Mitchell I. Steinberg

Angiotensin ii antagonists

Acute blockade of the renin-angiotensin system with the parenterally active angiotensin II antagonist saralasin has been shown to effectively lower blood pressure in a large fraction of patients with essential hypertension and to improve hemodynamics in some patients with congestive heart failure. It is now possible to antagonize chronically angiotensin II at its receptor using the non-peptide angiotensin II inhibitor losartan (DuP 753, MK 954). When administered by mouth, this compound induces a dose-dependent inhibition of the pressor response to exogenous angiotensin II. This effect is closely related to circulating levels of the active metabolite E3174. Preliminary studies performed in hypertensive patients suggest that losartan has a blood pressure lowering action equivalent to that of an ACE inhibitor. Whether this compound will compare favorably with ACE inhibitors requires however further investigation.

Losartan, a nonpeptide angiotensin II (Ang II) receptor antagonist, inhibits neointima formation following balloon injury to rat carotid arteries

Angiotensin-converting enzyme inhibitors have been shown to inhibit intimal thickening following balloon catheterization of rat carotid arteries. To assess the role of the renin-angiotensin pathway and the angiotensin type-I (AT1) receptor in this effect, the nonpeptide Ang II antagonist losartan (DuP 753) or vehicle was infused continuously i.v. in rats from two days before to two weeks after balloon injury to the left common carotid artery; drug effects upon intimal thickening were examined histologically. Losartan produced a dose-dependent reduction in cross-sectional area of intimal lesions determined two weeks post balloon injury. At 5 mg/kg/day a nonsignificant 23% reduction of intimal area was observed. At the higher dose of 15 mg/kg/day, losartan produced a 48% reduction in intimal area (P less than 0.05) compared to the vehicle-infused group. The cellular density of the neointima was not affected by losartan, indicating a probable effect of the drug upon migration and/or proliferation of smooth muscle cells. In separate groups of non-ballooned rats, losartan infusions of 5 and 15 mg/kg/day produced significant rightward shifts (averaging 6.4- and 55-fold, respectively) in curves relating increases in blood pressure to intravenous Ang II in pithed rats determined between 2 and 16 days following initiation of losartan infusion. Mean arterial blood pressure (determined under alpha-chloralose anesthesia) was reduced following continuous losartan infusion for 6 days from 128 +/- 8 mm Hg (vehicle) to 105 +/- 8 mm Hg at 5 mg/kg/day (P less than 0.05), and 106 +/- 4 mm Hg at 15 mg/kg/day (P less than 0.05). Thus, losartan attenuated the vascular response to balloon catheter injury, and this effect was associated with functional block of vascular AT1 receptors. The results support a role for Ang II, acting via AT1 receptors, in myointimal thickening subsequent to balloon injury of rat carotid arteries.

Ventricular refractory period extension caused by defibrillation shocks.

In pentobarbital-anesthetized dogs, transcardiac shocks of up to 30 J or pacing stimuli were delivered to myocardial tissue at different times in the electrical cycle. When delivered midway or later into electrical systole, shocks, but not pacing stimuli, greatly extended the refractory period as determined by left ventricular pacing. There was a positive correlation between both the shock energy and timing and the amount of delay. A 30-J shock given 10 msec before the end of the refractory period extended the refractory period by 63 +/- 15 msec (p less than 0.001), whereas the same shock given 40 msec earlier produced only 25 +/- 10 msec (p less than 0.001) of extension. By comparison, a 5-J shock given at those times produced 36 +/- 18 (p less than 0.005) and 10 +/- 8 msec (p less than 0.001) of extension, respectively. When delivered early into electrical systole, both a pacing stimulus and a shock had no substantial effect on the tissue refractory period. Because the tissue that is late in electrical systole would otherwise be the first to repolarize if no shock were given, the selective refractory period extension may create a period after the shock during which no tissue is repolarized to a level sufficient for wavefront propagation. Thus, the energy- and time-dependent refractory period extension may help explain the mechanism by which ventricular defibrillation occurs during transcardiac shocks.

Non-peptide angiotensin II receptor antagonists discriminate subtypes of 125I-angiotensin II binding sites in the rat brain.

We have utilized quantitative autoradiography to define subtypes of 125I-angiotensin II (AII) binding in rat brain. AII-1 binding (displaced by DuP 753) was found in the nucleus of the solitary tract and the hypothalamus, while AII-2 binding (displaced by WL 19) was found in the thalamus and lateral septum. These results indicate that subtypes of the AII receptor are present in the brain and the AII-1 receptor subtype is present in regions consistant with the known actions of angiotensin.

