Diketopyridylryanodine Has Three Concentration-dependent Effects on the Cardiac Calcium-release Channel/Ryanodine Receptor

• Published in: The Journal of biological chemistry 2003-04, Vol.278 (16), p.14237-14248
• Main Authors: Bidasee, Keshore R., Xu, Le, Meissner, Gerhard, Besch, Henry R.
• Format: Article
• Language: English
• Subjects: Algorithms; Animals; Binding Sites; Calcium - metabolism; diketopyridylryanodine; Dogs; Dose-Response Relationship, Drug; Inhibitory Concentration 50; Kinetics; Ligands; Models, Chemical; Models, Molecular; Myocardium - metabolism; Protein Binding; Ryanodine - analogs & derivatives; Ryanodine - chemistry; Ryanodine - metabolism; Ryanodine - pharmacology; Ryanodine Receptor Calcium Release Channel - metabolism; Sarcoplasmic Reticulum - metabolism; Time Factors
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M208372200

By interacting with more than one site, ryanoids induce multiple effects on calcium-release channels. To date, the kinetics of interaction of only one of these sites has been characterized. Using C4,C12-diketopyridylryanodine in both [3H]ryanodine binding and single channel experiments we characterized another site on the cardiac ryanodine receptor (RyR2) with which ryanoids interact. Competitive binding of this ryanoid to RyR2 implied a minimal two-site binding model. At the single channel level, C4,C12-diketopyridylryanodine induced three distinct effects. At nanomolar concentrations, it increased channel open probability severalfold without inducing a subconductance. This effect was independent of membrane holding potential. As for other ryanoids, low micromolar concentrations of C4,C12-diketopyridylryanodine readily induced a subconductance state. The major subconductance had a current amplitude of 52% of fully open, it was reversible, and its time to induction and duration were voltage- and concentration-dependent, affording Hill slopes of >2. At higher micromolar concentrations C4,C12-diketopyridylryanodine induced long lasting, yet reversible shut states. Using a pharmacological strategy we have discerned an additional ryanoid-binding site on RyR2 that triggers an increase in channel activity. This site likely resides outside the strict confines of the transmembrane conducting pathway.