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Biomedical subjects

D R Ferry

Publications and source records attributed to D R Ferry.

At least 55 records · Page 3Linked to original sources
PubMed1992

Characteristics, evaluation, and complications of prosthetic valves and valvular repair.

The availability of prosthetic heart valves and advances in techniques for their installation have drastically changed the outcome of valvular heart disease. Problems remain, however, in the assessment of prosthetic valve function, durability of prosthetic valves, preservation of left ventricular performance, and the treatment of complications of prosthetic dysfunction. Recent reports have addressed these issues.

Bioprosthesis↗
PubMed1988

Hemodynamic effects of prolonged intravenous therapy with enoximone in patients with severe congestive heart failure.

The hemodynamic differences between bolus administration and constant intravenous infusion over a 48-h period with enoximone, a new positive inotropic/vasodilator agent, were evaluated. Twenty-four patients were studied, 15 patients in the bolus group (Group A) and nine patients in the constant infusion group (Group B). The overall hemodynamic results were similar in both groups. Cardiac output increased in Group A from 3.1 +/- 0.71 to 5.5 +/- 1.3 L/min and in Group B from 3.6 +/- 1.0 to 5.9 +/- 1.2 L/min. Significant decreases occurred in pulmonary capillary wedge pressure (30 +/- 7 to 20 +/- 8 mm Hg and 37 +/- 5 to 21 +/- 11 mm Hg) and systemic vascular resistance (2184 +/- 456 to 1300 +/- 305 dyn.s.cm-5 and 1752 +/- 415 to 1035 +/- 130 dyn.s.cm-5). Group A required repeat drug boluses every 3-5 h to maintain these hemodynamic effects. The terminal blood half-life of enoximone derived following the continuous infusion in Group B was 10.6 +/- 7.0 h. In conclusion, intravenous enoximone produces acute salutary hemodynamic effects in patients with severe congestive heart failure that can be sustained for at least 48 h by intermittent boluses or a continuous infusion.

Adult↗
PubMed1987

Photoaffinity labelling of the cardiac calcium channel. (-)-[3H]azidopine labels a 165 kDa polypeptide, and evidence against a [3H]-1,4-dihydropyridine-isothiocyanate being a calcium-channel-specific affinity ligand.

The arylazide 1,4-dihydropyridine (-)-[3H]azidopine binds to a saturable population of sites in guinea-pig heart membranes with a dissociation constant (KD) of 30 +/- 7 pM and a density (Bmax.) of 670 +/- 97 fmol/mg of protein. This high-affinity binding site is assumed to reside on voltage-operated calcium channels because reversible binding is blocked stereoselectively by 1,4-dihydropyridine channel blockers and by the enantiomers of Bay K 8644. A low-affinity (KD 25 +/- 7 nM) high-capacity (Bmax. 21.6 +/- 9 pmol/mg of protein) site does not bind (-)- or (+)-Bay K 8644, but is blocked by high concentrations (greater than 500 nM) of dihydro-2,6-dimethyl-4-(2-isothiocyanatophenyl)-3,5-pyridinedicarboxy lic acid dimethyl ester (1,4-DHP-isothiocyanate) or, e.g., (+/-)-nicardipine. (-)-[3H]Azidopine was photoincorporated covalently into bands of 165 +/- 8, 39 +/- 2 and 35 +/- 3 kDa, as determined by SDS/polyacrylamide-gel electrophoresis. Labelling of the 165 kDa band is protected stereoselectively by 1,4-dihydropyridine enantiomers at low (nM) concentrations and by (-)- and (+)-Bay K 8644, whereas the lower-Mr bands are not. Thus, only the 165 kDa band is the calcium-channel-linked 1,4-dihydropyridine receptor. Photolabelling of the 39 or 35 kDa bands was only blocked by 10 microM-1,4-DHP-isothiocyanate or 50 microM-(+/-)-nicardipine but not by 10 microM-(-)-Bay K 8644. [3H]-1,4-DHP-isothiocyanate binds to guinea-pig heart membranes with a KD of 0.35 nM and dissociates with a k-1 of 0.2 min-1 at 30 degrees C. [3H]-1,4 DHP-isothiocyanate irreversibly labels bands of 39 and 35 kDa which are protected by greater than 10 microM-(+/-)-nicardipine or unlabelled ligand but not by 10 microM-(-)-Bay K 8644. Thus, [3H]-1,4-DHP-isothiocyanate is not an affinity probe for the calcium channel.

