(-)-[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.
D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D004110:Diltiazem / Q000378:metabolism; D006168:Guinea Pigs; D007473:Ion Channels / Q000378:metabolism*; D007700:Kinetics; D009132:Muscles / Q000378:metabolism*; D009539:Nicotinic Acids / Q000378:metabolism; D009553:Nimodipine; D013237:Stereoisomerism; D013347:Subcellular Fractions / Q000378:metabolism; D014700:Verapamil / Q000031:analogs & derivatives* / Q000378:metabolism
D007473:Ion Channels; D009539:Nicotinic Acids; D009553:Nimodipine; C043184:4-desmethoxyverapamil; D014700:Verapamil; D004110:Diltiazem; D002118:Calcium