphenyl-4-yl)methy l]2- imidazolin-5-one, trifluoroacetate), on angiotensin II-induced pressor responses were studied in the pithed rat in comparison to losartan, EXP 3174 and [Sar1,Val5,Ala8]angiotensin II. SR 47155A (1-10 mg/kg i.v.) and losartan (1-10 mg/kg i.v.) shifted dose dependently the dose-response curve of angiotensin II to the right without affecting the maximal response. SR 47436 (0.3-10 mg/kg i.v.), EXP 3174 (0.03-1 mg/kg i.v.) and [Sar1,Val5,Ala8]angiotensin II (0.03-1 mg/kg i.v.) induced, at least at high doses, a non-parallel shift to the right of the angiotensin II dose-response curve and this was associated with a reduction of the maximal response. During a 70 min period, the effect of [Sar1,Val5,Ala8]angiotensin II (1 mg/kg i.v.) on the angiotensin II (0.3 microgram/kg i.v.)-induced pressor response was shown to be reversible, the effect of SR 47155A (10 mg/kg i.v.) was partially reversible and the effect of SR 47436 (3 mg/kg i.v.), EXP 3174 (1 mg/kg i.v.) or losartan (6 mg/kg i.v.) was not reversed at the end of this 70 min period. Administration of SR 47155A (10 mg/kg i.v.) before SR 47436 (1-10 mg/kg i.v.) reversed the reduced angiotensin II-maximal response induced by SR 47436. Administration of SR 47436 (10 mg/kg i.v.) before SR 47155A (1-10 mg/kg i.v.) prevented the full development of the pressor response as observed in the absence of SR 47436. In the pithed rat, SR 47436 (30 mg/kg i.v.) and losartan (30 mg/kg i.v.) reduced the change in diastolic blood pressure induced by electrical stimulation of the spinal cord only at low stimulation rates. Taken together these results indicate that SR 47436, under in vivo conditions, is a potent non-peptide angiotensin receptor antagonist. The type of antagonism (partially insurmountable but selective) can be explained by different theoretical models which are discussed.

Effect of SR 47436, a novel angiotensin II AT1 receptor antagonist, on human vascular smooth muscle cells in vitro.

Proliferation of smooth muscle cells within the intima plays a key role in vascular occlusive disorders such as atherosclerosis and restenosis following balloon angioplasty. Among the factors that may be important in the development of vascular lesions, several authors have reported that the local angiotensin system participates in modulating the proliferation of smooth muscle cells after arterial injury. This study was therefore designed to characterize the antagonistic properties and to investigate the antiproliferative effect of a newly developed non-peptide angiotensin II AT1 receptor antagonist, SR 47436. This compound is a potent and competitive antagonist of the binding of [125I]angiotensin II to its receptor on cultured human aortic smooth muscle cells, exhibiting an IC50 value of 1.7 +/- 0.6 nM. SR 47436 was 10-fold more potent than DuP 753 (Losartan) (IC50 = 20.8 +/- 3.7 nM). In these same cells, SR 47436 potently inhibited the angiotensin II-induced [Ca2+]i increase (IC50 = 0.53 +/- 0.13 vs. 7.4 +/- 1.3 nM for DuP 753). Angiotensin II is a potent mitogen for human aortic smooth muscle cells in culture, exhibiting a maximum proliferative response at 1 microM. SR 47436 and Losartan prevented angiotensin II-induced proliferation of these cells in a dose-dependent manner (IC50 = 0.32 +/- 0.09 and 0.71 +/- 0.08 microM, respectively). SR 47436 displayed a marked in vitro inhibition of serum-induced smooth muscle cell proliferation (IC50 = 5.5 +/- 0.8 microM). A selective AT2 receptor antagonist, PD 123177 did not affect angiotensin II-induced responses in these cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Vascular effects of [Arg8]vasopressin in the isolated perfused rat kidney.

