Adaptation of single cardiac ryanodine receptor channels

Single cardiac ryanodine receptor (RyR) channel adaptation was previously defined with Ca2+ stimuli produced by flash photolysis of DM-nitrophen (caged-Ca+2). Photolysis of DM-nitrophen induced a very fast Ca+2 overshoot (Ca+2 spike) at the leading edge of the Ca+2 stimuli. It has been suggested that adaptation (tau approximately 1.3 s) may reflect Ca+2 slowly coming off the RyR Ca+2 activation sites following the faster Ca+2 spike (tau approximately 1 ms). This concern was addressed by defining the Ca2+ deactivation kinetics of single RyR channels in response to a rapid reduction in free Ca2+ concentration ([Ca2+]FREE). The [Ca2+]FREE was lowered by photolysis of Diazo-2. Single RyR channels deactivated (tau approximately 5.3 ms) quickly in response to the photolytically induced [Ca2+]FREE reduction. Improved estimates of the Ca2+ spike time course indicate that the Ca2+ spike is considerably faster (10-100-fold) than previously thought. Our data suggest that single RyRs are not significantly activated by fast Ca2+ spikes and that RyR adaptation is not due to deactivation following the fast Ca2+ spike. Thus, RyR adaptation may have an important impact on Ca2+ signaling in heart.