Affinity Labels↗
PubMed1987

Relationship between beta-adrenoceptors and calcium channels in human ventricular myocardium.

The stoichiometric relationship between adrenoceptors and saturable binding sites for 1,4-dihydropyridines in calcium channels was investigated in human ventricular myocardium. Membrane particles were prepared from heart specimens of patients undergoing open heart surgery. The patients suffered from hypertrophic obstructive cardiomyopathy (HOCM) or mitral valve disease. Using [3H]-prazosin and [125I]-2-beta-hydroxy-3-iodiphenyl-ethyl-aminoethyl tetralone ([125I]-HEAT) as labels we detected only a marginal density of alpha 1-adrenoceptors, regardless of disease. No alpha 2-adrenoceptors were detected with [3H]-rauwolscine. In HOCM patients we estimated 72 +/- 10 fmol mg-1 (n = 12) beta-adrenoceptors labelled with [3H]-(-)-dihydroalprenolol and 74 +/- 5 fmol mg-1 (n = 2) beta-adrenoceptors labelled with [125I]-(-)-iodocyanopindolol; the equilibrium dissociation constants KD, were 1.2 +/- 0.2 nmol l-1 for [3H]-(-)-dihydroalprenolol and 7 +/- 1 pmol l-1 for [125I]-(-)-iodocyanopindolol. In patients with mitral valve disease we estimated 84 +/- 11 fmol mg-1 (n = 3) labelled with [3H]-(-)-dihydroalprenolol and 66 +/- 13 fmol mg-1 (n = 2) labelled with [125I]-(-)-iodocyanopindolol. The KD values were 1.8 +/- 0.6 nmol l-1 for [3H]-(-)-dihydroalprenolol and 8 +/- 2 pmol l-1 for [125I]-(-)-iodocyanopindolol. In 14 HOCM patients we estimated 107 +/- 12 fmol mg-1 calcium channel sites labelled with [3H]-nimodipine with a KD of 280 +/- 4 pmol l-1. In 5 patients with mitral valve disease the density of calcium channel sites labelled with [3H]-nimodipine was 78 +/- 5 fmol mg-1 with a KD of 290 +/- 20 pmol l-1, In HOCM patients the density of calcium channel sites labelled with the benzoxadiazol 1,4-dihydropyridine ([3H]-(+)-PN 200-110) was 1.6 fold of that labelled with [3H]-nimodipine with a KD of 84 +/- 11 pmol l-1. In a group of 4 HOCM patients in which calcium channels were labelled with [125I]-iodipine, the density of sites was 1.37 +/- 0.07 fold the density of sites labelled by [3H]-(+)-PN 200-11-. The KD value of [125I]-iodipine was 246 +/- 16 pmol-1. (+)-PN 200-110 was approximately 100 fold more potent than (-)-PN 200-110 as a competitor of [125I]-iodipine binding. For the HOCM group a significant correlation was found between beta-adrenoceptor density and calcium channel density, whereas in the mitral valve group no such correlation was found. This does not prove that there is causal interaction leading to a relationship between the density of beta-adrenoceptors and calcium channels. However, because positive inotropic effects of catecholamines mediated by beta-adrenoceptors are associated with opening of calcium channels, this suggests that the density of both beta-adrenoceptors and calcium channels could be co-regulated.

Calcium Channel Blockers↗
PubMed1987

Interaction between calcium channel ligands and calcium channels.

Distinct drug receptors for 1,4-dihydropyridines, phenylalkylamines, and the benzothiazepine d-cis-diltiazem exist on voltage-dependent calcium channels. The drug receptors show reciprocal allosteric communication and are linked to calcium binding sites. The 1,4-dihydropyridine-selective receptor (probed with [3H]nimodipine) has a size (measured by radiation inactivation) identical in heart, skeletal muscle, and brain (180 kDa). To compare the sizes of 1,4-dihydropyridine receptors in different tissues, pure tritiated enantiomers of the arylazido photoaffinity probe [3H]azidopine were used to irreversibly label the purified 1,4-dihydropyridine receptor (155, 65, and 32 kDa) from guinea pig skeletal muscle transverse tubules and the membrane-bound cardiac receptor. The 155 kDa polypeptide region, but not the 65 or 32 kDa bands, was specifically labelled by (-)-[3H]azidopine. (+)-[3H]Azidopine did not label any of the polypeptides in the purified receptor preparation. In contrast with the results from other investigators, a 155 kDa polypeptide was also specifically labelled in cardiac membranes by (-)-[3H]azidopine. A 34 kDa photolabelled band carries a low-affinity 1,4-dihydropyridine binding site that has no obvious relation to the channel but is abundant in heart membranes and has apparently led to erroneous results in previous affinity or photoaffinity labelling experiments. Antibodies raised against the purified skeletal muscle channel precipitate the 125I-labelled 155 kDa channel polypeptide from skeletal muscle and precipitate the three drug receptor sites from both crude and purified channel preparations. We conclude that all three drug receptor sites are localized on this polypeptide.

