Ruthenium red modifies the cardiac and skeletal muscle Ca2+ release channels (ryanodine receptors) by multiple mechanisms

The effects of ruthenium red (RR) on the skeletal and cardiac muscle ryanodine receptors (RyRs) were studied in vesicle-Ca2+ flux, [H-3]ryanodine binding, and single channel measurements. In vesicle-Ca2+ flux measurements, RR was more effective in inhibiting RyRs at 0.2 mu M than 20 mu M free Ca2+. [H-3]Ryanodine binding measurements suggested noncompetitive interactions between RR inhibition and Ca2+ regulatory sites of RyRs. In symmetric 0.25 M KCl. with 10-20 mu M cytosolic Ca2+, cytosolic RR decreased single channel activities at positive and negative holding potentials. In close to fully activated skeletal (20 mu M Ca2+ + 2 mM ATP) and cardiac (200 mu M Ca2+) RyRs, cytosolic RR induced a predominant subconductance at a positive but not negative holding potential. Lumenal RR induced a major subconductance in cardiac RyR at negative but not positive holding potentials and several subconductances in skeletal RyR. The RR-related subconductances of cardiac RyR showed a nonlinear voltage dependence, and more than one RR molecule appeared to be involved in their formation, Cytosolic and lumenal RR also induced subconductances in Ca2+-conducting skeletal and cardiac RyRs recorded at 0 mV holding potential. These results suggest that RR inhibits RyRs and induces subconductances by binding to cytosolic and lumenal sites of skeletal and cardiac RyRs.