Bronchodilator activity of (3R)-3-[[[(3-fluorophenyl)[(3,4,5-trifluorophenyl)methyl]amino] carbonyl]oxy]-1-[2-oxo-2-(2-thienyl)ethyl]-1-azoniabicyclo[2.2.2]octane bromide (CHF5407), a potent, long-acting, and selective muscarinic M3 receptor antagonist.

The novel quaternary ammonium salt (3R)-3-[[[(3-fluorophenyl)[(3,4,5-trifluorophenyl)methyl]amino]carbonyl]oxy]-1-[2-oxo-2-(2-thienyl)ethyl]-1-azoniabicyclo[2.2.2]octane bromide (CHF5407) showed subnanomolar affinities for human muscarinic M1 (hM1), M2 (hM2), and M3 (hM3) receptors and dissociated very slowly from hM3 receptors (t(½) = 166 min) with a large part of the receptorial complex (54%) remaining undissociated at 32 h from radioligand washout. In contrast, [(3)H]CHF5407 dissociated quickly from hM2 receptors (t(½) = 31 min), whereas [(3)H]tiotropium dissociated slowly from both hM3 (t(½) = 163 min) and hM2 receptor (t(½) = 297 min). In the guinea pig isolated trachea and human isolated bronchus, CHF5407 produced a potent (pIC(50) = 9.0-9.6) and long-lasting (up to 24 h) inhibition of M3 receptor-mediated contractile responses to carbachol. In the guinea pig electrically driven left atrium, the M2 receptor-mediated inhibitory response to carbachol was recovered more quickly in CHF5407-pretreated than in tiotropium-pretreated preparations. CHF5407, administered intratracheally to anesthetized guinea pigs, potently inhibited acetylcholine (Ach)-induced bronchoconstriction with an ED(50) value of 0.15 nmol/kg. The effect was sustained over a period of 24 h, with a residual 57% inhibition 48 h after antagonist administration at 1 nmol/kg. In conscious guinea pigs, inhaled CHF5407 inhibited Ach-induced bronchoconstriction for at least 24 h as did tiotropium at similar dosages. Cardiovascular parameters in anesthetized guinea pigs were not significantly changed by CHF5407, up to 100 nmol/kg i.v. and up to 1000 nmol/kg i.t. In conclusion, CHF5407 shows a prolonged antibronchospastic activity both in vitro and in vivo, caused by a very slow dissociation from M3 receptors. In contrast, CHF5407 is markedly short-acting at M2 receptors, a behavior not shared by tiotropium.

Pharmacokinetic and pharmacologic properties of delapril, a lipophilic nonsulfhydryl angiotensin-converting enzyme inhibitor.

Delapril is a carboxy-alkyl-dipeptide mainly converted in animals and humans to an active diacid derivative (M-I), which in turn is converted to an active 5-hydroxy-indane diacid (M-III). In humans these metabolites are excreted in the urine. The presence of the indanyl-glycine moiety gives delapril a high lipophilicity, greater than several other angiotensin-converting enzyme (ACE) inhibitors, such as captopril and enalapril. Due to its greater lipophilicity, delapril has been shown to exert a more effective inhibition of vascular ACE than captopril and enalapril, both in vitro and in vivo. The activity of delapril on tissue ACE also lasts longer than on the circulating enzyme. At doses ranging from 1-10 mg/kg orally, delapril exerts a marked and long-lasting antihypertensive action in various experimental models of hypertension. The blood pressure reduction has been shown to be accompanied by suppression of angiotensin II release from the vascular wall. In stroke-prone spontaneously hypertensive rats (SHR-SP) and in SHR with chronic renal failure, besides reducing hypertension, delapril significantly improves survival rate and prevents the development of stroke, cardiac hypertrophy, and renal sclerosis. The ability of delapril to reduce hypertrophy in vascular and cardiac tissue has been demonstrated in both in vitro and in vivo experiments.

Beta 2-adrenoceptor blockade is the basis of guinea-pig bronchial hyper-responsiveness to leukotriene C4 and other agonists.