Separate receptors mediating the positive inotropic and chronotropic effect of histamine in guinea-pig atria☆

The direct positive inotropic effect of histamine was studied on paced left atrial preparation from guinea pigs. Histamine (10(-8) to 10(-4) M) increased the maximum tension developed in left atria incubated at 35degreesC and driven at 2 Hz. The maximum increase in tension was 60% of that observed with norepinephrine. Metiamide (3 X 10(-5) M, a specific H2-receptor antagonist) did not alter the inotropic response of left atria to histamine. However, tripelennamine (a typical H1-receptor antagonist) competitively shifted the histamine inotropic dose--response curve to the right at concentrations from 10(-8) to 10(-7) M. Higher concentrations (3 X 10(-7) and 10(-6) M) caused little further additional shift to the right. The positive chronotropic effect of histamine on spontaneously beating atria was competitively antagonized by metiamide (10(-6) and 3 X 10(-6) M). These results demonstrate that in guinea-pig atria histamine increases myocardial contractility by an interaction with receptors closely related to classical H1-receptors while its chronotropic effect is mediated by interaction with H2-receptors.

Clofilium1--a new antifibrillatory agent that selectively increases cellular refractoriness

Abstract Clofilium is the most promising member in a new series of antifibrillatory agents to selectively prolong cardiac action potential duration (APD) and effective refractory period (ERP). In normal superfused canine Purkinje fibers, clofilium prolonged APD and ERP by a maximum of 35% (ED 50 =1.3 × 10 −8 M). The effect of clofilium reached equilibrium in 61±3 min but APD did not return toward control during several hr of superfusion with drug-free medium. No change in rate of rise, amplitude, resting potential or rate of diastolic depolarization was noted in the presence of clofilium (3 × 10 −7 M). Clofilium increased the canine ventricular fibrillation threshold (VFT) measured using gated trains of electrical stimuli. This effect occurred in a dose-related fashion following a 30 min infusion of a total of 0.5 or 1.0 μmole/kg of clofilium. The increase was evident within 30 min after ending the infusion and persisted for at least 4 hr. Following the infusion of clofilium (1.0 μmole/kg) 22% of the episodes of ventricular fibrillation (VF) spontaneously reverted to normal sinus rhythm without the use of direct current countershock; this phenomenon did not occur in dextrose-infused dogs.

Cardiac electrophysiological effects of pinacidil and related pyridylcyanoguanidines: relationship to antihypertensive activity.

It has been suggested that certain direct acting vasodilators increase potassium conductance (gK) of smooth muscle membranes. A similar increase of gK in cardiac tissue would be expected to selectively shorten the action potential duration (APD). The present studies were conducted to determine whether a series of pyridyl-cyanoguanidine antihypertensive agents, including pinacidil, might shorten the APD of canine Purkinje fibers and ventricular muscle, and if so, whether this effect can be correlated with their antihypertensive activities in anesthetized spontaneously hypertensive rats (SHR). Cardiac Purkinje fiber APD was decreased by most compounds. P-1075, the most potent compound of the series (EC50 = 3.7 × 10−8 M), reduced the APD of Purkinje fibers by a maximum of 91 ± 2% at 10−6 M with no change in the maximum rate of rise (&OV0312;max) or conduction time of the action potential. Similar selective effects on APD were noted in ventricular muscle cells. Shortening of APD was not altered by muscarinic blockade with atropine (3 × 10−6 M) or adenosine receptor blockade with isobutyl methylxanthine (3 × 10−5 M). In chloralose-anesthetized SHR, the pyridylcyanoguanidines reduced mean arterial blood pressure (BP) and left ventricular dP/dtmax with an order of potency similar to their effect on APD95. The most potent compound was again P-1075 with an ED50 of 8 μg/kg i.v. for its effect on BP. When the ED50 for BP reduction in vivo was compared with the EC50 for Purkinje fiber APD shortening in vitro, an excellent correlation was obtained (r = 0.97, p < 0.005). In summary, the pyridylcyanoguanidines studied, including pinacidil, profoundly decrease cardiac APD without affecting &OV0312;max. Because the decrease in APD was correlated with their antihypertensive activity in anesthetized SHR, the results support the concept that a common mechanism, possibly involving an increase in gK, may underly both phenomena.

The Relation Between Vascular Relaxant and Cardiac Electrophysiological Effects of Pinacidil

