Data-based theoretical identification of subcellular calcium compartments and estimation of calcium dynamics in cardiac myocytes

In cardiac cells, Ca²⁺ release flux (J_rel) via ryanodine receptors (RyRs) from the sarcoplasmic reticulum (SR) has a complex effect on the action potential (AP). Coupling between J_rel and the AP occurs via L-type Ca²⁺ channels (I_Ca) and the Na⁺/Ca²⁺ exchanger (I_NCX). We used a combined experimental and modelling approach to study interactions between J_rel, I_Ca and I_NCX in porcine ventricular myocytes. We tested the hypothesis that during normal uniform J_rel, the interaction between these fluxes can be represented as occurring in two myoplasmic subcompartments for Ca²⁺ distribution, one (T-space) associated with RyR and enclosed by the junctional portion of the SR membrane and corresponding T-tubular portion of the sarcolemma, the other (M-space) encompassing the rest of the myoplasm. I_Ca and I_NCX were partitioned into subpopulations in the T-space and M-space sarcolemma. We denoted free Ca²⁺ concentrations in T-space and M-space Ca_t and Ca_m, respectively. Experiments were designed to allow separate measurements of I_Ca and I_NCX as a function of J_rel. Inclusion of T-space in the model allowed us to reproduce in silico the following important experimental results: (1) hysteresis of I_NCX dependence on Ca_m; (2) delay between peak I_NCX and peak Ca_m during caffeine application protocol; (3) delay between I_NCX and Ca_m during Ca²⁺-induced-Ca²⁺-release; (4) rapid I_Ca inactivation (within 2 ms) due to J_rel, with magnitude graded as a function of the SR Ca²⁺ content; (5) time delay between I_Ca inactivation due to J_rel and Ca_m. Partition of 25% NCX in T-space and 75% in M-space provided the best fit to the experimental data. Measured Ca_m and I_Ca or I_NCX were used as input to the model for estimating Ca_t. The actual model-computed Ca_t, obtained by simulating specific experimental protocols, was used as a gold standard for comparison. The model predicted peak Ca_t in the range of 6-25 μm, with time to equilibrium of Ca_t with Ca_m of ~350 ms. These Ca_t values are in the range of LCC and RyR sensitivity to Ca²⁺. An increase of the SR Ca²⁺ load increased the time to equilibrium. The I_Ca-based estimation method was most accurate during the ascending phase of Ca_t. The I_NCX-based method provided a good estimate for the descending phase of Ca_t. Thus, application of both methods in combination provides the best estimate of the entire Ca_t time course.