The renal vascular effects of [Arg8]vasopressin (vasopressin) were investigated in the isolated perfused rat kidney. Vasopressin (0.01-3 nM) elicited a dose-dependent vasoconstriction in kidneys from Sprague Dawley rats, with a EC50 value of 0.206 +/- 0.044 nM. Inhibition of nitric oxide synthase by N omega-nitro-L-arginine (100 microM) shifted the vasopressin-induced vasoconstrictor response curve to the left. Inhibition of cyclooxygenase by indomethacin (10 or 30 microM) blunted the constriction induced by low concentrations of the peptide. Vasopressin, like angiotensin II but not noradrenaline, induced tachyphylaxis, SR 49059 ((2S)1-[(2R,3S)-5-chloro-3-(2-chlorophenyl)-1-(3,4-dimethoxybenzene- sulfonyl)-3-hydroxy-2,3-dihydro-1H-indole-2-carbonyl]-pyrrolidine-2- carboxamide) (1-30 nM), a new potent and selective non-peptide vasopressin V1A receptor antagonist, shifted the concentration-response curve for vasopressin to the right without decreasing the maximum contraction. Antagonism became competitive with a pA2 value (+/- S.D.) of 9.72 +/- 0.20 during inhibition of nitric oxide release. [Mpa1,D-Arg8]Vasopressin (desmopressin; 0.1-100 nM), or vasopressin (0.01-1 nM) after blockade of the vasopressin V1A receptor by SR 49059, induced no vasopressin V2 receptor-related renal relaxation in kidneys with vascular tone previously restored by noradrenaline or prostaglandin F2 alpha. These findings indicate that in the isolated perfused rat kidney vasopressin is a potent renal vasoconstrictor. The constriction depends on activation of smooth muscle vasopressin V1A receptors and is modulated by endothelial nitric oxide but not by prostacyclin or vasopressin V2 receptor-related vasodilation.

Renal vascular reactivity to vasopressin in rats with diabetes mellitus.

We evaluated how renal vascular reactivity to vasopressin changes when nitric oxide (NO) synthesis varies, as has been reported to occur in the course of insulin-dependent diabetes mellitus. Renal vasoconstrictor responses to vasopressin were obtained in young and older Sprague-Dawley control rats (3 and 10 months old) and in age-matched diabetic rats that had been treated with streptozotocin (60 mg/kg i.v.) at the age of 2 months. In young rats, vasopressin (3-1000 ng/kg/min i.v.) induced in vivo a dose-dependent decrease in renal blood flow, which was diminished in streptozotocin diabetic rats (P<0.05). Similarly, in in vitro perfused kidneys, the concentration-response curve for vasopressin (0.03-10 nM) was shifted 3-fold to the right in kidneys isolated from young diabetic rats (P<0.05). This shift was abolished in the presence of an inhibitor of nitric oxide synthesis, N(G)-nitro-L-arginine (100 microM), in the perfusate. In 10-month-old rats, the in vivo renal vasoconstrictor dose-response curve to vasopressin was shifted 10-fold to the left as compared to that for young rats (P<0.001). This shift was similar in both control and diabetic rats. In conclusion, the present study documented the existence of hyporesponsiveness to vasopressin in the early stage of diabetes, possibly related to nitric oxide overproduction. In contrast, renal vascular hyperreactivity to vasopressin occurs with aging, whether the rats are diabetic or not.

Efficacy of SR 47436 (BMS-186295), a non-peptide angiotensin AT1 receptor antagonist in hypertensive rat models.

The efficacy of SR 47436 (BMS-186295), 2-n-butyl-3-[(2'-(1H-tetrazol-5-yl)- biphenyl-4-yl)methyl]-1,3-diaza-spiro[4,4]non-1-en-4-one, a non-peptide angiotensin AT1 receptor antagonist, was characterized in various conscious hypertensive rat models. In spontaneously hypertensive rats, single intravenous or oral doses of SR 47436 induced mild to modest antihypertensive effects. No tolerance of the antihypertensive effect was observed when the oral treatment was extended to 15 days. SR 47436 was highly effective to lower blood pressure in high renin-dependent hypertensive models such as two-kidney, one-clip renal hypertensive rats and renal artery-ligated hypertensive rats. In this last model, intravenous or oral administration of the angiotensin II antagonist produced a dose-dependent decrease in blood pressure. When injected after the maximal effective dose, enalapril did not induce any further decrease in blood pressure. Furthermore, the antihypertensive effect elicited after a single oral dose (10 mg/kg) was long-lasting (at least 24 h). The simultaneous blunting effect of the angiotensin II-induced blood pressure increase indicated clearly that the antihypertensive effect was due to the blockade of vascular angiotensin AT1 receptors. As expected, the angiotensin AT1 receptor antagonist did not show any efficacy in deoxycorticosterone acetate hypertensive rats, with a suppressed renin-angiotensin system. In genetic and renal hypertensive rats, the antihypertensive effect induced after acute dosing of SR 47436 was similar to that observed after losartan and enalapril. A reflex tachycardia accompanied the antihypertensive effect only after intravenous treatment with either SR 47436 or losartan. These results show that this angiotensin II antagonist, SR 47436, is an effective and long-lasting antihypertensive agent in rats.