Ryanodine

Ryanodine is found naturally in the stem and roots of the plant Ryania speciosa. The ryanodine alkaloid was first isolated as a potential insecticide¹ but was found to act as a potent paralytic on skeletal and cardiac muscles.² Ryanodine inhibits SR Ca²⁺ release by binding with high affinity to the ryanodine receptor (RyR) channel as a tetrameric complex.^3,4

The RyR channel has three isoforms and is a poorly selective Ca²⁺ channel (selectivity Ca²⁺/K⁺ ~6) with very high conductance (~700 pS with K⁺ as charge carrier and ~100 pS with Ca²⁺ as charge carrier). The channel is regulated by Ca²⁺, Mg²⁺, ATP, and caffeine. Application of ryanodine locks the channel in a slow-gating subconductance state. Opening of the ryanodine-bound channel is long-lived and gives rise to a smaller unit current (~1/3 or ~1/2) than the control amplitude. This action of ryanodine on single RyR channel function is quite distinctive and is often used for functional identification of the channel.^5-8

Ryanodine binds with high affinity to RyR proteins in a pore located in the carboxyl terminus of the protein.⁹ Binding induces a complex change in single RyR channel function. This change is similar in all three RyR channel isoforms, inducing open events of very long duration and simultaneous reduction of ion conductance through the pore.^10,7 This dual impact on single-channel function (i.e. slower gating and reduced conductance) suggests a complicated mode of action. This is also supported by the rather complicated ryanodine dose dependency. Low doses of ryanodine (~10 nM) are reported to increase the frequency of single RyR channel openings to the normal conductance level. Intermediate ryanodine doses (~1 µM) are reported to induce the classical ryanodine action described above. High doses of ryanodine (~100 µM) are reported to lock the channel in a closed configuration.^11,10

It is generally assumed that each RyR monomer contains one high-affinity ryanodine binding site (K_D<50 nM).^6,12 It is suggested that cooperative interaction between RyR monomers is what generates the complexity.¹³ This is supported by ryanodine binding studies, which suggest the existence of additional lower affinity ryanodine binding sites.^6,13