Phenylalkylamine Calcium Channel Blocker Research Articles

EFFICACY OF INTRALESIONAL VERAPAMIL HYDROCHLORIDE VERSUS INTRALESIONAL TRIAMCINOLONE ACETONIDE IN HYPERTROPHIC SCARS AND KELOIDS

Background: Keloids and hypertrophic scars are dermal fibro-proliferative disorders lead to disfigurement, pain and pruritus. Their management is still challenging as there is no universally accepted treatment regimen. Corticosteroid injections, most commonly triamcinolone acetonide, continue to play a major role in the management of keloids. Verapamil is a phenylalkylamine calcium channel blocker antiarrhythmic agent that has antifibrotic effect. Objective: To compare efficacy of intralesional verapamil hydrochloride and triamcinolone acetonide in hypertrophic scars and keloids. Patients and methods: Forty Egyptian patients with keloids or hypertrophic scars were divided into two equal groups. The patients were recruited from the Dermatology outpatient clinics of Al-Zahraa University Hospital during the period from May 2016 to May 2017. Informed written consents were obtained from all patients. Group A: Intralesional Verapamil Hydrochloride. Group B: Intralesional Triamcinolone Acetonide. The efficacy of treatment was evaluated by Digital photography and Vancouver scar scale before and after treatment. Two- fine millimeter punch biopsies were taken from 5 patients of the verapamil group before and after treatment to demonstrate the histopathological changes induced by verapamil. Results: Both drugs improved total Vancouver scar scale, vascularity score and pliability score of keloid or hypertrophic scar nearly equally with no statistical significant difference. Both drugs improved height of keloid or hypertrophic scar significantly, but triamcinolone showed better improvement. Verapamil highly improved pigmentation score of keloids and hypertrophic scars, but triamcinolone showed non- significant improvement. Side effects were reported in 4 patients of triamcinolone group with no side effects in verapamil group. The histopathological examination after treatment with verapamil injection showed marked reduction of collagen deposition and alteration of the fibroblast shape (from elongated to spherical), and these changes are similar to histopathological changes which occur after corticosteroids injection. Conclusion: Verapamil was among the several therapeutic modalities, have an option for keloids and hypertrophic scars with an extremely low cost and fewer adverse effects.

Moderate hypothermia and responses to calcium channel blockers - Role of the nitric oxide.

Moderate hypothermia (25-31°C) may have a significant influence on vascular tone. At present, very little is known about the role of endothelial nitric oxide on the hypothermia-induced responses. In this study, we investigated the effect of hypothermia (to 28°C) on the vasodilatation induced by verapamil, a phenylalkylamine calcium channel blocker (10-9-3 × 10-4M) and dihydropyridines, amlodipine (10-9-3 × 10-4M), and benidipine (10-9-10-3M) on 5-hydroxytryptamine (5-HT or serotonin) precontracted calf cardiac veins. Furthermore, the role of nitric oxide in the hypothermia-induced responses was analyzed. Ring preparations of veins obtained from calf hearts were suspended in organ baths containing 15ml of Krebs-Henseleit solution, maintained at 37°C, and continuously gassed with 95% O2-5% CO2. After a resting period, verapamil, amlodipine, and benidipine were applied cumulatively on serotonin (10-6M) precontracted calf cardiac vein rings and induced concentration-dependent relaxations. In another part of the study, the medium temperature was decreased to 28°C after the preparations were contracted with 5-HT, then cumulative concentrations of verapamil, amlodipine, or benidipine were added. During hypothermia, the pIC50 value, but not the maximal response, to all blockers were significantly higher than at 37°C. Hypothermia in the presence of NG-nitro-l-arginine methyl ester (L-NAME, 10-4M) decreased the pIC50 and Emax values to verapamil, amlodipine, and benidipine. Only one blocker was tested in each preparation. These results suggest that nitric oxide may play a role in the hypothermia-induced changes in vasodilation caused by verapamil, amlodipine, and benidipine in calf cardiac vein, but further research is needed to explain the complete mechanism.