Four beta-adrenoceptor antagonists, namely (-)-propranolol, (+)-propranolol, ICI-118551 and (+/-)-practolol, were investigated for their effects on leukotriene C4 (LTC4)-induced bronchoconstriction in the anesthetized guinea-pig. (-)-Propranolol was also investigated for its effects on acetylcholine and histamine bronchospasm in the anaesthetized guinea-pig, and on LTC4-induced contractions of guinea-pig isolated trachea and lung parenchyma. The various beta-adrenoceptor antagonists potentiated, dose-dependently, the bronchoconstriction induced by threshold doses of LTC4 and the intensity of the potentiation correlated with the beta 2-blocking capacity possessed by the drugs. (-)-Propranolol potentiated the bronchospasm induced by threshold doses of acetylcholine and histamine but to a lesser degree than the LTC4-induced bronchospasm. The airway hyper-responsiveness induced by (-)-propranolol was unaffected by pretreatment with mepyramine, cyproheptadine, phenoxybenzamine, atropine or indomethacin. The airway hyper-responsiveness induced by (-)-propranolol persisted even in adrenalectomized or reserpine-treated guinea-pigs, although adrenalectomy induced some increase in airway responsiveness. (-)-Propranolol had no effect on LTC4, histamine and acetylcholine-induced contractions of isolated trachea and lung parenchyma. The results show that the airway hyper-responsiveness induced by beta-adrenoceptor antagonists generally correlates with their beta 2-blocking activity. The possibility remains that some other unknown mechanism(s) may also be implicated.

The involvement of the release of nitric oxide in the pharmacological activity of the new furoxan derivative CHF 2363.

1. The mechanism of action and the pharmacological effects of the new furoxan derivative, CHF 2363 (4-ethoxy-3-phenylsulphonylfuroxan), were investigated. 2. Pre-incubation of CHF 2363 with human platelet-rich plasma produced a concentration-dependent inhibition of the platelet aggregation induced by collagen, adenosine diphosphate (ADP) and platelet activating factor (PAF). The test compound was about 5 times more potent than sodium nitroprusside. 3-Isobutyl-1-methyl-xanthine (IBMX) potentiated the antiaggregating effect of CHF 2363. 3. CHF 2363 was a potent inhibitor of rubbed endothelium rabbit aortic ring contraction induced by noradrenaline. Comparison of IC50 values showed that CHF 2363 was as potent as glyceryl trinitrate (GTN). 4. Increasing concentrations of CHF 2363 elevated platelet guanosine 3':5'-cyclic monophosphate (cyclic GMP) levels. Adenosine 3':5'-cyclic monophosphate (cyclic AMP) levels were unaffected. 5. Oxyhaemoglobin reduced all the pharmacological actions of the test compound. Moreover, CHF 2363 concentration-dependently released nitric oxide (NO) in platelet-rich plasma. The NO release was correlated to its ability to increase platelet cyclic GMP levels. 6. After exposure of rat aortic strips to supramaximal concentrations of GTN (550 microM), the vasorelaxant activity of CHF 2363 did not change, although that of GTN decreased about 55 fold. 7. It has been concluded that the new furoxan derivative CHF 2363 exerts a potent antiaggregating and vasorelaxant activity via NO release and increase of cyclic GMP levels. No in vitro cross tolerance between GTN and CHF 2363 was observed.

Beta-adrenoceptor and cyclo-oxygenase block as a tool for evoking the direct bronchoconstrictor effect of leukotriene C4 in the guinea-pig.

The intravenous administration of LTC4 to anaesthetized guinea-pigs induced dose-dependent bronchoconstriction, which was maximal at the dose of 1.6 nmol kg-1. The response was abolished by indomethacin. The maximal dose of LTC4, preceded by both (-)-propranolol and indomethacin, induced maximal, slow in onset and reversible broncho-constriction. This direct effect of LTC4 was markedly and long-lastingly antagonized by FPL-55712. The beta-blocker did not appreciably affect the systemic blood pressure changes induced by LTC4. The procedure described is suggested as a suitable tool for investigating the per se effect of leukotrienes on the guinea-pig airways in vivo, as well as the efficacy of their receptor antagonists.

Effects of alkyl analogs of histamine and metiamide on gastric acid secretion.

