Effects of phosphocreatine on SR Ca 2+ regulation in isolated saponin‐permeabilized rat cardiac myocytes

The effects of phosphocreatine (PCr) on sarcoplasmic reticulum (SR) Ca 2+ regulation were investigated in saponin‐permeabilized rat ventricular myocytes. Cells were perfused continuously with weakly Ca 2+ ‐buffered solutions approximating to the intracellular milieu. Ca 2+ release from the SR was detected using Fura‐2 or Fluo‐3. Withdrawal of PCr reduced the frequency of spontaneous Ca 2+ release by 12.8 ± 3.4 % ( n = 9 ) and the amplitude of the spontaneous Ca 2+ transient by 17.4 ± 3.1 % ( n = 9 ). Stepwise reductions in [PCr] progressively increased the time for the spontaneous Ca 2+ transient to rise from 25 to 100 % of the maximum value (TP75) and to fall by 75 % of the peak level (DT75). Following complete PCr withdrawal, the TP75 and the DT75 were 147.1 ± 13.2 and 174.8 ± 23.2 % of the control values, respectively. Experiments involving confocal microscopy showed that PCr withdrawal decreased the propagation velocity of spontaneous Ca 2+ waves. PCr withdrawal also reduced the frequency and amplitude, but increased the duration of spontaneous Ca 2+ sparks. Rapid application of 20 m m caffeine was used to assess the SR Ca 2+ content at the point of spontaneous Ca 2+ release. In the absence of PCr, the amplitude of the caffeine‐induced Ca 2+ transient was 18.4 ± 2.7 % ( n = 9 ) lower than in the presence of 10 m m PCr. This suggests that PCr withdrawal reduces the maximum SR Ca 2+ content that can be sustained before spontaneous Ca 2+ release occurs. These results suggest that local ADP buffering by PCr is essential for normal Ca 2+ regulation by the SR. Prolongation of the descending phase of the spontaneous Ca 2+ transient is consistent with a reduction in the efficiency of the SR Ca 2+ pump due to ADP accumulation. The fact that spontaneous Ca 2+ release occurs at a lower SR Ca 2+ content in the absence of PCr suggests that the Ca 2+ release mechanism may also be affected. These effects may be of relevance in circumstances where PCr depletion and Ca 2+ overload occur, such as myocardial ischaemia or anoxia.