Enhanced Ca²⁺-induced calcium release by isolated sarcoplasmic reticulum vesicles from malignant hyperthermia susceptible pig muscle

To further define the possible involvement of sarcoplasmic reticulum calcium accumulation and release in the skeletal muscle disorder malignant hyperthermia (MH), we have examined various properties of sarcoplasmic reticulum fractions isolated from normal and MH-susceptible pig muscle. A sarcoplasmic reticulum preparation enriched in vesicles derived from the terminal cisternae, was further fractionated on discontinuous sucrose density gradients (Meissner, G. (1984) J. Biol. Chem. 259, 2365-2374). The resultant MH-susceptible and normal sarcoplasmic reticulum fractions, designated F₀ F₄, did not differ in yield, cholesterol and phospholipid content, or nitrendipine binding capacity. Calcium accumulation (0.27 μmol Ca/mg per min at 22°C), Ca²⁺-ATPase activity (0.98 μmol P_i/mg per min at 22°C), and calsequestrin content were also similar for MH-susceptible and normal sarcoplasmic reticulum fraction F₃. To examine sarcoplasmic reticulum calcium release, fraction F₃ vesicles were passively loaded with ⁴⁵Ca (approx. 40 nmol Ca/mg), and rapidly diluted into a medium of defined Ca²⁺ concentration. Upon dilution into 1 μM Ca²⁺, the extent of Ca²⁺-dependent calcium release measured after 5 s was significantly greater for MH-susceptible than for normal sarcoplasmic reticulum, 65.9 ± 2.8% vs. 47.7 ± 3.9% of the loaded calcium, respectively. The C _{1 2} for Ca²⁺ stimulation of this calcium release (5 s value) from MH-susceptible sarcoplasmic reticulum also appeared to be shifted towards a higher Ca²⁺-sensitivity when compared to normal sarcoplasmic reticulum. Dantrolene had no effect on calcium release from fraction F₃, however, halothane (0.1-0.5 mM) increased the extent of calcium release (5 s) similarly in both MH-susceptible and normal sarcoplasmic reticulum. Furthermore, Mg²⁺ was less effective at inhibiting, while ATP and caffeine were more effective in stimulating, this Ca²⁺-dependent release of calcium from MH-susceptible, when compared to normal sarcoplasmic reticulum. Our results demonstrate that while sarcoplasmic reticulum calcium-accumulation appears unaffected in MH, aspect(s) of the sarcoplasmic reticulum Ca²⁺-induced calcium release mechanism are altered. Although the role of the Ca²⁺-induced calcium release mechanism of sarcoplasmic reticulum in situ is not yet clear, our results suggest that an abnormality in the regulation of sarcoplasmic reticulum calcium release may play an important role in the MH syndrome.