Ca²⁺ buffering in the heart: Ca²⁺ binding to and activation of cardiac myofibrils

The measurement of cardiac Ca²⁺ transients using spectroscopic Ca²⁺ indicators is significantly affected by the buffering properties of the indicators. The aim of the present study was to construct a model of cardiac Ca²⁺ buffering that satisfied the kinetic constraints imposed by the maximum attainable rates of cardiac contraction and relaxation on the Ca²⁺ dissociation rate constants and which would account for the observed effects of 19F-NMR indicators on the cardiac Ca²⁺ transient in the Langendorff-perfused ferret heart. It is generally assumed that the Ca²⁺ dependency of myofibril activation in cardiac myocytes is mediated by a single Ca²⁺ binding site on troponin-C. A model based on 1:1 Ca²⁺ binding to the myofilaments, however, was unable to reproduce our experimental data, but a model in which we assumed ATP-dependent cooperative Ca²⁺ binding to the myofilaments was able to reproduce these data. This model was used to calculate the concentration and dissociation constant of the ATP-independent myofilament Ca²⁺ binding, giving 58µM and 2.0µM respectively. In addition to reproducing our experimental data on the concentration of free Ca²⁺ ions in the cytoplasm ([Ca²⁺]_i), the resulting Ca²⁺ and ATP affinities given by fitting of the model also provided good predictions of the Ca²⁺ dependence of the myofibrillar ATPase activity measured under in vitro conditions. Solutions to the model also indicate that the Ca²⁺ mobilized during each beat remains unchanged in the presence of the additional buffering load from Ca²⁺ indicators. The new model was used to estimate the extent of perturbation of the Ca²⁺ transient caused by different concentrations of indicators. As little as 10µM of a Ca²⁺ indicator with a dissociation constant of 200nM will cause 20% reduction in peak-systolic [Ca²⁺]_i and 30µM will cause ≈50% reduction in the peak-systolic [Ca²⁺]_i in a heart paced at 1.0Hz.