Histamine and metiamide analogs with alkyl, methyl, ethyl or isopropyl groups on the imidazole ring in position 5, not previously tested on an in vivo preparation, were studied for their effects on gastric acid secretion in cats with gastric fistulas. Our findings confirmed that the methyl group appears to be optimal for an effect on gastric acid secretion as regards both agonism and antagonism. Ethyl substitution apparently did not change the pharmacological activity of histamine but modified the inhibitory effect of metiamide from competitive to non-competitive kinetics. Isopropyl analogs were inactive on gastric secretion under our experimental conditions.

CHF 2206, a new potent vasodilating and antiaggregating drug as potential nitric oxide donor.

The effects of the new 3,4-disubstituted furoxan, CHF 2206, on vascular tone, platelet aggregation and platelet cyclic 3',5'-guanosine monophosphate (cGMP) levels were investigated. The compound was a potent inhibitor of rabbit aortic ring contraction induced by norepinephrine, the stable prostaglandin F2 alpha analogue, U-46619, and KCl; this activity was independent of endothelium integrity. When pre-incubated with platelet-rich plasma, CHF 2206 potently reduced in a dose-dependent manner the aggregation induced by the threshold aggregating concentration of collagen, ADP or platelet activating factor (PAF). In the same experimental conditions, the test compound increased the platelet cGMP levels and this rise was clearly associated with the inhibition of platelet aggregation. Moreover, 3-isobutyl-1-methyl-xanthine (IBMX) 5 microM potentiated the antiaggregating activity of CHF 2206. These results indicate that the increase in cGMP plays a key role in the CHF 2206-elicited responses. Oxyhemoglobin reduced all the pharmacological actions of the test compound. This evidence, along with the capacity of CHF 2206 to cause inorganic nitrite production in the presence of platelet-rich plasma, strongly suggests that nitric oxide (NO) may be a common reactive intermediate responsible for the effects induced by the drug. In conclusion, the furoxan derivative CHF 2206 exerts a potent antiaggregating and vasodilating activity with a pharmacological profile similar to the one described for NO-donating pro-drugs.

Cardiovascular effects of nicardipine in anesthetized open-chest dogs in the absence and presence of beta-adrenergic receptor blockade: a comparison with nifedipine and verapamil.

Nicardipine, a new 1,4-dihydropyridine calcium-entry blocker, was assessed for cardiovascular effects against nifedipine and verapamil in anesthetized open-chest beagle dogs, in the absence and presence of beta-adrenoceptor blockade (propranolol 1 mg/kg i.v.). In control conditions, intravenous nicardipine and nifedipine (30 nmol/kg) produced decreases in blood pressure of similar magnitude without evidence of cardiodepression. An equihypotensive dose of verapamil (300 nmol/kg), by contrast, induced long-lasting negative chronotropic and inotropic effects, which, when combined with the effects of propranolol, were so marked as to cause the death of two of five dogs (atrioventricular block). Dihydropyridine derivatives at higher doses (100 nmol/kg) produced different responses: nifedipine depressed cardiac performance significantly more than nicardipine and, in the presence of beta-adrenergic blockade, caused the death of three of six dogs due to cardiovascular failure. In the nicardipine group, on the contrary, cardiac function was adequately preserved even in the presence of beta-blocker, and no animals died. The results of the present study confirm a previous in vitro observation: nicardipine is a calcium-entry blocker devoid of remarkable cardiodepressant effects and particularly selective for the vascular smooth muscle.

In vitro effects of nicardipine on vascular and cardiac muscle preparations.

Nicardipine, a new 1,4-dihydropyridine derivative with Ca++-antagonist activity, was tested on rabbit aortic strips contracted with K+ and NA, and on an isolated guinea pig heart perfused at constant flow. It inhibited K+-contraction in the aortic strips and reduced coronary resistance with the same potency as nifedipine resulting about 10 time less potent than the latter in depressing the myocardial contractility. The effects of nicardipine appear to be related essentially to its Ca++-antagonist activity rather than to any antiphosphodiesterase activity.

Regional vasodilating and cardiac effects of nicardipine in anaesthetized open chest dog.

