Identification of Changes in the Functional Profile of the Cardiac Ryanodine Receptor Caused by the Coupled Gating Phenomenon

Abstract

The objective of this work was to identify and further characterize potential changes in the functional profile of the cardiac ryanodine receptor (RyR2) channel caused by the coupled gating phenomenon. By reconstituting an ion channel into a planar lipid membrane, we showed that coupled RyR2 channels were activated by cytosolic Ca²⁺ with similar efficacy and potency as reported for the single RyR2 channel. In contrast, all examined parameters of gating kinetics were affected by the functional interaction between channels. Ignoring brief closings during main open events, the average open and closed times were considerably prolonged and the frequency of opening was reduced. Interestingly, when luminal Ca²⁺ was used as a charge carrier, Ca²⁺-activated coupled RyR2 channels did not exhibit a sudden switch from slow to fast gating kinetics at an open probability of 0.5 as reported for the single RyR2 channel. Regarding flicker gating, the average closed time was significantly shorter and the frequency of closing was greatly enhanced. Furthermore, in contrast to the single RyR2 channel, both parameters for coupled channels were independent of cytosolic Ca²⁺. Selected permeation properties of coupled RyR2 channels were comparable to those found for the single RyR2 channel. The Ca²⁺ current amplitude-luminal Ca²⁺ relationship displayed a simple saturation and the channel selectivity for Ba²⁺ and Ca²⁺ ions was similar. Our results suggest that the major targets influenced by coupled gating are likely the gates of individual RyR2 channels recruited into a functional complex, thus ensuring the correlation of Ca²⁺ fluxes.