beta-Adrenergic stimulation of the heart is thought to increase cardiac muscle contractility by activation of cyclic AMP-dependent protein kinase and concomitant increase in the phosphorylation of certain proteins (for refs see refs 1-6). Electrophysiological studies have shown that the stimulation of cardiac beta-adrenoreceptors, the external application of cyclic AMP or its analogues to Purkinje fibres, or the injection of cyclic AMP into single myocytes can increase the slow inward current (Isi) during the plateau phase of the action potential (AP). In heart muscle this current is mainly carried by Ca2+ (refs 10, 11) and it has been suggested that cyclic AMP-dependent phosphorylation of some component of the calcium channel increases the amount of Ca2+ which enters the cell during depolarization. We have investigated this hypothesis by examining the electrical responses of isolated guinea pig ventricular myocytes to pressure injections of subunits of the cyclic AMP-dependent protein kinase. We report here that injection of the catalytic subunit (C) resulted in a lengthening of the action potential duration (APD) and an increase in the height of the plateau as well as the amplitude of Isi. By contrast, the injection of regulatory subunit (R) shortened the APD of fast and slow response APs, an effect which was reversed by adrenaline.

In bovine, cat and guinea pig myocytes the effect of bath application of adrenaline or isoprenaline and of injection of cAMP on the Ca channel was studied with the patch clamp (Hamill et al. 1981), and the following results were obtained. On beta-adrenergic stimulation more activity of the single channel on repeated depolarizations and less records without activity were observed. Correspondingly, the average currents were increased. When the patch contained only one channel as judged from the lack of superpositions during all depolarizations, beta-adrenergic stimulation never produced superpositions indicating that the total number of channels did not increase. Also, it was never possible to activate a channel in the mute patch. The single channel conductance was not changed by catecholamines or cAMP. Increase in probability during depolarization of the channel to be in the open state was proven by non-stationary fluctuation analysis. The kinetic analysis showed a prolongation of the open times and shortening of the shut times by catecholamines, indicating that the rate constants in a three state model C1 in equilibrium C2 in equilibrium O are changed in such a way that the equilibrium shifted towards the open state. In some patches clusters of channels were observed, and activity was greatly increased by adrenaline, isoprenaline and cAMP-injection. The decay of the mean current was either mono-exponential or, in most cases, double-exponential. When the decay was mono-exponential, fluctuation analysis showed an increase in open probability on beta-stimulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Angiotensin converting enzyme inhibition markedly suppresses neointima formation in response to balloon catheter-induced vascular injury of the rat carotid artery. To determine whether this effect was mediated through the vasoactive peptide angiotensin II (Ang II), two approaches were followed. First, the balloon model was used to compare the effects of continuous infusion of Ang II, with and without concurrent converting enzyme inhibition by cilazapril; second, the effects of the orally active nonpeptidic Ang II receptor antagonist DuP 753 were analyzed. Morphometric analysis was performed at 14 days after balloon injury. Animals that received continuous infusion of Ang II (0.3 micrograms/min/rat) were found to have significantly greater neointima formation in response to balloon injury than controls. Animals treated with cilazapril (10 mg/kg/day) had markedly reduced neointima formation, but in animals receiving infusion of Ang II, treatment with cilazapril did not suppress development of neointimal lesions. In the second group of experiments, DuP 753 (10 mg/kg twice daily) was as effective to prevent neointima formation as cilazapril. These data support the conclusions that converting enzyme inhibition prevents neointima formation after vascular injury through inhibition of Ang II generation.

1. Voltage-clamp experiments were conducted on small specimens of rabbit sinoatrial node. In the same preparation the dose-response curve of the potassium current induced by application of different concentrations of acetylcholine (ACh), the time constant of relaxation and the current fluctuations were measured. From these measurements the apparent dissociation constant and the rate constants for the opening and closing of the ACh-activated potassium channel were estimated. 2. In the presence of neostigmine a measurable response was recorded at around 10(-8) M ACh, the saturation was reached at 10(-4) M, and the half saturation was attained at around 10(-6) M. 3. The time constant of relaxation at --35 mV decreased from 100 ms at 10(-8) M to 45 ms at 10(-4) M ACh. 4. The variance of the fluctuations of the ACh-activated current increased with increasing ACh concentration to a peak value of around 10(-5) M. 5. From the above 3 kinds of measurements, opening and closing rate constants of about 12s-1 and 10s-1, respectively, and a dissociation constant of 1.7 microM were calculated. 6. The Katz-Miledi model was considered to be appropriate to describe the reaction of ACh with the muscarinic receptor in the S-A node. 7. The current on ionophoretic application of ACh was computed using the rate constants and taking into account diffusion in the S-A node in which the density of receptors is low. The computed response had a similar time course to the recorded current.

Key Words: Ro 40-5967--calcium antagonists-hypertension-coronary heart diseaseCalcium antagonists are now widely used for the treatment of angina pectoris and hypertension (for review, see ref. 25). Despite different chemical structures, they all inhibit the slow Ca2+ inward current (7,15). Three main classes of calcium antagonists have been described: dihydropyridines, represented by nifedipine; phenylalkylamines, represented by verapamil; and benzothiazepines, represented by diltiazem. Each of these compounds has its own advantages and disadvantages. Dihydropyridine-type calcium antagonists are very potent peripheral vasodilators and can therefore produce headache, ankle edema, and reflex tachycardia (24). Verapamil has a very potent negative inotropic effect (29) that makes this compound dangerous in patients with a compromised myocardium (8,27). Diltiazem is also negatively inotropic, can produce severe bradycardia, and has also been shown to increase the mortality of patients with prior myocardial infarction and clinical signs of heart failure (30).Finally, all existing calcium antagonists (except the novel dihydropyridine, amlodipine) have a poor bioavailability due to a large first-pass effect and a rather short half-life requiring slow-release formulation for once-a-day dosing. Therefore, the purpose of the calcium antagonist program at F. Hoffmann-La Roche Ltd, Basel was to find a compound with the following properties: (a) high bioavailability; (b) long half-life, allowing once-a-day dosing; (c) lack of biologically relevant negative inotropism; and (d) no strong peripheral vasodilation in normotensive subjects.Ro 40-5967, a novel calcium antagonist, which was selected in animal experiments, fulfilled these criteria. The first clinical studies confirmed the preclinical findings. CHEMISTRYThe chemical name of Ro 40-5967 (see Fig. 1) is (lS,2S)-2-(2-[[3-(2-benzimidazolyl)propyl]methylamino]ethyl)-6-fluoro-1,2,3 ,Ctetrahydro-1 -isopropy1-2-naphthylAddress conespondence and reprint requests to Dr.