Effects of Azumolene on Ryanodine Binging to Sarcoplasmic Reticulum of Normal and Malignant Hyperthermia Sucseptible Swine Skeletal Muscles

DOantrolene is a primary specific therapeutic drug for prevention and treatment of malignant hyperthermia symptoms. The mechanisms underlying the therapeutic effects of the drug are not well understood. The present study aimed at the characterization of the effects of azumolene, a water soluble dantrolene analogue, on ryanodine binding to sarcoplasmic reticulum (SR) from normal and malign::lnt hyperthermia susceptible (MHS) swine muscles. Characteristics of $[^3H]ryanodine$ binding were clearly different between the two types of SR. Kinetic analysis of eH]ryanodine binding to SR in the presence of $2{\mu}M$ $Ca^{2+}$ showed that association constant $(K_{ryanodine}_7$ is significantly higher in MHS than normal muscle SR $(2.83 vs. 1.32{\times}10^7 M^{-1}$, whereas the maximal ryanodine binding capacity $(B_{max})$ is similar between the two types of SR. Addition of azumolene $(e.g. 400{\mu}M)$ did not significantly alter both $K_{ryanodine}$ and $B_{max}$ of $[^3H]$ryanodine binding in both types of SR, indicating that the azumolene effect was not on the ryanodine binding sites. Addition of caffeine activated $[^3H]$ ryanodine binding in both types of SR, and caffeine sensitivity was significantly higher in MHS muscle SR than normal muscle SR $(K_{caffeine}:3.24 vs. 0.82 {\times} 10^2 M^{-l}). Addition of azumolene $(e.g.400{\mu}M)$ decreased Kcaffeine without significant change in $B_{max}$ in both types of SR suggesting that azumolene competes with caffeine binding site(s). These results suggest that malignant hyperthermia symptoms are caused at least in part by greater sensitivity of the MHS muscle SR to the $Ca^{2+}$ release drug(s), and that azumolene can reverse the symptoms by reducing the drug affinity to $Ca^{2+}$ release channels.