Previous studies have shown that the skeletal muscle ryanodine receptor (RyR) is strongly inhibited by Mg²⁺ at levels within the normal cytosolic range (0.8-1 mM). This may explain why high levels of caffeine (40 mM) do not induce a maximal release of Ca²⁺ from the SR unless [Mg²⁺] is simultaneously reduced (Lamb, 2000).

In the present study we have investigated the effects of cytosolic Mg²⁺ on halothane-induced Ca²⁺ release from the SR. Rats (200-250 g) were humanely killed (Schedule 1). Single extensor digitorium longus (EDL) muscle fibres were mechanically skinned and perfused with solutions approximating to the intracellular milieu containing (mM): KCl, 100; Hepes, 25; EGTA, 0.15; phosphocreatine, 10; ATP, 5. The free [Ca²⁺] was 100 nM and the free [Mg²⁺] was 1 or 0.1 mM (pH 7.0, 22 °C). Changes in [Ca²⁺] within the fibre were detected using fura-2. Preparations were initially perfused with a solution containing 1 mM Mg²⁺. Following a 2 min Ca²⁺ loading period, application of 1 mM halothane failed to induce Ca²⁺ release from the SR. Indeed, in the presence of 1 mM Mg²⁺, levels of halothane as high as 20 mM did not induce Ca²⁺ release (n = 12). However, when [Mg²⁺] was reduced to 0.1 mM, application of 1 mM halothane induced a substantial release of Ca²⁺ from the SR. The amplitude of the halothane-induced fluorescence transient was 60 ± 9 % (n = 6, mean ± S.E.M.) of that induced by a maximal response to 40 mM caffeine. A similar release was obtained when [Mg²⁺] was reduced to 0.1 mM in the halothane-containing solution, while the [Mg²⁺] of the Ca²⁺ loading solution was maintained at 1 mM. Therefore, the effect of reducing [Mg²⁺] appears to involve facilitation of RyR activation by halothane, rather than an indirect action on the SR Ca²⁺ uptake mechanism.

These results suggest that even during deep anaesthesia or induction, when the [halothane] may reach 1.2 mM (Franks & Lieb, 1996), SR Ca²⁺ release is unlikely to be significant due to the potent inhibitory effect of cytosolic Mg²⁺ on the RyR. However, in malignant hyperthermia (MH) clinical levels of halothane or other volatile anaesthetics can induce a SR Ca²⁺ release, resulting in a sustained contracture. Interestingly, recent work suggests that inhibition of the RyR by Mg²⁺ is substantially reduced in porcine (Owen et al. 1997) and human (Duke & Steele, 2002) MH. The present study suggests that a decrease in Mg²⁺ inhibition may substantially increase the sensitivity of the RyR to halothane.

This work was supported by the The Wellcome Trust.

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