Affinity Labels↗
PubMed1986

Stereoselective photoaffinity labelling of the purified 1,4-dihydropyridine receptor of the voltage-dependent calcium channel.

The voltage-dependent calcium channel from guinea-pig skeletal muscle T-tubules has been isolated with a rapid, two-step purification procedure. Reversible postlabelling of the channel-linked 1,4-dihydropyridine receptor and stereoselective photolabelling as a novel approach were employed to assess purity. A 135-fold purification to a specific activity of 1311 +/- 194 pmol/mg protein (determined by reversible equilibrium binding with (+)-[3H]PN200-110) was achieved. Three polypeptides of 155 kDa, 65 kDa and 32 kDa were identified in the purified preparation. The 155-kDa band is a glycoprotein. The arylazide photoaffinity probe (-)-[3H]azidopine bound with high affinity to solubilized membranes (Kd = 0.7 +/- 0.2 nM) and highly purified fractions (Kd = 3.1 +/- 2 nM), whereas the optical antipode (+)-azidopine was of much lower affinity. Irradiation of (-)-[3H]azidopine and (+)-[3H]azidopine receptor complexes with ultraviolet light led to preferential incorporation of the (-) enantiomer into the 155-kDa polypeptide in crude solubilized and purified preparations. The pharmacological profile of irreversible labelling of the 155-kDa glycoprotein by (-)-[3H]azidopine is identical to that found in reversible binding experiments. Specific photolabelling of the 155-kDa band by (-)-[3H]azidopine per milligram of protein increases 150-fold upon purification, whereas incorporation into non-specific bands in the crude solubilized material is identical for both, (-) and (+)-[3H]azidopine.

Affinity Labels↗
PubMed1985

Subunit composition of skeletal muscle transverse tubule calcium channels evaluated with the 1,4-dihydropyridine photoaffinity probe, [3H]azidopine.

The arylazide 1,4-dihydropyridine, [3H]azidopine, binds with high affinity to calcium channels in partially purified guinea-pig skeletal muscle transverse tubule membranes. Upon brief exposure to u.v. light, [3H]azidopine incorporates covalently into transverse tubule membrane proteins, as judged by SDS-PAGE. After alkylation of sulfhydryl groups with N-ethylmaleimide three specifically labelled bands of mol wts. 240 kd, 158 kd and 99 kd are always observed with fluorography after one-dimensional SDS-PAGE. Two other specific bands with mol. wts. of 52 kd and 55 kd, respectively, were sometimes observed. Two-dimensional SDS-PAGE (non-reduced but alkylated in the first dimension and reduced in the second dimension) revealed that the 240-kd band after reduction migrates with a mol. wt. of 99 kd. The 158-kd and 99-kd bands do not change in mobility. It is suggested that [3H]azidopine binds in such a way that the arylazide moiety of the ligand comes into contact with at least three calcium channel components: the A component of mol. wt. 240 kd, the B component of mol. wt. 158 kd and a C component of mol. wt. 99 kd. B and C are non-covalently bonded subunits of the channel, whereas A could be a heterodimer consisting of B and C, linked by disulfide bonds. Subunits of smaller mol. wt. may be also part of the ionic pore. Photolabelling of transverse tubule membranes after high energy irradiation with 10 MeV electrons supports this interpretation.

Affinity Labels↗
PubMed1985

Molecular approach to the calcium channel.

