Received for publication December 12, 2005.
Revised March 15, 2006.
Accepted for publication March 15, 2006.

The Interaction of an impermeant cation with the sheep Cardiac RyR Channel Alters Ryanoid Association with the channel

Abstract

In earlier 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 the tetraethylammonium (TEA⁺) as a blocker of K⁺ translocation (Tanna et al., 2001). In this communication we have 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-9-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-9-hydroxyryanodine with its receptor, whilst 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.

Key words: Ion channel regulation, Receptor binding studies, Single channel kinetics, Protein targets