In this study the effects of intravenous nicardipine on the cardiovascular system in anaesthetized open chest dogs were assessed. In the dose range of 3-30 nmol/kg it induced a dose-dependent and long lasting reduction of the vertebral, coronary and femoral vascular resistance. Nicardipine proved to be equipotent, but longer lasting than nifedipine, and about 10 and 100 times more potent than verapamil and papaverine respectively. Unlike papaverine, calcium entry blockers caused a preferential vasodilation of the vertebral and coronary arteries. Hypotension and a reduction of LVSP and dP/dt appeared only at the highest dose tested. However, for both dihydropyridine derivatives, in contrast to verapamil, the cardiodepressant effects were short-lived and resulted less marked for nicardipine than for nifedipine.

Stimulation of acid and pepsin secretions in fistulated cats by dimaprit-like compounds.

In the present paper the Authors describe the effects of dimaprit analogs on cat gastric acid and pepsin secretions. Such compounds are only weakly active as stimulants of H2-receptors, and studied as antagonists fail to inhibit acid and pepsin secretions evoked by dimaprit. The different mechanisms of action of dimaprit and histamine on H2-receptors are discussed.

Omeprazole-like compounds on histamine-stimulated acid and peptic secretions in conscious dog and cat.

A newly synthetized aniline derivative series has been tested on histamine-stimulated gastric acid secretion in conscious cats and dogs and compared to the proton pump inhibitors omeprazole and NC 1300. The compounds markedly and long-lastingly inhibited acid output with a potency lower than omeprazole but very close to the reference compound NC 1300. The results show that the introduction of a nitrogen atom, irrespective of the position in the benzimidazole ring does not seem to influence the gastric antisecretory activity.

Lung gas volumes and expiratory time constant in immature newborn rabbits treated with natural or synthetic surfactant or detergents.

Immature newborn rabbits delivered at a gestational age of 27 days were tracheotomized and treated, via the tracheal cannula, with clinically recommended doses of natural or synthetic surfactant (Curosurf and Exosurf, respectively). Littermates received 0.1% tyloxapol, 5% Tween 20, or saline. The dose volume of Curosurf was 2.5 ml/kg, that of the other materials 5 ml/kg. Animals were kept in a multiplethysmograph system and ventilated for 30 min with a standardized sequence of insufflation pressures. End-expiratory lung gas volume was calculated at the end of the experiment from measurements of lung weight and total lung volume. Tidal volumes were significantly improved in all groups of animals receiving surfactant or detergents. However, expiratory time constant (determined from the tidal volume tracing) was significantly longer, and end-expiratory gas volume significantly larger, in animals treated with Curosurf than in those receiving Exosurf or detergents. These differences were confirmed by semiquantitative evaluation of alveolar air expansion in histological sections. In addition, airway epithelial necrosis was reduced in animals receiving Curosurf, Exosurf, or Tween 20, but not in animals treated with tyloxapol. The discrepancy between improvements in tidal volume, expiratory time constant, and end-expiratory gas volume reflects failure of lung stabilization in animals treated with Exosurf or detergents, probably due to absence of specific hydrophobic proteins in the synthetic products.

Evaluation of potentially injurious effects of exogenous surfactant lysophospholipids on the alveolar epithelium and pulmonary mechanics.

Although lysophospholipids are normally found in the lung and their presence is connected to the metabolic pathway of surfactant phospholipids, several studies have reported that their intratracheal instillation is able to induce severe alveolar epithelial injury. Since lysophospholipids are normally present in exogenous surfactants as a consequence of the nonenzymatic hydrolysis of parent phospholipids during their production and shelf-life, the aim of this study was to test the potential toxicity of surfactant lysophospholipids in artificially ventilated newborn rabbits in comparison with that of pure lysophosphatidylcholine (Lyso-PC) suspensions. In premature (surfactant-deficient) animals, a commercially available Curosurf batch (0.56 mg Lyso-PC/ml) improved lung-thorax compliance and reduced lactate dehydrogenase (LDH), total protein and hemoglobin contents in bronchoalveolar lavage (BAL) fluid. The same batch submitted to thermal stress in order to increase the Lyso-PC content (10.2 mg Lyso-PC/ml) failed to improve lung mechanics but did not induce any significant change in biochemical markers in BAL fluid. When suspended in saline, pure Lyso-PC had a dramatic and dose-dependent tissue-damaging effect with increased LDH, hemoglobin and protein contents in BAL and a fall in the lungthorax compliance, in both immature and mature (near-term) animals. The lack of toxicity of Lyso-PC in Curosurf might be explained by an interaction with surfactant phospholipids.