Tritiated 1,4-dihydropyridines (nimodipine, nitrendipine, nifedipine, PN 200-110) and [3H]D-cis-diltiazem as well as [3H]verapamil were employed to directly identify calcium channels in membranes for excitable tissues. The channels, when probed with 1,4-dihydropyridines, exhibit the following properties: 1,4-Dihydropyridine calcium channel blockers bind in a temperature-dependent, reversible manner and with high affinity (dissociation constants 0.2-2 nM at 37 degrees C) to a finite number of sites. For chiral 1,4-dihydropyridines, the binding is stereoselective. Hill slopes of approximately 1.0 are observed. In brain, heart, and solubilized skeletal-muscle membranes, an absolute requirement for certain divalent cations exists in order to bind the ligands with high affinity. Cooperativity (negative and positive) between Me2+ and 1,4-dihydropyridine binding sites is observed. 1,4-Dihydropyridine binding sites are down-regulated in a complex manner by the optically pure enantiomers of D-600 and verapamil. These channel blockers induce, to a different extent, a low-affinity state of the 1,4-dihydropyridine binding site. It is postulated that this allosteric site, at which these blockers act, is closely coupled to the 1,4-dihydropyridine binding site and that a spectrum of compounds exists that differ in their affinity as well as their intrinsic activity to induce the down-regulation. The 1,4-dihydropyridine binding sites are up-regulated by D-cis-diltiazem and KB-944. The up-regulation is temperature-dependent and induces a high-affinity state for 1,4-dihydropyridine channel blockers, accompanied by distinct alterations of the kinetics as well as the pharmacological profile of the 1,4-dihydropyridine binding sites. Complex interactions exist between the channel blockers that induce up-regulation and those that induce down-regulation of the binding. For a given radiolabeled 1,4-dihydropyridine, a tissue-specific (but not species-specific) equilibrium binding dissociation constant is observed. Thus, all heart (human, rat, guinea pig, frog, bovine) have the same KD (0.25 nM at 37 degrees C) for, [3H]nimodipine. The same is observed for brain (KD = 0.5 nM) and for skeletal muscle (KD = 1-2 nM). Three subtypes of channels can be distinguished on the basis of the KD and the tissue-specific up-regulation by D-cis-diltiazem. Subtype-selective drugs exist; e.g., AQA 39 is an inhibitor of [3H]nimodipine binding at skeletal-muscle calcium channels, but not at brain channels. Despite their different pharmacological and kinetic profiles, calcium channels in skeletal muscle and brain have the same molecular size (Mr) when probed by radiation inactivation.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1985

Regional coronary vasoconstriction after combined beta-adrenergic and calcium channel blockade in patients with coronary artery disease.

The beta-adrenergic and calcium channel blocking drugs, which individually and combined have proven efficacious in the treatment of angina pectoris, appear to have opposing effects on coronary artery vasomotion. Previous studies have shown that beta-adrenergic blockade may potentiate and calcium channel blockade reverse coronary vasoconstriction during adrenergic cold stimulation in patients with coronary artery disease. To assess the coronary hemodynamic effects of combined drug therapy, thermodilution coronary sinus and great cardiac vein flow and mean arterial pressure were measured during serial cold pressor testing, both before and after 0.1 mg/kg of intravenous propranolol and again after the addition of 10 mg of sublingual nifedipine in 21 patients (9 without [group A1] and 12 with [group A2] greater than 50% narrowing of the left anterior descending coronary artery). In an additional 15 patients (6 patients without [group B1] and 9 with [group B2] left anterior descending artery stenosis), serial cold pressor testing was performed reversing the drug order. Despite significant increases in mean arterial pressure (p less than 0.01) during cold pressor testing, coronary sinus resistance responses after propranolol plus nifedipine were not statistically significant for any group. However, regional coronary resistance responses differed between patients with and without left anterior descending artery stenosis. In group A1, great cardiac vein resistance was unchanged after propranolol plus nifedipine. In group A2, great cardiac vein flow decreased significantly after propranolol plus nifedipine from 8 +/- 17 to -4 +/- 12% (p less than 0.01 versus control), and great cardiac vein resistance increased from 4 +/- 21 to 15 +/- 19% (p less than 0.01 versus control). A similar significant response was observed for groups B1 and B2. Regional coronary vasoconstriction during adrenergic stimulation after combined drug therapy was only observed in patients with significant left anterior descending artery stenosis. These data suggest that in some patients with severe coronary artery disease, combined beta-adrenergic and calcium channel blockade modified regional coronary responses to adrenergic stimulation with an inhomogeneous distribution of blood flow to potentially ischemic regions without affecting total coronary blood flow. These data also imply that an improvement in anginal symptoms after combined drug therapy may be due primarily to mechanisms that reduce myocardial oxygen demand rather than to improved myocardial oxygen supply.

Adrenergic beta-Antagonists↗
PubMed1985

Relationship between the stereoselective negative inotropic effects of verapamil enantiomers and their binding to putative calcium channels in human heart.

