Paradoxical SR Ca2+ release in guinea-pig cardiac myocytes after β-adrenergic stimulation revealed by two-photon photolysis of caged Ca2+

In heart muscle the amplification and shaping of Ca2+ signals governing contraction are orchestrated by recruiting a variable number of Ca2+ sparks. Sparks reflect Ca2+ release from the sarcoplasmic reticulum (SR) via Ca2+ release channels (ryanodine receptors, RyRs). RyRs are activated by Ca2+ influx via L-type Ca2+ channels with a specific probability that may depend on regulatory mechanisms (e.g. beta-adrenergic stimulation) or diseased states (e.g. heart failure). Changes of RyR phosphorylation may be critical for both regulation and impaired function in disease. Using UV flash photolysis of caged Ca2+ and short applications of caffeine in guinea-pig ventricular myocytes, we found that Ca2+ release signals on the cellular level were largely governed by global SR content. During beta-adrenergic stimulation resting myocytes exhibited smaller SR Ca2+ release signals when activated by photolysis (62.3% of control), resulting from reduced SR Ca2+ content under these conditions (58.6% of control). In contrast, local signals triggered with diffraction limited two-photon photolysis displayed the opposite behaviour, exhibiting a larger Ca2+ release (164% of control) despite reduced global and local SR Ca2+ content. This apparent paradox implies changes of RyR open probabilities after beta-adrenergic stimulation, enhancing local regenerativity and reliability of Ca2+ signalling. Thus, our results underscore the importance of phosphorylation of RyRs (or of a related protein), as a regulatory physiological mechanism that may also provide new therapeutic avenues to recover impaired Ca2+ signalling during cardiac disease.