Inhibition of Ca²⁺ sparks by ruthenium red in permeabilized rat ventricular myocytes

We have compared the effects of the sarcoplasmic reticulum (SR) Ca²⁺ release inhibitor, ruthenium red (RR), on single ryanodine receptor (RyR) channels in lipid bilayers, and on Ca²⁺ sparks in permeabilized rat ventricular myocytes. Ruthenium red at 5 μM inhibited the open probability (Po) of RyRs ~20-50-fold, without significantly affecting the conductance or mean open time of the channel. At the same concentration, RR inhibited the frequency of Ca²⁺ sparks in permeabilized myocytes by ~10-fold, and reduced the amplitude of large amplitude events (with most probable localization on the line scan) by ~3-fold. According to our theoretical simulations, performed with a numerical model of Ca²⁺ spark formation, this reduction in Ca²⁺ spark amplitude corresponds to an ~4-fold decrease in Ca²⁺ release flux underlying Ca²⁺ sparks. Ruthenium red (5 μM) increased the SR Ca²⁺ content by ~2-fold (from 151 to 312 μmol/l cytosol). Considering the degree of inhibition of local Ca²⁺ release events, the increase in SR Ca²⁺ load by RR, and the lack of effects of RR on single RyR open time and conductance, we have estimated that Ca²⁺ sparks under normal conditions are generated by openings of at least 10 single RyRs.