In cardiac cells, Ca2+ release flux (Jrel) via ryanodine receptors (RyRs) from the sarcoplasmic reticulum (SR) has a complex effect on the action potential (AP). Coupling between Jrel and the AP occurs via L-type Ca2+ channels (ICa) and the Na+/Ca2+ exchanger (INCX). We used a combined experimental and modelling approach to study interactions between Jrel, ICa and INCX in porcine ventricular myocytes. We tested the hypothesis that during normal uniform Jrel, 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. ICa and INCX were partitioned into subpopulations in the T-space and M-space sarcolemma. We denoted free Ca2+ concentrations in T-space and M-space Cat and Cam, respectively. Experiments were designed to allow separate measurements of ICa and INCX as a function of Jrel. Inclusion of T-space in the model allowed us to reproduce in silico the following important experimental results: (1) hysteresis of INCX dependence on Cam; (2) delay between peak INCX and peak Cam during caffeine application protocol; (3) delay between INCX and Cam during Ca2+-induced-Ca2+-release; (4) rapid ICa inactivation (within 2 ms) due to Jrel, with magnitude graded as a function of the SR Ca2+ content; (5) time delay between ICa inactivation due to Jrel and Cam. Partition of 25% NCX in T-space and 75% in M-space provided the best fit to the experimental data. Measured Cam and ICa or INCX were used as input to the model for estimating Cat. The actual model-computed Cat, obtained by simulating specific experimental protocols, was used as a gold standard for comparison. The model predicted peak Cat in the range of 6-25 μm, with time to equilibrium of Cat with Cam of ~350 ms. These Cat 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 ICa-based estimation method was most accurate during the ascending phase of Cat. The INCX-based method provided a good estimate for the descending phase of Cat. Thus, application of both methods in combination provides the best estimate of the entire Cat time course.
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