Specific binding of [3H]LY186126, an analogue of indolidan (LY195115), to cardiac membranes enriched in sarcoplasmic reticulum vesicles.
LY186126 was found to be a potent inhibitor of type IV cyclic AMP phosphodiesterase located in the sarcoplasmic reticulum of canine cardiac muscle. This compound, a close structural analogue of indolidan (LY195115), was prepared in high specific activity, tritiated form to study the positive inotropic receptor(s) for cardiotonic phosphodiesterase inhibitors such as indolidan and milrinone. A high-affinity binding site for [3H]LY186126 was observed (Kd = 4 nM) in purified preparations of canine cardiac sarcoplasmic reticulum vesicles. Binding was proportional to vesicle protein, was inactivated by subjecting membranes to proteolysis or boiling, and was dependent on added Mg2+. Scatchard analysis suggested the presence of a single class of binding sites in the membrane preparation. Indolidan, milrinone, and LY186126 (all at 1 microM) produced essentially complete displacement of bound [3H]LY186126, while nifedipine, propranolol, and prazosin had little or no effect at this concentration. This represents the first reported use of a radioactive analogue to label the inotropic receptor for cardiotonic phosphodiesterase inhibitors. The results suggest that [3H]LY186126 is a useful radioligand for examining the subcellular site(s) responsible for positive inotropic effects of these drugs.
Kauffman
R F
RF
Department of Cardiovascular Pharmacology, Eli Lilly and Co., Indianapolis, Indiana 46285.
Utterback
B G
BG
Robertson
D W
DW
D000818:Animals; D001665:Binding Sites; D002316:Cardiotonic Agents / Q000378:metabolism*; D004285:Dogs; D007211:Indoles / Q000378:metabolism*; D008861:Microsomes / Q000378:metabolism; D009206:Myocardium / Q000201:enzymology / Q000378:metabolism*; D000078183:Oxindoles; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D010727:Phosphoric Diester Hydrolases / Q000032:analysis; D011724:Pyridazines / Q000378:metabolism*; D012519:Sarcoplasmic Reticulum / Q000201:enzymology / Q000378:metabolism*; D014316:Tritium
D002316:Cardiotonic Agents; D007211:Indoles; D000078183:Oxindoles; D010726:Phosphodiesterase Inhibitors; D011724:Pyridazines; C050325:indolidan; D014316:Tritium; C059371:LY 186126; D010727:Phosphoric Diester Hydrolases