Abstract

In previous studies, we have demonstrated that the interaction of ryanoids with the sarcoplasmic reticulum Ca²⁺-release channel [ryanodine receptor (RyR)] incorporated into planar lipid bilayers reduced the effectiveness of tetraethylammonium (TEA⁺) as a blocker of K⁺ translocation (J Gen Physiol 117: 385-393, 2001). In the current study, we investigated both the effect of TEA⁺ on [³H]ryanodine binding and the actions of this impermeant cation on the interaction of the reversible ryanoid 21-amino-9α-hydroxyryanodine with individual, voltage-clamped RyR channels. A dose-dependent inhibition of [³H]ryanodine binding was observed in the presence of TEA⁺, suggesting that the cation and alkaloid compete for access to a common site of interaction. Single channel studies gave further insights into the mechanism of the competition between the two classes of ligands. TEA⁺ decreases the association rate of 21-amino-9α-hydroxyryanodine with its receptor, whereas the dissociation rate of the ryanoid from the channel was unaffected. Our results demonstrate that TEA⁺ inhibits both K⁺ translocation through RyR, and ryanoid interaction at the high affinity ryanodine site on the channel. These actions involve binding of TEA⁺ to different, but weakly interacting, sites in the RyR channel.