Ventricular preparations from patients with mitral disease and hypertrophic obstructive cardiomyopathy (HOCM) were set up to contract isometrically. Ventricular membrane particles were also prepared and putative calcium channels were labelled with [3H]-nimodipine. Positive staircase was induced by varying the rate of stimulation of isolated strips from 6 min-1 to 120 min-1 in the presence of 6-60 microM (-)-adrenaline or (-)-noradrenaline. (-)-Verapamil 3-5 microM or (+)-verapamil 20-30 microM reversed the force-frequency relationship (i.e. caused negative staircase) in preparations from patients with mitral disease or HOCM. In subendocardial strips of ventricular septum from 5 patients with HOCM paced at 60 min-1, both (-)-verapamil and (+)-verapamil caused cardiodepression. Half-maximal cardiodepression was observed with 0.4 microM (-)-verapamil and with 3 microM (+)-verapamil. [3H]-nimodipine bound to ventricular membrane particles in a saturable, reversible fashion to a high affinity site with an equilibrium dissociation constant of 0.23 nM. The density of these sites was 95 fmol mg-1 of membrane protein. Binding of the tritiated 1,4-dihydropyridine was stereoselectively inhibited by 1,4-dihydropyridine enantiomers and nifedipine. (-)-Verapamil and (+)-verapamil inhibited high affinity [3H]-nimodipine binding in a negative heterotropic allosteric manner with (-)-verapamil being 5 times more potent than (+)-verapamil on an IC50 basis. At a given [3H]-nimodipine concentration, (+)-verapamil inhibited a greater fraction of specific [3H]-nimodipine binding. The allosteric mode of (+)-verapamil inhibition of [3H]-nimodipine binding was confirmed by kinetic studies. (-)-Verapamil shifted (+)-verapamil-binding inhibition curves to the right in an apparently competitive fashion. The inversion of staircase caused by both verapamil enantiomers suggests that they cause a use-dependent channel blockade. The similar potency ratios for binding and for cardiodepression are indicative of a common locus of action for both verapamil enantiomers within the calcium channel.

Aged↗
PubMed1985

Calcium channels: basic properties as revealed by radioligand binding studies.

The structural features and molecular properties of calcium channels have been explored with 1,4-dihydropyridines, d-cis diltiazem and verapamil in radiolabeled form. The concept of multiple (at least three) drug receptor sites of the channel (sites 1, 2, and 3) has been proposed. Site 1 is labeled by 1,4 dihydropyridines, site 2 by (+/-) verapamil (in skeletal muscle) and by (-) desmethoxyverapamil (in brain, skeletal muscle, and heart), site 3 by d-cis diltiazem (in skeletal muscle). The three distinct drug receptor sites communicate with each other via reciprocal allosterism and are linked to divalent cation binding sites. The concept of the reciprocal allosteric communication between the receptor sites has been proven by individual labeling of each of the drug receptor sites in the skeletal muscle t-tubule membrane where the density of calcium channels is extremely high. In brain and heart, labeling of the receptor sites 1 and 2 has also been successful. Three novel ligands for the calcium channel are presented: 125I-iodipine, a 1,4 dihydropyridine ligand, which has an exceptionally high specific activity (2,200 Ci/mmol), 3H-azidopine, a high-affinity, photoaffinity ligand and finally (-) 3H-desmethoxyverapamil, a high-affinity radioligand for receptor site 2. These ligands will aid in the isolation and final characterisation of the calcium channel. 125I-iodipine, 3H-azidopine and (-) 3H-desmethoxyverapamil bind to detergent-solubilized channels from skeletal muscle t-tubules. Whereas the 1,4-dihydropyridines label only a 12.9 S macromolecule, (-) 3H-desmethoxyverapamil additionally labels a approximately 5.0 S peak, where almost no high-affinity 1,4 dihydropyridine binding is found. Taken together with the results from target-size analysis experiments and photoaffinity labeling, an oligomeric structure of the channel is most likely. One polypeptide chain (Mr = 145,000) in reduced form has been unambiguously identified and is currently further characterized. It remains to be seen if the all of the earlier postulated four elements of the channel (alpha, beta, delta, gamma) are on different polypeptide chains or if the 145,000 peptide corresponds to the (beta, delta) or the (delta, gamma) element.

Affinity Labels↗
PubMed1985

Calcium channels and calcium channel drugs: recent biochemical and biophysical findings.