The Effects of Emopamil on Neurologic and Neuropathologic Outcomes Following Transient Global Ischemia in Rabbits

Background: Previous studies have suggested that the phenylalkylamine calcium channel blocker, emopamil, may have neuroprotective properties. This study was designed to examine the effects of emopamil on the neurologic and neuropathologic outcome after transient global cerebral ischemia. Methods: New Zealand white rabbits were randomly assigned to one of the following groups: emopamil group (n = 6; emopamil 1 mg/kg bolus prior to ischemia and then 0.1 mg/kg/min for 1 hour), hypothermia group (n = 7; 29oC prior to, during and until 1 hour after ischemia), or control group (n = 7). All rabbits were subjected to 6.25 minutes of global cerebral ischemia produced by a neck tourniquet inflation (20 psi) combined with hypotension during halothane anesthesia. Neurologic deficit scoring was done at 24 hours and 48 hours after ischemia. The rabbits were then euthanized and perfused with 10% formalin. Coronal sections of brain (6m) were stained with hematoxylin and eosin. The sections were scored for evidence of ischemic injury by two observers blinded as to the experimental group. Results: The hypothermia group had significantly less evidence of neurologic and neuropathologic injury when compared to the control group (P 0.05). There were no significant differences between the control and emopamil groups in neurologic or neuropathologic outcome. Conclusions: These results suggest that the peri-ischemic administration of emopamil had no effect on their neurologic or neuropathologic outcome in this model, but, as anticipated, hypothermia was neuroprotective.

Modulation of calcium mobilization in aortic rings of pregnant rats: Contribution of extracellular calcium and of voltage-operated calcium channels.

Pregnancy is associated with decreased vascular responsiveness to vasopressor stimuli. We have tested the involvement of Ca2+ mobilization in myotropic responses of aortic rings obtained from pregnant and virgin rats. Contractions of the rings to phenylephrine, in the absence of calcium in the bathing medium, were lower in tissues from virgin than from pregnant rats. Concentration-response curves to CaCl2 that were measured after stimulation by phenylephrine in the absence of Ca2+ were shifted to higher levels of contraction. This was not observed when KCl was used to prestimulate the aorta. D-600, a phenylalkylamine calcium channel blocker, similarly inhibited these responses to CaCl2 in tissues from both pregnant and virgin animals. D-600 exerted a concentration-dependent inhibition of responses to phenylephrine and KCl. However, the calcium antagonist was less effective in aortic rings of pregnant than of virgin rats. Basal 45Ca2+ uptake was lower in aortic rings from pregnant than from virgin rats, and Bay K 8644 was unable to reverse this difference. The time course of basal and stimulated (KCl) 45Ca2+ influx was lower in aorta of pregnant rats at all times studied. Moreover, when the intracellular calcium pools were emptied with phenylephrine, the refilling of these pools was delayed in aortic rings of pregnant rats. These results indicate an altered extracellular calcium mobilization of aortic rings from pregnant rats. These changes may be due to a functional alteration of the voltage-operated calcium channels during pregnancy.

Methapyrilene hepatotoxicity is associated with oxidative stress, mitochondrial disfunction and is prevented by the Ca 2+ channel blocker verapamil

