Figure 8 – Increasing surface/inner CRU RyR expression ratio inhibits SCRs, has biphasic effects on DADs but does not affect SAP.

A) Increasing surface/inner CRU RyR expression ratio (i.e., increasing RyR density in surface CRUs without changing whole-cell RyR expression) monotonically increases the rate threshold of SCRs (i), whereas DADs (ii) display biphasic dependence and SAP (iii) remains absent in all conditions. The effect on SCRs is greater in cells with sparse and intermediate tubular structures vs cells with dense tubules, whereas the biphasic effect on DADs is less pronounced in cells with sparse tubular structures vs cells with intermediate and dense tubules. B) Effect of altered surface/inner CRU RyR expression ratio on voltage (i) and global cytosolic Ca²⁺ concentration (ii) in cells with sparse (left), intermediate (middle), and dense (right) tubules following pacing at 3 Hz to examine the occurrence of DADs and SCRs. C) Mechanism underlying increasing surface/inner CRU RyR expression ratio inhibiting SCRs, having biphasic effects on DADs but not affecting SAP. Biomarkers were determined from the first 100 ms of no-stimulation period and normalized to those of cells with a retained surface/inner CRU RyR expression ratio of 1.0. Increasing surface/inner CRU RyR expression ratio means more RyRs are located closer to NCX, resulting in increased NCX-RyR coupling. This is associated with increased NCX contribution to Ca²⁺ extrusion (i) and higher SCR-induced voltage changes/SCR amplitude ratio (ii). Enhanced NCX extrusion results in lower cleft Ca²⁺ concentration, smaller RyR P_o and leak leading to milder SCRs. While this in itself limits DADs, DAD likelihood is promoted by the increased SCR-induced changes in V_m, thus explaining the biphasic effects of increasing surface/inner CRU RyR expression ratio.