Quenching the spark: Termination of CICR in the submicroscopic space of the dyad

Figure 1.. Possible CICR termination mechanisms. (A) SR Ca2+ depletion. A reduction in SR Ca2+ levels will reduce release flux regardless of RyR gating (as described by their open probability, PO). However, fateful termination by this mechanism alone is problematic, because the jSR lumen is continually refilled from the rest of the SR. (B) SR luminal control of RyR gating may be modulated either by a direct effect on the RyR itself (red) and/or via an accessory protein such as CSQ (green). However, the extent to which these mechanisms could reduce RyR PO sufficiently to terminate release is unclear. (C) Stochastic attrition. If all RyRs close simultaneously, then the release flux is terminated. However, it is unlikely that this will occur within the timescale of a Ca2+ spark. Stochastic attrition could be accelerated by coupled gating between RyRs, either by direct contact or by a protein linker (X). (D) Induction decay. After CRU activation, jSR Ca2+ levels decline which results in a decreasing release flux. The local cytoplasmic Ca2+ level is proportional to the release flux, and this is transduced via the steep Ca2+ dependence of the RyR closed time. As the closed time becomes longer, it…

Figure 2.. A method for calculating jSR release fluxes from spark records. (A) A simplified diagram illustrating the relative sizes of the T-tubules, jSR, network SR (nSR), myofilaments (myo), and a typical microscope PSF that blurs the Ca2+ signals. (B) By using spherical geometry for the model, the computational complexity is greatly reduced while still retaining the volume fractions and concentrations of each component that contribute to Ca2+ fluxes. Effectively, the spatial geometries of each component are spread over each computational compartment, making transport from the jSR isotropic at the length scale of interest (200 nm). The center element is dedicated to the dyadic space (including the jSR with CSQ; Kong et al., 2013), and all other compartments include reaction and diffusion fluxes in the cytosol and SR as indicated in the bottom part of the figure. Having calculated the spatial Fluo-4 and Fluo-5N signals, these are blurred by convolution with the microscope PSF, and the resulting signal is fitted to typical Ca2+ sparks by altering the time course of the jSR permeability change basis function (Kong et al., 2013). Because large numbers of simulations are performed to allow the…

Figure 3.. Estimating jSR Ca2+ depletion and RyR gating time course by flux reconstructions. (A) The 3-D model of Ca2+ reactions, diffusion, and microscope blurring shown in Fig. 2 generates a Ca2+ spark record, which is fitted to experimental data by varying a basis function for the jSR permeability time course (Kong et al., 2013). (B) Although not fitted, the model can also calculate the corresponding SR Ca2+ depletion signals, which are similar to those recorded experimentally (Brochet et al., 2005). (C) From the calculated release flux (red) and the jSR Ca2+ levels, the jSR permeability time course was derived (normalized flux in red and normalized nPo [number of open channels] in blue). (D) Normalized release flux (red) and permeability (blue) changes from the induction decay model of Cannell et al. (2013). Note the concordance of flux estimates by both models, although the permeability time course is dependent on species-dependent RyR Ca2+ sensitivity.