Unambiguous observation of blocked states reveals altered, blocker-induced, cardiac ryanodine receptor gating

The flow of ions through membrane channels is precisely regulated by gates. The architecture and function of these elements have been studied extensively, shedding light on the mechanisms underlying gating. Recent investigations have focused on ion occupancy of the channel's selectivity filter and its ability to alter gating, with most studies involving prokaryotic K⁺ channels. Some studies used large quaternary ammonium blocker molecules to examine the effects of altered ionic flux on gating. However, the absence of blocking events that are visibly distinct from closing events in K⁺ channels makes unambiguous interpretation of data from single channel recordings difficult. In this study, the large K⁺ conductance of the RyR2 channel permits direct observation of blocking events as distinct subconductance states and for the first time demonstrates the differential effects of blocker molecules on channel gating. This experimental platform provides valuable insights into mechanisms of blocker-induced modulation of ion channel gating.