Methapyrilene (MP) is an unusual hepatotoxin in that it causes periportal necrosis in rats. The mechanism of acute methapyrilene hepatotoxicity has, therefore, been investigated in cultured male rat hepatocytes. Addition of methapyrilene to rat hepatocytes resulted in a time- and dose-dependent loss in cell viability between 4 and 8 h of incubation as judged by cellular enzyme leakage. The cytochrome P450 (CYP) inhibitor metyrapone protected against methapyrilene-mediated toxicity suggesting that MP is metabolised by CYP for toxicity. The concentration-dependent protection from methapyrilene toxicity afforded by metyrapone correlated with an inhibition of microsomal CYP2C11-associated androstenedione 16α hydroxylase activity, and hepatocytes prepared from hypophysectomised rats (containing reduced levels of microsomal immunodetectable CYP2C11 and associated androstenedione 16α hydroxylase activity) showed resistance to the toxic effects of methapyrilene. These data suggest that the toxicity of methapyrilene is predominantly dependent on the CYP2C11 isoform. Treatment of hepatocytes with a toxic concentration of MP caused oxidative stress as indicated by increases in NADP + levels within 2 h and cellular thiol oxidation as evidenced by a reduction—but not complete loss—in glutathione levels. Methapyrilene hepatotoxicity was associated with an early loss in mitochondrial function, as indicated by mitochondrial swelling and significant losses in cellular ATP within 2 h. Co-incubation of methapyrilene-treated hepatocytes with inhibitors of inner mitochondrial transition permeability pore opening—cyclosporin A or the thiol reductant dithiothreitol—abrogated cell death suggesting that pore opening and loss of mitochondrial Ca 2+ homeostasis play a significant role in methapyrilene-mediated cell death. Co-incubation of methapyrilene-treated hepatocytes with the phenylalkylamine calcium channel blocker verapamil—but not by treating cells in a nominally calcium-free medium—also abrogated cell death, suggesting that if Ca 2+ is involved in cell killing then it is dependent on an intracellular Ca 2+ pool. Pre-treatment of hepatocytes for 1 h with verapamil—to inhibit intracellular Ca 2+ pool filling—increased the potency of verapamil protection against methapyrilene toxicity by approximately 100-fold. Taken together, these data indicate that methapyrilene intoxication leads to mitochondrial disfunction and suggest a critical role for a loss of mitochondrial Ca 2+ homeostasis in this model of hepatocyte death.

ChemInform Abstract: Convenient Synthesis of (2R)‐ and (2S)‐2‐(1‐Methylethyl)‐5‐oxo‐2‐ phenylpentanenitrile, Intermediates in the Preparation of Phenylalkylamine Calcium Channel Blockers.

AbstractChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

Mechanism of verapamil calcium channel blockade-induced hyperprolactinemia.

Verapamil, a phenylalkylamine calcium channel blocker, causes a doubling of serum prolactin (PRL) levels in humans. To determine whether the mechanism involved a decrease in the PRL response to dopamine (DA), we infused low doses of DA, finding that the percent inhibition of PRL was not affected by verapamil (max %decrements for 0.003, 0.01, and 0.03 microgram.kg-1.min-1 doses of DA, respectively, 86.7 +/- 19.1, 73.2 +/- 24.8, and 65.2 +/- 20.0% without verapamil and 93.4 +/- 24.6, 79.7 +/- 14.9, and 58.0 +/- 18.1% with verapamil). To determine whether the PRL elevation was due to a decrease in hypothalamic generation of DA, we measured the inhibition of PRL by L-dopa before and after inhibition of peripheral decarboxylase activity with carbidopa. Without verapamil, L-dopa alone and carbidopa-L-dopa caused similar maximum decreases in PRL levels of 83.2 +/- 2.5 and 80.3 +/- 2.0%, respectively. With verapamil, the PRL maximum decrement with L-dopa was 85.2 +/- 2.7% and with carbidopa-L-dopa was 76.3 +/- 1.9% (P < 0.01). We also found that dihydropyridine and benzothiazepine calcium channel blockers had no effect on PRL. These results suggest that verapamil acts by decreasing central DA generation, possibly through N-type calcium channels.

Convenient synthesis of (2R)- and (2S)-2-(1-methylethyl)-5-oxo-2-phenylpentanenitrile, intermediates in the preparation of phenylalkylamine calcium channel blockers

The multigram synthesis of (2S)- and (2R)-2-(1-methylethyl)-5-oxo-2-phenylpentanenitriles 9a and 9b is described, using either (4R)-2,2-dimethyl-1,3-dioxolan-4-ylmethanol or (2R)-butane-1,2,4-triol as chiral auxiliary. The configuration of an intermediate dioxolane 10b is assigned by X-ray crystallography. The synthetic utility of the aldehydes is demonstrated by conversion to both enantiomers of the calcium antagonist noremopamil in >98% enantiomeric excess(ee). The enantiomeric purity of the final amines is assayed by 1H NMR spectroscopy in the presence of the chiral solvating agent (1R)-(–)-2,2,2-trifluoro-1-(9-anthryl)ethanol.