Protective effects of delapril, indapamide and their combination on stroke occurrence and lifespan in salt-loaded stroke-prone spontaneously hypertensive rats.

The effects of long-term oral administration of delapril (CAS 83435-67-0), indapamide (CAS 26807-65-8) and their combination on the occurrence of stroke and on mortality were investigated in young salt-loaded stroke-prone spontaneously hypertensive rats (SHRsp) for 31 weeks of treatment (8th-39th week of age) and up to 8 weeks thereafter. Body weight and saline consumption were investigated at regular intervals and cerebrovascular lesions, renal and heart weight were assessed after sacrifice. Untreated SHRsp served as controls. About 50% of control animals died within 6 weeks of saline administration and in 56% of surviving animals cerebral lesions were present at sacrifice, while no death and no cerebral lesions were observed in animals drinking saline, to which delapril, indapamide and their combination had been added, up to the end of treatment. This protective effect was maintained even in the withdrawal period. All treatments induced a highly significant (p < 0.001) reduction of heart weight/body weight and kidney weight/body weight ratios.

Lack of cardiostimulant effects of bamifylline in in vitro and in vivo models.

The cardiac effects of bamifylline (Bamifix), an alkylxanthine derivative, were evaluated on in vivo and in vitro cardiac preparations and compared with those of theophylline and enprofylline. In isolated rat and guinea-pig right atria bamifylline, in contrast to the other xanthines, left the rate of contraction unchanged or reduced it. In isolated rat and guinea-pig left atria the compound induced only slight inotropic effects showing an intrinsic activity significantly lower than that of theophylline and enprofylline. In isolated rabbit papillary muscles bamifylline administration was without effect up to 10(-3) mol/l and decreased the contractile force at a higher concentration, while theophylline dose-dependently increased inotropism. Also in anaesthetized rabbits intravenous administration of bamifylline produced negative chronotropic and dromotropic effects that were associated, at the highest dose, with a reduction of myocardial contractility. Theophylline and enprofylline, at the same equimolar doses, induced positive chronotropic and inotropic effects. The results show that bamifylline is devoid of cardiostimulant effects in respect to other xanthine derivatives. Moreover, they suggest that bamifylline acts on cardiac muscle by a mechanism, probably linked to the Ca+(+)-movements across the cardiac membrane, able to outweigh the effects due to its antiphosphodiesterase activity.

Comparative efficacy of a DA2/alpha2 agonist and a beta-blocker in reducing adrenergic drive and cardiac fibrosis in an experimental model of left ventricular dysfunction after coronary artery occlusion.

Attenuation of neuroendocrine activation may be beneficial in congestive heart failure. Sympathetic nervous system overactivity can be reduced by receptors blockade or by reducing norepinephrine (NE) spillover. This study evaluated and compared the effects of a DA2-dopaminergic receptor/alpha2-adrenoceptor agonist (CHF-1024) and a beta1-adrenoreceptor antagonist in terms of hemodynamics, ventricular remodeling, beta-adrenergic drive, and cardiac fibrosis after myocardial infarction (MI) in rats. MI was induced by left coronary artery ligation in 213 rats, whereas 12 were left unoperated on. After 2 months, the operated-on animals were treated for 1 more month with CHF-1024 at either 0.33 mg/kg/day (low dose) or 1 mg/kg/day (high dose) or with metoprolol (10 mg/kg/day), delivered through implanted osmotic minipumps. Plasma concentration and urinary excretion of NE were measured before the rats were killed. Hemodynamic variables were measured and morphometric analysis was done on the diastole-arrested hearts to quantify left ventricular remodeling and interstitial collagen density. Metoprolol treatment tended to normalize LV end-diastolic pressure (LVEDP). CHF-1024 at either dose, and metoprolol, significantly reduced collagen deposition in LV of infarcted animals (from 8.8 +/- 0.5% LV area in vehicle-treated rats to 6.6 +/- 0.2% or 6.4 +/- 0.2% after the low or high dose of CHF-1024, respectively; p < 0.05). Similarly, CHF-1024 at either dose reduced the plasma concentration of NE (from 224 +/- 53 pg/ml to 60 +/- 7 pg/ml or 87 +/- 13 pg/ml; p < 0.05) and urinary excretion of NE in rats with MI, whereas beta-blockade did not affect these variables. In conclusion, CHF-1024 infused for 1 month to rats with LV dysfunction reduced heart rate, NE spillover, and collagen deposition, without unwanted effects, only appearing at the higher dose. Effective beta-blockade with metoprotol reduced LVEDP with no effects on heart function. Neither DA2/alpha2 stimulation nor beta-blockade altered LV remodeling after coronary artery ligation.

