High-affinity binding of [3H]DTZ323 to the diltiazem-binding site of L-type Ca2+ channels

D-cis-[N-Methyl-H-3]-3-(acetyloxy)-5-[2-[[2-(3,4-dimethoxyphenyl)ethyl]-methylamino]ethyl]-2,3-dihydro-2-(4-methoxyphenyl)- 1,5-benzothiazepine-4(H-5)-one ([H-3]DTZ323), a novel 1,5-benzothiazepine radioligand, was characterized in a ligand-receptor binding study. Specific binding of [H-3]DTZ323 to rabbit skeletal muscle T-tubule membranes was saturable and reversible. Scatchard. analysis indicated a single binding site with a K-d value of 1.4 and 1.8 nM at 25 and 37 degreesC, respectively. DTZ323 and diltiazem derivatives inhibited specific [H-3]DTZ323 binding with a rank order of DTZ323>DTZ417 (quaternary ammonium derivative of DTZ323)>diltiazem>L-cis-DTZ323. The affinity of DTZ323 was 51 times higher than that of diltiazem. [H-3]DTZ323 binding was also completely inhibited by verapamil and tetrandrine, thus revealing the unique nature of the diltiazem-binding site. Specific [H-3]DTZ323 binding to crude guinea pig ventricular membranes was inhibited by diltiazem, DTZ323 and its derivatives with IC50 values close to those previously reported for the blockade of L-type Ca2+ channel currents. These results indicate that [H-3]DTZ323 is a potent and selective radioligand for the diltiazem-binding site of L-type Ca2+ channels. (C) 2003 Elsevier Science B.V All rights reserved.