Inhibition of Ca 2+ Sparks by Ruthenium Red in Permeabilized Rat Ventricular Myocytes

Abstract

We have compared the effects of the sarcoplasmic reticulum (SR) Ca 2+ release inhibitor, ruthenium red (RR), on single ryanodine receptor (RyR) channels in lipid bilayers, and on Ca 2+ sparks in permeabilized rat ventricular myocytes. Ruthenium red at 5 μM inhibited the open probability ( P o) 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 Ca 2+ 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 Ca 2+ spark formation, this reduction in Ca 2+ spark amplitude corresponds to an ∼4-fold decrease in Ca 2+ release flux underlying Ca 2+ sparks. Ruthenium red (5 μM) increased the SR Ca 2+ content by ∼2-fold (from 151 to 312 μmol/l cytosol). Considering the degree of inhibition of local Ca 2+ release events, the increase in SR Ca 2+ load by RR, and the lack of effects of RR on single RyR open time and conductance, we have estimated that Ca 2+ sparks under normal conditions are generated by openings of at least 10 single RyRs.