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

Abstract

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 co-operative 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 58 and 2.0 μM 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 10 μM of a Ca2+ indicator with a dissociation constant of 200 nM will cause a 20% reduction in peak-systolic [Ca2+]i and 30 μM will cause approx. 50% reduction in the peak-systolic [Ca2+]i in a heart paced at 1.0 Hz.