Effects of a DA2/alpha2 agonist and a beta1-blocker in combination with an ACE inhibitor on adrenergic activity and left ventricular remodeling in an experimental model of left ventricular dysfunction after coronary artery occlusion.

Renin-angiotensin-aldosterone and sympathetic nervous systems overactivity play a major role in worsening the extent of heart failure. Attenuation of neurohumoral activation with angiotensin-converting enzyme (ACE) inhibitors and beta-blockers has proven beneficial in congestive heart failure. Because ACE inhibition is a recommended treatment for heart failure, this study was designed to test the effects on neurohumoral activation, hemodynamics, and left ventricular (LV) volume of the combination of an ACE inhibitor (delapril) with a DA2-dopaminergic receptor/alpha2-adrenoceptor agonist (CHF-1024) or a beta1-adrenoceptor antagonist (metoprolol) after a moderate to large myocardial infarction (MI) in rats. MI was induced by left coronary artery ligation in 134 rats, and six were not operated on. After 2 months, the animals with ECG evidence of MI were treated for 1 more month with CHF- 1024, 0.33 mg/kg/day or with metoprolol (10 mg/kg/day), delivered through implanted osmotic minipumps, in addition to delapril (6 mg/kg/day) in the drinking water. Daily urinary excretion of norepinephrine (NE) and circulating concentration were measured. Hemodynamic variables were measured, and three-dimensional morphometric analysis was done on the diastole-arrested hearts to quantify infarct size and LV geometry. In conscious animals, delapril alone or with CHF-1024 or metropolol did not modify heart rate or systolic blood pressure. Both combination treatments, however, significantly reduced heart rate in anesthetized animals compared with the group receiving vehicle. Infarct size was not different between treatments, averaging 20-22% of LV volume. The threefold increase of LV chamber volume in infarcted rats was significantly attenuated by delapril alone or with CHF-1024 or metoprolol (-37 to -44%, p<0.05). Treatment with a combination of the ACEi and CHF-1024 tended to normalize the shape of the LV cavity. Urinary NE excretion was unaffected by delapril alone but was reduced by the addition of CHF-1024 or metoprolol. In conclusion, 1 month of treatment with doses of delapril having no hemodynamic effect, reduced LV volume in a model of chronic heart failure. When CHF-1024 or metoprolol was given with delapril, sympathetic activation decreased with no unwanted effects, such as excessive hypotension.

CHF-1024, a DA2/alpha2 agonist, blunts norepinephrine excretion and cardiac fibrosis in pressure overload.

We compared the effects of an ACE inhibitor, captopril, with those of a DA2-dopaminergic/alpha2-adrenergic receptor agonist (CHF-1024) on neuroendocrine activation and cardiac fibrosis in a model of pressure-overload hypertrophy. Interrenal aortic stenosis was performed in 89 rats, treated with CHF-1024 (0.33, 2 or 6 mg kg(-1) day(-1)), or captopril (1 g/L). Hemodynamic variables were recorded. Cardiac and renal weights, plasma aldosterone, renin activity and urinary catecholamine excretion were measured, as well as cardiac collagen. Blood pressure was lower in stenotic animals treated with CHF-1024 compared to vehicle (161 +/- 10 vs 219 +/- 10 mmHg, p < 0.01), but LV weight was similar. CHF-1024 elicited a marked dose-dependent attenuation of urinary norepinephrine excretion (1.80 +/- 0.18 in controls compared to 0.40 +/- 0.14 microg/24 h at the highest dose, p < 0.01) and of LV perivascular fibrosis. Captopril provoked a marked hypotension, reduced cardiac and body weights, plasma aldosterone concentration, dopamine excretion and perivascular collagen. The DA2/alpha2 agonist CHF-1024 effectively blunts adrenergic drive and cardiac fibrosis in a rat model of pressure overload.