Coupled gating modifies the regulation of cardiac ryanodine receptors by luminal Ca(2+)

Cardiac ryanodine receptors (RYR2s) infrequently exhibit coupled gating that is manifested by synchronous opening and closing. To better characterize this phenomenon, we investigated the regulation of coupled RYR2 channels by luminal Ca(2+) focusing on effects that are likely mediated by the true luminal activation mechanism. By reconstituting an ion channel into a planar lipid bilayer and using substantially lower concentration of luminal Ba(2+) (8mM, the virtual absence of Ca(2+)) and luminal Ca(2+) (8mM), we show that response of coupled RYR2 channels to caffeine at a diastolic cytosolic Ca(2+) (90nM) was affected by luminal Ca(2+) in a similar manner as for the single RYR2 channel except the gating behavior. Whereas, the single RYR2 channel responded to luminal Ca(2+) by prolongation in open and closed times, coupled RYR2 channels seemed to be resistant in this respect. In summary, we conclude that the class of Ca(2+) sites located on the luminal face of coupled RYR2 channels that is responsible for the channel potentiation by luminal Ca(2+) is functional and not structurally hindered by the channel coupling. Thus, the idea about non-functional luminal Ca(2+) sites as a source of the apparent gating resistance of coupled RYR2 channels to luminal Ca(2+) appears to be ruled out.