Pinacidil may represent an example of a new class of vasodilators that act by increasing membrane permeability to potassium ions. In the present study, the cardiac electrophysiological and venorelaxant effects of a series of pinacidil analogs in canine tissues in vitro were examined. Piacidil (3 x 10(-5) M) markedly reduced action potential duration in Purkinje fibers (82 +/- 3% decrease) and ventricular muscle (54 +/- 2% decrease) without significantly affecting maximal upstroke velocity of the action potential or conduction time. The EC50 for the reduction in Purkinje fiber action potential duration was 2.6 +/- 0.5 microM. Pinacidil also decreased barium-induced automaticity in Purkinje fibers; the concentration that decreased the rate of firing by 50% was identical to the EC50 for decreasing action potential duration. In some preparations, high concentrations of pinacidil (greater than or equal to 3 x 10(-5) M) were associated with the appearance of spontaneous action potentials that were closely coupled to the preceding driven action potential. The EC50 for pinacidil in relaxing phenylephrine-contracted cephalic veins was 0.43 +/- 0.09 microM, and in isolated cat papillary muscle, pinacidil had a direct negative inotropic effect with an EC50 of 4.1 +/- 0.7 microM. Thus, pinacidil was 6 and 10 times more potent in relaxing phenylephrine-contracted veins than in shortening action potential or decreasing cardiac contractility. There was an excellent correlation (r = 0.933, p = 0.002) between decreases in action potential duration and venorelaxation for all pinacidil analogs, as well as for BRL 34915 and nicorandil, two purported potassium channel openers. Significant correlations were also obtained between negative inotropic effects and reductions in action potential duration for the pinacidil series. Pinacidil (10(-5) M) also inhibited the venoconstrictor responses to the selective alpha 2 agonist, B-HT 920, to a greater extent than the alpha 1 agonist, methoxamine. Since a good correlation exists in vitro among all the compounds studied in reducing action potential duration, relaxing vascular tissue, and decreasing cardiac contractility, it is concluded that pinacidil as well as nicorandil and BRL 34915 affect vascular and cardiac tissues by similar mechanisms, possibly by increases in potassium ion permeability, although other mechanisms may also play a role.

Characterization of distinct angiotensin II binding sites in rat adrenal gland and bovine cerebellum using selective nonpeptide antagonists.

We investigated the characteristics of 125I-AII binding to rat adrenal and bovine cerebellar membranes in the presence and absence of new nonpeptide angiotensin II (All) receptor ligands. The imidazole All ligands, DUP753 and WL19, both produced biphasic competition curves to 125I-All binding in rat adrenal glomerulosa and adrenal medulla particles, suggesting the existence of two distinct All binding sites. Antagonist affinity (Ki) and binding capacity (Bmax) for each binding site was determined using nonlinear analysis of competition data fit to a two-site model. The high capacity site (68% of total specific 125I-All bound) in glomerulosa had high affinity for DUP753 (4.6 ± 0.8 nM) and low affinity for WL19 (29 ± 3 μM), and the low capacity site had high affinity for WL 19 (3.3 ± 1.4 nM) and low affinity for DUP753 (51 ± 9 μM). Conversely, in medulla, the high capacity site (77% total binding) had high affinity for WL 19 (19 ± 6 nM) and low affinity for DUP753 (29 ± 8 μM), and the low capacity site had low affinity for WL19 (25 ± 7 μM) but a high affinity for DUP753 (2.8 ± 2.0 nM). In glomerulosa, binding parameters for the nonpeptide ligands at each site derived from monophasic competition curves obtained in the presence of either 0.3 μM DUP753 or WL 19 to selectively block the high or low capacity binding site, respectively, were similar to values determined from the biphasic competition curves. Unlike the nonpeptide inhibitors, unlabeled All yielded monophasic inhibition curves. WL19 and DUP753 also blocked 125I-All binding to bovine cerebellar membranes (Ki = 0.14 ± 0.03 and 170 ± 21 μM, respectively), and both displayed monophasic competition curves. All-induced contraction of rabbit aorta was potently and competitively blocked by DUP753 (K = 3.9 ± 0.4 nM) but not by WL 19. Thus, the Ki values for DUP753 at the DUP753-sensitive binding site in rat adrenal glomerulosa and medulla are similar to the affinity of DUP753 for All receptors in rabbit aorta, suggesting a similarity among these sites. The Ki values for WL19 at the WL19-sensitive site in rat adrenal gland and bovine cerebellum differ from 7–40-fold and represent binding site(s) distinct from the DUP753-sensitive site.

Electrophysiologic properties of propoxyphene and norpropoxyphene in canine cardiac conducting tissues in vitro and in vivo.

Abstract The centrally-acting analgesic, d-propoxyphene, and its N-desmethyl metabolite, d-norpropoxyphene, depressed cardiac conduction in canine myocardial tissues in vitro and in vivo. In Purkinje fibers superfused with propoxyphene or norpropoxyphene (10−6–10−4 m ), there was a decrease in the maximal rate of rise of the upstroke of the action potential (Vmax) and a decrease in total action potential duration and cellular refractoriness. Both agents decreased the amplitude of the compound action potential recorded from isolated rat sympathetic trunk. A significant correlation was shown between the depression of Purkinje fiber Vmax and local anesthetic activity in vitro for propoxyphene, norpropoxyphene, and several typical membrane-active antidysrhythmic agents. In isolated guinea pig atria, both agents (10−6–10−4 m ) decreased sinus frequency and contractility. In conscious dogs both agents (2.1–21 μmol/kg, iv) prolonged P-R interval of the ECG. His bundle electrograms recorded in anesthetized dogs revealed a 10% prolongation in H-V and A-H intervals by norproxyphene in doses of 7.5±2.8 and 6.6±1.3 μmol/kg, respectively. The corresponding values for propoxyphene were 17.1±1.2 and 3.7±0.4 μmol/kg. These observations suggest that cardiac conduction depression may be a factor in some of the cardiac toxicities associated with propoxyphene overdosage.