Magnesium Ions Moderate Calcium-Induced Calcium Release in Cardiac Calcium Release Sites by Binding to Ryanodine Receptor Activation and Inhibition Sites

Figures

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  FIGURE 1. The geometric model of the calcium release site. (A) The arrangement of ryanodine receptors (RyRs) in the model CRS. The empty squares with dots represent RyRs with a central ion channel, the gated point source of Ca2+ current. (B) The distribution of RyR vicinities in the model CRS. The box diagram shows the 25% and 75% percentile, whiskers show minimum and maximum, the black square depicts the mean, and the horizontal line represents the median.

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  FIGURE 2. The gating scheme of the MWC-Ca/Mg RyR model. The scheme depicts transitions of four allosterically coupled monomeric activation sites and one common inhibitory site of the RyR channel. The competitive binding of Ca2+ and Mg2+ ions to RyR activation sites is shown by horizontal (thin lines) and vertical (thick lines) transitions, respectively. The RyR open/close transitions (dashed lines) are independent of ion binding/unbinding. The RyR inhibitory site binds two Mg2+ ions simultaneously and switches the RyR channel to a non-conducting state without affecting the activation sites. Squares – closed states; Circles – open states; Gray – inhibited states. The three numerals in the symbols denote (from left to right): the number of Ca2+ ions bound to the activation site; the number of Mg2+ ions bound to the activation site; and the number of Mg2+ ions bound to the inhibition site. All RyR transitions between states are reversible and obey the principle of detailed balance.

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  FIGURE 3. Comparison of the MWC-Ca/Mg RyR model predictions with experimental data. The model predictions (lines) were calculated using Eqs. 14 (A), 2 (B), and 4 (C) and the reference set of parameter values (Table 1) for the same ion conditions as the experimental data (points). (A) The calcium dependence of open probability for 0 mM Mg2+ and 0.5 mM total ATP (Tencerova et al., 2012) (black), and for 0.6, 0.94, and 1.33 mM free Mg2+ (blue, red, and green, respectively) and 3 mM total ATP (Zahradnikova et al., 2003). (B) The calcium dependence of mean open time for 0 mM total Mg2+ and 0.5 mM ATP (Tencerova et al., 2012) (black), for 0.6 mM free Mg2+, 3 mM total ATP (Zahradnikova et al., 2003) (blue), and for 1 mM free Mg2+ and 2 mM total ATP (Cannell et al., 2013) (red). (C) The simulated time course of RyR responses to a step Ca2+ increase from 100 nM to 20 μM in the absence of Mg2+ (black), and from 1.5 to 10 μM Ca2+ in the presence of 0.6 and 0.94 mM free Mg2+ and 3 mM total ATP (blue and red lines, respectively). The 63.2% activation time is indicated by diamonds. (D) Comparison of the calculated (diamonds) and experimental (dots with error bars) 63.2% activation times obtained under…

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  FIGURE 4. Characteristics of calcium release events (CREs) simulated with the MWC-Ca/Mg RyR model and the reference set of parameters. (A) Top panel – the time courses of simulated CREs representing a typical quark, blip, and spark for iCa = 0.6 pA; Bottom panel – the fluorescence calcium signals emulating laser line-scan images in a confocal microscope corresponding to CREs in panel (A). (B) The distribution of the peak number of open RyRs, NOp⁢e⁢a⁢k, in the simulated calcium release events. The red, blue, and green columns correspond to peaks of respective traces shown in panel (A). (C) The surface plot of the time to peak vs. NOp⁢e⁢a⁢k; dotted lines indicate the NOp⁢e⁢a⁢k values used for discrimination between quarks, blips, and sparks. (D) Relationships between iCa values and characteristics of simulated quarks, blips, and sparks; nCa – the amount of Ca2+ released per CRE.

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  FIGURE 5. Simulated dependence of RyR single-channel characteristics on Mg2+ binding. PO([Ca2+]), tO([Ca2+]), tC([Ca2+]), and PO(t) were calculated with Eqs. 14, 2, 2, and 4, respectively. (A) Variation of Mg-binding to the RyR activation site. (B) Variation of Mg-binding to the RyR inhibitory site. Bottom panels: time courses of PO evolution after a stepwise increase of [Ca2+] from 0.1 to 20 μM. Color-coding is described in the legend. The conditions that had the same effect are shown in solid, dashed, or dotted lines. Note the logarithmic scale of PO and tC.

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  FIGURE 6. The effect of Mg-binding parameters on CREs. Simulations were performed at 100 nM Ca2+ and iCa of 0.6 pA. (A,D) The effect of Mg-binding parameters at the activation site on the time course (A) and the fraction (D) of CREs. (B,E) The effects of the allosteric coefficient fMg on the time course (B) and the fraction (E) of CREs. (C,F) The effect of Mg-binding parameters at the inhibition site on the time course (C) and the fraction (F) of CREs. Individual conditions are color-coded as indicated in the legend.

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  FIGURE 7. Calcium release event characteristics related to the effective coupling strength. (A) The whole dataset (14240 points per panel) for various Mg-binding parameters of the RyR activation site (orange and pink) and the RyR inhibition site (blue), which were obtained for all RyR vicinities and all single-channel calcium currents (not indicated). The dots are semitransparent to better visualize the data. The circles, triangles, and diamonds mark results for decreased (×0.5; lighter color) or increased (×2; darker color) Mg-binding parameter values (see legend), relative to the reference parameter values (black pentagons). The POmax colormap refers solely to the NO⁢Sp⁢e⁢a⁢k graph to indicate the relation of the NO⁢Sp⁢e⁢a⁢k increase to POmax increase when varying Mg-binding at the inhibitory site, not present in the case of blips. (B) The dependence of the fraction of quarks, blips, and sparks on the reciprocally changed rate constants kAonMg (top axis) and kAoffMg (bottom axis), so that the equilibrium constant KAMg stayed at the reference value of 92 μM. Squares, circles, up triangles and down triangles denote values of iCa = 0.15, 0.25, 0.4, and 0.6 pA, respectively.