Effects of azumolene on Ryanodine binding to sarcoplasmic reticulum of normal and malignant hyperthermia susceptible swine skeletal muscles

Abstract

Dantrolene 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 malignant hyperthermia susceptible (MHS) swine muscles. Characteristics of [³H]ryanodine binding were clearly different between the two types of SR. Kinetic analysis of [³H]ryanodine binding to SR in the presence of 2 μM Ca²⁺ showed that association constant (K_ryanodine) is significantly higher in MHS than normal muscle SR (2.83 vs. 1.32×10⁷ M^‐1), whereas the maximal ryanodine binding capacity (B_max) is similar between the two types of SR. Addition of azumolene (e.g. 400μM) 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. 0.82×10² M^‐1). Addition of azumolene (e.g. 400 μM) decreased K_caffeine 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 Ca²⁺ release drug(s), and that azumolene can reverse the symptoms by reducing the drug affinity to Ca²⁺ release channels.

Key words:

• Calcium release channel
• Ryanodine receptor
• Dantrolene sodium
• Excitation‐contraction coupling
• Caffeine

Notes

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