(S)-emopamil attenuates acute reduction in regional cerebral blood flow following experimental brain injury.

We examined the effects of (S)-emopamil, a phenylalkylamine calcium channel blocker with serotonin receptor antagonist properties, on regional cerebral blood flow (rCBF) following experimental brain injury in the rat. Animals were subjected to fluid percussion brain injury of moderate severity (2.1 atm), and received (S)-emopamil (20 mg/kg, i.p., n = 10) or saline (n = 10) at 20 minutes postinjury and 2.5 hours after the first injection of the drug. Consecutive rCBF measurements were performed: (1) prior to injury, (2) 15 minutes, (3) 90 minutes, and (4) 4 hours postinjury, using the radiolabeled microsphere technique. Brain injury produced an acute and significant reduction of rCBF at 15 minutes postinjury in all the regions examined (p < 0.05). At 90 minutes postinjury, rCBF remained significantly depressed in the forebrain regions. All brain regions showed a recovery of rCBF to normal by 4 hours following injury in saline-treated animals, with the exception of injured left parietal cortex and bilateral hippocampi, where rCBF remained significantly depressed. A significant attenuation of the trauma-induced reduction in rCBF was observed at 70 minutes after the first administration of (S)-emopamil in the forebrain regions and cerebellum (p < 0.05). Following the second (S)-emopamil injection, the significant improvement in rCBF observed in left injured cortex was maintained. These results suggest that (S)-emopamil may be efficacious in reversing post-traumatic alterations in rCBF, which may contribute to the post-traumatic pathophysiologic sequelae.

Combined Therapy with Dizocilpine and Levemopamil Does Not Reduce Ischemic Neuronal Injury in Rats

Excessive intracellular accumulation of calcium has been postulated to result in ischemic neuronal death. Reduction of intracellular calcium entry should therefore be expected to reduce ischemic neuronal injury. Two pathways through which extracellular calcium ions can enter neurons are voltage-sensitive and N-methyl-D-aspartate receptor-linked cation pores. Combined blockade of both these types of channels might be more effective in reducing intracellular calcium accumulation than the blockade of either channel alone. We therefore evaluated the cerebroprotective effects of dizocilpine, an N-methyl-D-aspartate receptor antagonist, and levemopamil, a phenylalkylamine calcium channel blocker, administered singly or in combination, in a model of forebrain ischemia in the rat. Four groups of rats (n = 8 each) were studied. In the first group, dizocilpine, 5 mg/kg, was administered before ischemia. In the second group, levemopamil, 5 mg/kg, was given both preischemia and 2 h postischemia. In the third group, both dizocilpine (5 mg/kg) and levemopamil (5 mg/kg) were given preischemia and levemopamil (5 mg/kg) was given postischemia. The control group received saline placebo. The rats were subjected to forebrain ischemia by bilateral carotid artery occlusion for 10 min with simultaneous hypotension to 35 mm Hg. Neuronal injury was evaluated 3 days after ischemia. Dizocilpine reduced postischemia neuronal injury in the ventral hippocampus (p = 0.045). Levemopamil and the combination of levemopamil and dizocilpine did not protect neurons from ischemic injury. The present study does not provide support for the strategy of combined therapy with dizocilpine (administered before ischemia) and levemopamil (administered before and after ischemia) to protect neurons from injury produced by severe incomplete forebrain ischemia.

Calpain activity in organophosphorus-induced delayed neuropathy (OPIDN): effects of a phenylalkylamine calcium channel blocker.

Calpain activity in organophosphorus-induced delayed neuropathy (OPIDN): effects of a phenylalkylamine calcium channel blocker.

