Porcine malignant hyperthermia: Genotype and contractile threshold of immature muscles

Abstract

We investigated whether malignant hyperthermia (MH)-related contractile abnormalities, such as lowered contractile threshold, were expressed in MH-susceptible (MHS) immature muscles and myotubes. Muscles from neonatal piglets homozygous for Arg6I5 (normal) or for Cys6I5 (MHS) ryanodine receptor alleles, and heterozygotes were used. Intact cell bundles from piglet muscles generally were similar in contractile properties to adult muscles of the same genotype. Thresholds for K contractures in normal, heterozygous, and MHS piglet muscles (40 mmol/L, 25 mmol/L and 15 mmol/L K', respectively) differed significantly. Cultured myotubes were subjected to a series of square pulses of varying strengths (-50 to +50 mV) and durations (25-300 ms) using whole cell patch-clamp techniques. Threshold for contraction differed significantly among the three genotypes, for example, with 300 msec pulses thresholds were -6.9 5 0.9, -12.4 5 1.6, and -22.6 ? 2.6 mV for normal, heterozygous and MHS myotubes, respectively. Thus a significantly lower than normal threshold for contraction was expressed in MHS and heterozygous piglet muscles and myotubes. Further, these developmentally immature preparations are likely to express other differences characteristic of adult MHS muscles, and thus provide suitable preparations for clinically relevant studies of MHrelated cellular abnormalities.

A retrograde signal from RyR1 alters DHP receptor inactivation and limits window Ca 2+ release in muscle fibers of Y522S RyR1 knock-in mice

“…The question of whether, and if so how, MH mutations in RyR1 affect the two DHPR-controlled Ca 2ϩ fluxes has been assessed in myotubes derived from primary cultured satellite cells containing either native or expressed MH mutant RyR1 channels. A consistent finding from these studies was a lowered threshold for depolarization-induced activation of Ca 2ϩ release (6)(7)(8)(9).…”

The action of perchlorate on malignant-hyperthermia-susceptible muscle

“…A Twitch tension relative to control twitch (mean ± SE) for eight normal (l) and six MHS (q) muscle bundles was determined at various concentrations of perchlorate. for MHS (0.21 ± 0.02 normalized to tetanus) and normal bundles (0.13 ± 0.02 normalized to tetanus) were significantly different (P < 0.02) as reported previously [8,9]. Tetanic tension was not altered significantly by 20 mM perchlorate (data not shown).…”

“…(Muscles from MHS piglets have been previously shown to have in vitro responses which are similar to those of adult MHS muscle [2,9]). A small forelimb muscle (the lateral head of the common digital extensor) was quickly removed and placed in oxygenated porcine saline solution (PSS) containing (in mM): 135 NaCl, 4 KCl, 2.35 CaCl 2 , 0.85 MgCl 2 , 1 NaH 2 PO 4 , 18 NaHCO 3 , and 5.5 glucose, pH 7.2-7.3 when aerated with 5%CO 2 /95%O 2 .…”

Enhanced Excitation-coupled Calcium Entry in Myotubes Is Associated with Expression of RyR1 Malignant Hyperthermia Mutations

“…Its distinctive properties are: 1) it is initiated by depolarization; 2) it is dependent on the presence of both RyR1 and DHPR; 3) it is independent of the DHPR L-type Ca 2ϩ current; 4) it does not depend on SR Ca 2ϩ release; and 5) it can be effectively inhibited by La 3ϩ or SKF96365 (24). Previous studies of KCl responses in MH muscle cells showed significantly larger muscle contractures and lower KCl contracture thresholds in muscle fibers dissected from MH susceptible swine compared with WT (42)(43)(44). In addition, it has been reported that the total calcium transient in MH susceptible muscle fibers was larger than control fibers when exposed to subcontracture and contracture concentrations of KCl (30,43,45,46 This is most easily detectable during the sustained phase that follows the peak of the Ca 2ϩ transient.…”