EC-coupling and calcium (Ca) signaling in cardiac muscle relies on Ca-induced Ca release (CICR) from the sarcoplasmic reticulum (SR), an amplification process exhibiting a high degree of positive feedback. To avoid instabilities of this feedback system, Ca release is highly localized to elementary Ca signaling events, Ca sparks. The amplitude of the cellular Ca transient can be modulated by recruiting fewer or more Ca sparks, each of which normally remains under local control by a L-type Ca channel located in the sarcolemma. For proper relaxation between cardiac beats and for the system to be stable, the Ca release process itself has to terminate. However, the processes limiting Ca release from the SR and terminating Ca sparks are still poorly understood. We used UV-laser flash photolysis and two-photon excitation photolysis of caged Ca (DM-nitrophen) to activate CICR in isolated cardiac myocytes with a paired pulse protocol, while simultaneously recording Na-Ca exchange currents with he patch-clamp technique or Ca concentration with fluo-3 and a confocal microscope. Using this combination of techniques, we observed a genuine refractoriness of global CICR that recovered slowly (within about 1 second (DelPrincipe et al. 1999)). Based on discrepancies between global refractoriness and fast recovery of local refractoriness we had proposed that the SR Ca content may be an important regulator of RyR Ca sensitivity and that recovery may be related to SR refilling after a release. To test our hypothesis that SR Ca refilling via the SR Ca pump (SERCA) may be rate limiting for recovery from refractoriness, we used pharmacological tools to stimulate or slow down the pump (isoproterenol, thapsigargin or CPA). Indeed, isoproterenol accelerated and CPA slowed recovery from refractoriness. From these results we conclude that refilling of the SR via the SERCA is the rate limiting step for recovery from refractoriness. Most likely, this refractoriness results from functional Ca depletion of the SR during release, leading to deactivation and termination of the Ca release by desensitization of the RyRs for trigger Ca on the cytosolic side of the channel (Sobie et al. 2002; Terentyev et al. 2003). This raises the question, how the SR Ca content or free SR Ca concentration can be detected by the EC-coupling machinery. A Ca receptor is required in the lumen of the SR, and a pathway signaling information about SR Ca content back to the RyR. This pathway might involve recently identified small accessory proteins located in the dyadic region.