Effect of verapamil on the cyclic AMP-mediated pathway for amylase secretion in rat pancreatic acini.

In dispersed acini from rat pancreas, verapamil (a phenylalkylamine calcium channel blocker) potentiated amylase secretion stimulated by vasoactive intestinal peptide (VIP), secretin, peptide histidine isoleucine, helodermin, forskolin, and 8-bromocyclic AMP. The action of verapamil on VIP-stimulated amylase secretion was detectable at 10 microM verapamil and maximal at 100 microM verapamil. Verapamil did not alter binding of 125I-VIP, basal cAMP, the increase in cAMP caused by VIP, or the increase in cAMP-dependent protein kinase caused by VIP. The effects of verapamil on stimulated amylase secretion were fully reversible and could be reproduced by nicardipine (a 1,4-dihydropyridine calcium channel blocker) and diltiazem (a benzothiazepine calcium channel blocker), but not by cinnarizine (a piperazine calcium channel blocker). Although 300 microM verapamil increased outflux of 45Ca, 100 microM verapamil, the concentration that produced maximal potentiation of VIP-stimulated amylase secretion, did not alter 45Ca outflux. Our results indicate that the action of verapamil to potentiate amylase secretion stimulated by secretagogues that activate the cAMP pathway occurs at a step that is distal to the activation of cAMP-dependent protein kinase.

Calcium channel blockers modulate airway constriction in the canine lung periphery

We studied the effect of two voltage-sensitive calcium channel blockers on Na2EDTA-induced bronchoconstriction in the canine lung periphery. A wedged bronchoscope technique was used to measure collateral system resistance before and after challenges with aerosolized Na2EDTA, hypocapnia, aerosolized acetylcholine, and increased flow of dry air in anesthetized mongrel dogs. Nifedipine, a dihydropyridine calcium channel blocker, reduced hypocapnia-induced bronchoconstriction by 88 +/- 6% (SE) but did not alter Na2EDTA-induced constriction. Verapamil, a phenylalkylamine calcium channel blocker, attenuated hypocapnia- and Na2EDTA-induced bronchoconstriction by 69 +/- 6 and 44 +/- 7%, respectively, but did not significantly alter responses to either acetylcholine or dry air challenge. We conclude that calcium influx through voltage-sensitive calcium channels, perhaps of the T subtype, has a limited role in the initiation of Na2EDTA-induced bronchoconstriction in the canine lung periphery.

The molecular and pharmacological basis for using calcium antagonists in hypertension

The interaction of verapamil and other phenylalkylamine calcium channelblockers with the 1,4-dihydropyridine receptor was examined. Studies characterizing the interaction and relationship between calcium channel blocking potency and binding affinity were performed in rat myocardium. The 1,4-dihydropyridines, nifedipine and nitrendipine, interacted competitively. The apparent Kd and Bmax of nitrendipine were 270±140 pm and 390±76 fmol/mg protein, respectively. In contrast, the interaction of the phenylalkylamines with the 1,4-dihydropyridine receptor was not competitive. At a 3H-nitrendipine concentration of 0.12 nm, verapamil displaced only 60% of specifically bound radioactivity and progressively less as the concentration of 3H-nitrendipine increased. Kinetic data indicated that the interaction of both D600 and verapamil with the 1,4-dihydropyridine receptor was not cooperative. At infinite dilution the dissociation rate constant (k−1) was not altered in the presence of 10−5 m D600. We examined the hypothesis that 3H-nitrendipine interacts at several sites with similar affinities and that the phenylalkylamines interact at one of these sites. Although D600 could not further displace 3H-nitrendipine in the presence of a maximally displacing concentration of nifedipine (10−6 m), nifedipine could further displace 3H-nitrendipine in the presence of a maximally displacing concentration of D600 (10−5 m). These results are consistent with competitive interactions at multiple sites but do not explain the diminished ability of the phenylalkylamines to displace progressively less radioactivity at increasing 3H-nitrendipine concentrations. No relationship between binding affinity and pharmacologic potency of the phenylalkylamines was found suggesting that the interaction of the phenylalkylamines with the 1,4-dihydropyridine receptor is not responsible for their calcium channel blocking effects.

