John F. French

Dual Inhibition of Angiotensin-Converting Enzyme and Neutral Endopeptidase in Rats with Hypertension

Summary: Angiotensin converting enzyme (ACE) and neutral endopeptidase (NEP), are two mechanistically similar enzymes involved in the metabolism of several vasoactive peptides. Selective inhibitors of ACE are effective antihypertensive agents in high-renin, renovascular rats and normal-renin, spontaneously hypertensive rats (SHR), but are not effective in the low-renin, deoxy-corticosterone acetate (DOCA)-salt hypertensive rats. In contrast, NEP inhibitors are only effective in the low-renin model of hypertension. Treatment with a combination of selective inhibitors or with a dual inhibitor of both enzymes produces an antihypertensive response regardless of basal plasma renin activity. In this study, we compared the activities of MDL 100,173, a novel subnanomolar inhibitor of both ACE and NEP, with those of equimolar doses of captopril, a selective ACE inhibitor, following intravenous administration in these three rat models of hypertension. Treatment with MDL 100,173 significantly lowered blood pressure compared to vehicle treatment in all three models, whereas captopril treatment lowered blood pressure in the renovascular and SHR models only. Administration of MDL 100,173 also significantly elevated diuresis and natriuresis compared to either vehicle or captopril treatment in the SHR and DOCA-salt rats. Urinary excretion of atrial natriuretic peptide (ANP) was increased by MDL 100,173 treatment in all three models of hypertension. Treatment with captopril did not alter urine, sodium, or ANP excretion in any of the models. However, plasma-renin activity was elevated by both MDL 100,173 and captopril administration in all three groups. These results indicate that MDL 100,173 is an effective antihypertensive agent across a spectrum of rat models of hypertension, regardless of basal plasma-renin activity, whereas captopril is effective only in high- or normal-renin hypertensive rats. In addition, MDL 100,173 also possesses diuretic and natriuretic activity as a result of its ability to inhibit NEP.

Benzodiazepine Ro 5-4864 increases coronary flow

Ro 5-4864 (chlorodiazepam) increased coronary flow in isolated retrograde perfused Langendorff rat heart preparations without affecting heart rate and left ventricular contractility (dP/dt). On the other hand Ro 5-4023 (clonazepam) produced very little effect. PK 11195 which has been shown to inhibit the binding of Ro 5-4864 to cardiac muscle did not antagonize this vasodilatory effect of Ro 5-4864 but increased coronary flow by itself. The data indicate a specific vasodilatory effect of certain benzodiazepines. The mechanism of action remains unknown.

Possible mechanism of benzodiazepine-induced relaxation of vascular smooth muscle

This study investigated the mechanism of benzodiazepine-induced relaxation of vascular smooth muscle. The ability of several benzodiazepine and isoquinolinecarboxamide compounds, including a pair of enantiomers, to inhibit [3H]Ro5–4864 binding to the peripheral-type benzodiazepine binding site in rat aortic smooth muscle was compared with their relative ability to induce relaxation of rat aortic rings. The binding was performed in a membrane fraction obtained from a pellet centrifuged at 11,400 g and enriched with high-affinity [3H]Ro5–4864 binding. The rank order of potency (Ki for inhibition of [3H]Ro5–4864 binding to isolated membranes was: (−)PK 14067 (6.4 ± 0.7 nM) = PK 11195 (6.6 ± 0.8 nM) > Ro5–4864 (17.6 ± 2.1 nM) > diazepam (600 ± 180 nM) = (+)PK 14068 (530 ± 70 nM)> clonazepam (14,300 ± 2,100 nM). However, micromolar concentrations of these agents were required to induce relaxation of rat aortic rings contracted with KC1 and/or norepinephrine (NE). Moreover, the relaxations induced by these agents were not stereoselective. The rank order of potency (IC50) for relaxation of KCl-induced contracted muscle was: Ro5–4864 (6.6 ± 0.3 μM) = PK 11195 (6.7 ± 0.9 μM) = (−)PK 14067(11.6 ± 0.7 μM) = (+)PK 14068 (7.6 ± 1.1 μM)> diazepam (47.4 ± 5.3 μM) = clonazepam (47.5 ± 5.7 μM). Further investigation of the mechanism of ben-zodiazepine-induced relaxation showed that (−)PK 14067 and (+)PK 14068 inhibited CaCl2-induced contractions. The benzodiazepines relaxed muscle contracted with KC1 to a greater magnitude than those contracted with NE or prostaglandin F2α (PGF2α). These data suggest that the relaxant effects of these agents are unrelated to the high-affinity benzodiazepine binding site and are probably mediated through inhibition of the voltage-operated calcium channel.

Vasodilatory action of amlodipine on rat aorta, pig coronary artery, human coronary artery, and on isolated Langendorff rat heart preparations.

Amlodipine inhibited contractions of rat aortic rings induced by 40 mM KCl (IC50 = 7.5 x 10(-9) M). The time to attain the maximum inhibitory effect of KCl-induced contractions was long (hours) and dependent on the concentration of amlodipine. After 6 h of washing in drug-free normal Krebs-Ringer solution the contractions recovered only partially. The KCl-induced contractions appeared to be more sensitive to inhibition by amlodipine than were norepinephrine-induced contractions. CaCl2-induced contraction of KCl-depolarized aortic rings was inhibited by amlodipine in a complex manner. Amlodipine not only increased ED50 but also inhibited the maximal tension induced by CaCl2. Amlodipine also inhibited 35 mM KCl-induced contractions of pig coronary artery rings (IC50 = 2.2 x 10(-8) M) and human coronary artery rings (IC50 = 2.1 x 10(-8) M). In Langendorff rat heart preparations, low concentrations of amlodipine increased coronary flow (ED50, 10(-9) M) whereas higher concentrations (greater than 10(-7) M) decreased coronary flow. Amlodipine also decreased the rate of contraction (+ dP/dt, IC50 = 3 x 10(-7) M) and the rate of relaxation (-dP/dt, IC50 = 1.2 x 10(-7) M). Amlodipine decreased heart rate but only at high concentrations (greater than 300 nM). The results of this study indicate that amlodipine is a potent vasodilator with similar cardiovascular actions to other dihydropyridines except that its effects are slower in onset and longer lasting.

Dihydropyridine Ca2+ channel agonists and antagonists potentiate ultraviolet light-induced relaxation through cyclic GMP formation in porcine coronary artery

Summary: We investigated the mechanism of dihydropyridine Ca2+ channel agonist potentiation of ultraviolet (UV) light-induced smooth muscle relaxation in porcine coronary artery rings. Rings contracted with the dihydropyridine Ca2+ channel agonist, (+)-S-202–791, were more sensitive to relaxation in response to UV light than were rings contracted with KCl or histamine. Relaxation of (+)-S-202–791-contracted rings was independent of the presence of endothelium and was associated with cyclic GMP formation. Methylene blue (MB) prevented UV light-induced relaxation and cyclic GMP formation. UV light-induced relaxation of histamine and KG contracted rings and cyclic GMP formation were potentiated by (+ )-S-202–791 or the Ca2+ channel antagonist, (-)-R-202–791. Exposure of (+)-5–202-791 to UV light decreased its contractile potency. The data suggest that UV light-induced relaxation of vascular smooth muscle (VSM) is mediated through cyclic GMP formation and that potentiation of UV light-induced relaxation by dihydropyridine Ca2+ channel agonists results from their breakdown to a compound(s) that activates guanylate cyclase.