cADP ribose and [Ca²⁺](i) regulation in rat cardiac myocytes

cADP ribose (cADPR)-induced intracellular Ca²⁺ concentration ([Ca²⁺](i)) responses were assessed in acutely dissociated adult rat ventricular myocytes using real-time confocal microscopy. In quiescent single myocytes, injection of cADPR (0.1-10 μM) induced sustained, concentration-dependent [Ca²⁺](i) responses ranging from 50 to 500 nM, which were completely inhibited by 20 μM 8-amino-cADPR, a specific blocker of the cADPR receptor. In myocytes displaying spontaneous [Ca²⁺](i) waves, increasing concentrations of cADPR increased wave frequency up to ~ 31250% of control. In electrically paced myocytes (0.5 Hz, 5-ms duration), cADPR increased the amplitude of [Ca²⁺](i) transients in a concentration-dependent fashion, up to 150% of control. Administration of 8-amino-cADPR inhibited both spontaneous waves as well as [Ca²⁺](i) responses to electrical stimulation, even in the absence of exogenous cADPR. However, subsequent [Ca²⁺](i) responses to 5 mM caffeine were only partially inhibited by 8-amino-cADPR. In contrast, even under conditions where ryanodine receptor (RyR) channels were blocked with ryanodine, high cADPR concentrations still induced an [Ca²⁺](i) response. These results indicate that in cardiac myocytes, cADPR induces Ca²⁺ release from the sarcoplasmic reticulum through both RyR channels and via mechanisms independent of RyR channels.