Native and detergent-solubilized membrane extracts from Drosophila heads contain binding sites for phenylalkylamine calcium channel blockers

Membrane extracts from Drosophila melanogaster heads contain binding sites for [ 3H]D-600, a verapamil-like calcium channel blocker. Scatchard analysis suggested a two binding site model with a dissociation constant ( K d) of 3.3 ± 0.28 nM and a B max of 1.3 ± 0.14 pmol/mg protein for the high affinity site and a K d of 150 ± 28 nM and a B max of 29 ± 10 pmol/mg protein for the low affinity site. The rank order of affinity of phenylalkylamine binding to the high affinity site was (−)-D-888 > (±)-verapamil > (±)-D-600. Displacement of (−)-[ 3H])D-888 binding by phenylalkylamines is stereoselective. The diphenylpiperazine cinnarizine is an effective inhibitor of [ 3H]D-600 binding. However, [ 3H]D-600 binding is only partially inhibited by diltiazem and bepridil at 10 −4M concentrations, and is not affected by dihydropyridine calcium channel drugs at concentrations < 10 −5M. Moreover, only extremely low specific [ 3H]nitrendipine, (+)-[ 3H]PN200-110, or d-(cis)- [ 3H]diltiazem binding was detectable. The phenylalkylamine photoaffinity probe, [N-methyl- 3H]LU 49888, labeled two Proteinase-K sensitive proteins of 136,000 and 27,000 Da. The high molecular weight, trypsin-sensitive band behaves like the high affinity phenylalkylamine receptor, while the low molecular weight, trypsin-insensitive band has properties of the low affinity receptor. Digitonin-solubilized binding activity has a sedimentation coefficient of 12 S on a sucrose gradient.

Photoaffinity labeling of the multidrug-resistance-related P-glycoprotein with photoactive analogs of verapamil.

Verapamil, a phenylalkylamine calcium channel blocker, has been shown to reverse multidrug resistance in tumor cells, possibly by increasing drug retention through interaction with an outward drug transporter of the resistant cells. In this study two photoactive radioactive analogs of verapamil, N-(p-azido[3,5-3H]benzoyl)aminomethyl verapamil and N-(p-azido[3-125I]salicyl)aminomethyl verapamil, were synthesized and used to identify the possible biochemical target(s) for verapamil in multidrug-resistant DC-3F/VCRd-5L Chinese hamster lung cells selected for resistance to vincristine. The results show that a specifically labeled 150- to 180-kDa membrane protein in resistant cells was immunoprecipitated with a monoclonal antibody specific for P-glycoprotein. Phenylalkylamine binding specificity was established by competitive blocking of specific photolabeling with the nonradioactive photoactive analogs as well as with verapamil. Photoaffinity labeling was also inhibited by 50 microM concentrations of the calcium channel blockers nimodipine, nifedipine, nicardipine, azidopine, bepridil, and diltiazem and partially by prenylamine. Bay K8644, a calcium channel agonist, also inhibited P-glycoprotein photolabeling. Moreover, P-glycoprotein labeling was inhibited in a dose-dependent manner by vinblastine with half-maximal inhibition at 0.2 microM compared to that by verapamil at 8 microM. Photolabeling was also partially inhibited by two of the drugs to which these cells are cross-resistant, doxorubicin and actinomycin D, at 100 microM, but not by colchicine. These data provide direct evidence that P-glycoprotein has broad drug recognition capacity and that it serves as a molecular target for calcium channel blocker action in reversing multidrug resistance.

Correlation between the negative inotropic potency and binding parameters of 1,4-dihydropyridine and phenylalkylamine calcium channel blockers in cat heart.

