Figure 5.

E189D increases the propensity for SOICR. Stable, inducible HEK293 cells expressing RyR2 wt or the E189D mutant were grown on glass coverslips. Cells were induced with 1 µg/ml tetracycline for 24 hours and loaded with 5 µM fura-2-AM in KRH buffer for 20 min at room temperature. Cells were perfused continuously with KRH buffer containing 0, 0.1, 0.2, 0.3, 0.5, 1.0 mM CaCl₂ or 1.0 mM CaCl₂ plus 5 mM caffeine. (A) Typical images of oscillating cells expressing RyR2 wt or E189D at various [Ca²⁺]. Oscillating cells are shown as red. (B) Typical recording of Fura-2 ratios from RyR2 wt (green trace) and E189D (red trace). (C) Percentage of cells expressing RyR2 wt (open circle) or E189D (closed triangle) oscillating at various [Ca²⁺]. Oscillation frequency of RyR2 wt and E189D at 1.0 mM CaCl₂ and the store Ca²⁺ content, determined from the amplitude of caffeine (5 mM) induced Ca²⁺ release. Values are normalized to RyR2 wt level (100%). Data shown are mean ± seM from 3 or 16 separate experiments.