The biochemical and biophysical features of the voltage-dependent calcium channels, as discovered in vitro by means of radiolabelled drugs, are presented. The concept of distinct but reciprocally allosterically coupled drug receptor domains linked to calcium binding sites is explained. The evidence for the existence of isochannels (and isoreceptors) is reviewed and the voltage-dependence of 1,4-dihydropyridine binding and action is discussed. The structure of the channel is investigated by radiation-inactivation and by photoaffinity labelling. Low affinity binding sites for calcium channel drugs are shown to reside on the nucleoside carrier.

Adenosine↗
PubMed1984

(-)-[3H]Desmethoxyverapamil, a novel Ca2+ channel probe. Binding characteristics and target size analysis of its receptor in skeletal muscle.

(-)-[3H]Desmethoxyverapamil (2,7-dimethyl-3-(3,4-dimethoxyphenyl)-3-cyan- 7-aza-9-(3-methoxyphenyl)-nonanhydrochloride) was used to label putative Ca2+ channels in guinea pig skeletal muscle. The binding sites for (-)-[3H]desmethoxyverapamil co-purified with t-tubule membrane markers in an established subcellular fractionation procedure. (-)-[3H]Desmethoxyverapamil bound to partially purified t-tubule membranes with a KD of 2.2 +/- 0.1 nM and a Bmax of 18 +/- 4 pmol/mg membrane protein at 25 degrees C. Binding was stereoselectively inhibited by phenylalkylamine Ca2+ antagonists and in a mixed, non-competitive fashion by the benzothiazepine Ca2+ antagonist d-cis-diltiazem and the 1,4-dihydropyridine Ca2+ antagonist (+)-PN 200-110. Target size analysis of the (-)-[3H]desmethoxyverapamil drug receptor site revealed a molecular mass of 107 +/- 2 kDa. In contrast, the target size of the allosterically coupled benzothiazepine drug receptor site, labelled by d-cis-[3H]diltiazem, was 130.5 +/- 4 kDa (p less than 0.01) and of the 1,4-dihydropyridine binding site 179 kDa, when labelled with [3H]nimodipine. It is concluded that (-)-[3H]desmethoxyverapamil is an extremely useful radioligand for the phenylalkylamine-selective receptor site of the t-tubule localized Ca2+ channel which is allosterically linked to two other distinct drug receptor sites.

Animals↗
PubMed1984

Target size analysis and molecular properties of Ca2+ channels labelled with [3H]verapamil.

[3H]Verapamil was employed to label the drug receptor sites within the Ca2+-channel in skeletal muscle microsomes which are coupled in a negative heterotropic allosteric manner to the previously characterized 1,4-dihydropyridine receptors. At 2 degrees C the Kd of a high-affinity receptor site is 45 nM and the maximum density of binding sites is 37 pmol/mg of protein. Established subcellular fractionation procedures were used to isolate transverse tubule membranes from rabbit and guinea-pig skeletal muscle. [3H]Verapamil, d-cis-[3H]diltiazem as well as 1,4-[3H]dihydropyridine receptors copurify with t-tubule membranes. The ratio of high-affinity verapamil: 1,4-dihydropyridine d-cis-diltiazem Ca2+ channel receptor sites is 4:2:1. The verapamil drug receptors are heat-labile and have essential sulfhydryl groups since they are inactivated by p-chloromercuriphenylsulfonic acid and N-ethylmaleimide. The receptors recognize the main classes of Ca2+ antagonists and agonists in a stereoselective manner. Divalent cations (Mn2+ greater than Ca2+ greater than Mg2+) are inhibitory. Target size analysis with high-energy electrons was performed and the Mr of the verapamil drug receptor site is 110000.

Alamethicin↗
PubMed1984

Photoaffinity labelling of Ca2+ channels with [3H]azidopine.

A 1,4-dihydroypyridine arylazide photoaffinity ligand, [3H]azidopine (50.6 Ci/mmol), has been synthesized. [3H]Azidopine binds reversibly with a Kd of 350 pM to guinea-pig skeletal muscle membranes in the absence of ultraviolet light. The reversible [3H]azidopine binding is inhibited steroselectively by 1,4-dihydropyridines, phenylalkylamine Ca2+ channel blockers and La3+. Covalent incorporation into membrane proteins after photolysis was investigated by sodium dodecyl sulfate polyacrylamide slab gel electrophoresis. [3H]Azidopine is photoincorporated specifically into a protein of Mr approximately 145 000. The covalent labelling of the Mr approximately 145 000 band is inhibited stereoselectively by drugs and cations which block the reversible [3H]azidopine binding. It is suggested that [3H]azidopine is photoincorporated into a subunit of the putative Ca2+ channel.

Affinity Labels↗