Partially purified plasma membranes were prepared from cat ventricle. The purification factors for the calcium channel ligands (+)-3H-PN 200-110, 3H-nimodipine (1,4-dihydropyridines) and (-)-3-H-desmethoxyverapamil (a phenylalkylamine) were 3.1-, 3.4- and 2.9-fold, respectively, whilst beta-adrenoceptors labelled with (-)-3H-dihydroalprenolol were purified 3.0-fold. (+)-3H-PN 200-110 bound to 930 +/- 140 fmol/mg of membrane protein with a dissociation constant of 70 pmol/l at 25 degrees C. Under the same conditions 3H-nimodipine bound to 490 +/- 24 fmol/mg of sites with a KD of 120 pmol/l. (-)-3-H-desmethoxyverapamil bound to 530 +/- 55 fmol/mg of sites with a KD of 2.47 nmol/l. Twelve 1,4-dihydropyridines were evaluated for binding inhibition constants (Ki) with (+)-3H-PN 200-110 and 13 phenylalkylamines with (-)-3-H-desmethoxyverapamil in radioligand binding assays. Of the twelve 1,4-dihydropyridines evaluated (+/-)-nitrendipine was the most potent with a Ki-value of 280 pmol/l, nifedipine had a Ki-value of 500 pmol/l and the weakest drug tested, (+/-)-Bay b 4328, had a Ki-value of 14.3 nmol/l. The EC50-values of the same 1,4-dihydropyridines to inhibit the electrically driven cat papillary muscle were 77- to 3,450-fold higher and little correlation existed between Ki and EC50-values. Thirteen phenylalkylamines were tested for their potency to inhibit (-)-3-H-desmethoxyverapamil binding. The most potent phenylalkylamine with respect to negative inotropy was (+/-)-D 595 with an EC50-value of 794 nmol/l, the least potent substance was (+/-)-Sz 45 with an EC50-value of 39.8 mumol/l. The binding inhibition constants for the phenylalkylamines were 13- to 322-fold lower than the values for negative inotropy, but a significant positive correlation between the Ki and EC50-values (n = 12, r = 0.84) was observed. The absolute differences may reflect the state-dependent binding of phenylalkylamines to the channel. QSAR analysis revealed nearly identical correlations between physicochemical substituent properties on the one hand and binding affinities or functional potency on the other hand. In both cases the electronic properties (F-constant) of ring substituents mainly determine the variance in potency.

The interaction of phenylalkylamine calcium channel blockers with the 1,4-dihydropyridine binding site

The interaction of verapamil and other phenylalkylamine calcium channel blockers with the 1,4-dihydropyridine receptor was examined. Studies characterizing the interaction and relationship between calcium channel blocking potency and binding affinity were performed in rat myocardium. The 1,4-dihydropyridines, nifedipine and nitrendipine, interacted competitively. The apparent Kd and Bmax of nitrendipine were 270 +/- 140 pM and 390 +/- 76 fmol/mg protein, respectively. In contrast, the interaction of the phenylalkylamines with the 1,4-dihydropyridine receptor was not competitive. At a 3H-nitrendipine concentration of 0.12 nM, verapamil displaced only 60% of specifically bound radioactivity and progressively less as the concentration of 3H-nitrendipine increased. Kinetic data indicated that the interaction of both D600 and verapamil with the 1,4-dihydropyridine receptor was not cooperative. At infinite dilution the dissociation rate constant (k-1) was not altered in the presence of 10(-5) M D600. We examined the hypothesis that 3H-nitrendipine interacts at several sites with similar affinities and that the phenylalkylamines interact at one of these sites. Although D600 could not further displace 3H-nitrendipine in the presence of a maximally displacing concentration of nifedipine (10(-6) M), nifedipine could further displace 3H-nitrendipine in the presence of a maximally displacing concentration of D600 (10(-5) M). These results are consistent with competitive interactions at multiple sites but do not explain the diminished ability of the phenylalkylamines to displace progressively less radioactivity at increasing 3H-nitrendipine concentrations. No relationship between binding affinity and pharmacologic potency of the phenylalkylamines was found suggesting that the interaction of the phenylalkylamines with the 1,4-dihydropyridine receptor is not responsible for their calcium channel blocking effects.