Enhanced Excitation-Coupled Calcium Entry in Myotubes Expressing Malignant Hyperthermia Mutation R163C Is Attenuated by Dantrolene

Abstract

Dantrolene is the drug of choice for the treatment of malignant hyperthermia (MH) and is also useful for treatment of spasticity or muscle spasms associated with several clinical conditions. The current study examines the mechanisms of dantrolene's action on skeletal muscle and shows that one of dantrolene's mechanisms of action is to block excitation-coupled calcium entry (ECCE) in both adult mouse flexor digitorum brevis fibers and primary myotubes. A second important new finding is that myotubes isolated from mice heterozygous and homozygous for the ryanodine receptor type 1 R163C MH susceptibility mutation show significantly enhanced ECCE rates that could be restored to those measured in wild-type cells after exposure to clinical concentrations of dantrolene. We propose that this gain of ECCE function is an important etiological component of MH susceptibility and possibly contributes to the fulminant MH episode. The inhibitory potency of dantrolene on ECCE found in wild-type and MH-susceptible muscle is consistent with the drug's clinical potency for reversing the MH syndrome and is incomplete as predicted by its efficacy as a muscle relaxant.

Malignant hyperthermia susceptibility arising from altered resting coupling between the skeletal muscle L-type Ca 2+ channel and the type 1 ryanodine receptor

“…To determine whether the R174W mutation altered EC coupling, as has been demonstrated for other MH-causing mutations in either RyR1 (14)(15)(16)(17)(18)(19)(20)(25)(26)(27) or Ca V 1.1 (21), we measured myoplasmic Ca 2+ transients in the whole-cell configuration. As shown in Fig.…”

“…Previously characterized mutations in either channel have been found to promote activation of RyR1 during EC coupling and/or to enhance Ca 2+ entry via Ca V 1.1 (13)(14)(15)(16)(17)(18)(19)(20)(21). Thus, it has been proposed that augmented Ca 2+ movements via the EC coupling pathway or via the L-type current underlie the fulminant response to MH triggers.…”

Ca2+ handling abnormalities in early-onset muscle diseases: Novel concepts and perspectives

“…In primary myotubes, depolarizing stimuli that do not substantially deplete SR Ca 2+ stores can rapidly activate extracellular Ca 2+ entry by a mechanism called excitation coupled Ca 2+ entry (ECCE) that occurs through a complex made up of RyR1 and DHPR, the latter having dual functions not only as the voltage sensor for EC coupling but also as an L-type Ca2+ channel [61,62]. Experimental evidence supporting a pathogenic role of enhanced ECCE in MHS was provided by Cherednichenko et al [63] in mice knocked in for the RYR1 p.R163C mutation. Studies on skeletal muscles from these mice demonstrated: (i) that they exhibited enhanced depolarization-induced Ca 2+ entry, (ii) that this influx of Ca 2+ did not occur via store activated Ca 2+ entry, but via ECCE and (iii) that it is sensitive to dantrolene, the drug that is clinically used to revert MH reactions.…”

RGK protein-mediated impairment of slow depolarization- dependent Ca2+entry into developing myotubes

“…In this regard, RGK proteins may regulate a variety of cellular processes that have been linked to depolarization-dependent Ca 2+ entry such as maintenance of myoplasmic Ca 2+ levels during activity, 4 activation of transcription via NFAT translocation 20 and regulation of muscle strength 19 and mass (see below). Moreover, altered RGK protein-mediated modulation of depolarization-dependent Ca 2+ entry may potentially be involved in the pathophysiology of malignant hyperthermia, [21][22][23] central core disease, 20 and rippling muscle disease 24 since enhanced depolarization-dependent Ca 2+ entry is characteristic of these muscle disorders. The similar degree of inhibition between depolarizationdependent Ca 2+ entry and L-type current represents yet another commonality between these 2 modes of Ca 2+ flux into skeletal muscle (please see Table 4 of ref.…”