Identification of a Dantrolene-binding Sequence on the Skeletal Muscle Ryanodine Receptor

Paul-Pletzer, Kalanethee; Yamamoto, Takakazu; Bhat, Manjunatha B.; Ma, Jianjie; Ikemoto, Noriaki; Jimenez, Leslie S.; Morimoto, Hiromi; Williams, Philip G.; Parness, Jerome

doi:10.1074/jbc.m205487200

Cited by 145 publications

(106 citation statements)

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“…Fourth, we show that substantial inhibition of RyR1-coupled SOCE by azumolene occurs only when cells are treated with this drug prior to C/R-induced RyR1 activation and SR Ca 2ϩ depletion. Because C/R treatment produces prolonged RyR1 channel opening, the inability of azumolene to inhibit SOCE when added after C/R treatment is consistent with previously published in vitro studies demonstrating that dantrolene interacts preferentially with the closed state of RyR1 (36,37). Therefore, we hypothesize that the inhibitory effect of azumolene and, by extension, of dantrolene on SOCE results from drug binding to the closed state of RyR1 The discordance between the ability of azumolene to inhibit SOCE versus SR Ca 2ϩ release in FDB muscle fibers suggests that Ca 2ϩ itself is not the direct signal from RyR1 that stimulates SOCE.…”

Section: Discussionsupporting

confidence: 75%

Azumolene Inhibits a Component of Store-operated Calcium Entry Coupled to the Skeletal Muscle Ryanodine Receptor

Zhao

Weisleder

Han

et al. 2006

Journal of Biological Chemistry

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Dantrolene reduces the elevated myoplasmic Ca2؉ generated during malignant hyperthermia, a pharmacogenetic crisis triggered by volatile anesthetics. Although specific binding of dantrolene to the type 1 ryanodine receptor (RyR1), the Ca 2؉ release channel of skeletal muscle sarcoplasmic reticulum, has been demonstrated, there is little evidence for direct dantrolene inhibition of RyR1 channel function. Recent studies suggest storeoperated Ca 2؉ entry (SOCE) contributes to skeletal muscle function, but the effect of dantrolene on this pathway has not been examined. Here we show that azumolene, an equipotent dantrolene analog, inhibits a component of SOCE coupled to activation of RyR1 by caffeine and ryanodine, whereas the SOCE component induced by thapsigargin is not affected. Our data suggest that azumolene distinguishes between two mechanisms of cellular signaling to SOCE in skeletal muscle, one that is coupled to and one independent from RyR1. Malignant hyperthermia (MH)2 is a potentially fatal pharmacogenetic syndrome in which exposure to volatile anesthetics triggers uncontrolled elevation of myoplasmic Ca 2ϩ concentrations ([Ca 2ϩ ] i ), skeletal muscle hypercontracture, and hypermetabolism, resulting in a dramatic rise in body temperature (1, 2). Mutations in the type 1 ryanodine receptor (RyR1), the major Ca 2ϩ release channel in skeletal muscle, are linked to MH susceptibility in pigs in an autosomal recessive manner (3-5). In humans, MH is transmitted as an autosomal dominant trait with incomplete penetrance, and the more than 80 mutations in RyR1 that have been identified appear in only about 50% of affected families (6, 7). Muscle bundles from MH-susceptible patients are hypersensitive to RyR1 agonists, including caffeine, Ca 2ϩ , and halothane (8, 9), the latter a member of the class of volatile anesthetics that triggers MH. Thus, the loss of control of RyR1-mediated Ca 2ϩ release likely contributes to the elevation of [Ca 2ϩ ] i observed in MH patients. To date, the only effective treatment for MH is dantrolene sodium, a skeletal muscle relaxant, which suppresses the uncontrolled rise in myoplasmic Ca 2ϩ , presumably by targeting RyR1 and suppressing its Ca 2ϩ channel activity (10 -12). Azumolene sodium is a structurally similar, equipotent analog of dantrolene, with an ϳ30-fold greater water solubility (13,14).Whereas hyperactivity or leakiness of the RyR1 channel has been described as the primary physiological defect in MH-susceptible muscle, the molecular mechanism underlying the suppression of [ -sensitive fluorescence measurements of RyR1-dependent intracellular Ca 2ϩ transients and extracellular Ca 2ϩ entry via SOCE, we show that azumolene inhibits SOCE both in skeletal muscle fibers and in cultured cells expressing RyR1. Our results reveal two modes of SOCE activation. One mode is RyR1-dependent and can be inhibited by the action of azumolene; the other is RyR1-independent and is insensitive to azumolene. EXPERIMENTAL PROCEDURESCell Culture-C1148 cells, a Chinese hamster ovary (CHO) ce...

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Section: Discussionsupporting

confidence: 75%

Azumolene Inhibits a Component of Store-operated Calcium Entry Coupled to the Skeletal Muscle Ryanodine Receptor

Zhao

Weisleder

Han

et al. 2006

Journal of Biological Chemistry

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“…An MH episode is characterized by elevations in body temperature, metabolic acidosis, hypoxia, tachycardia, skeletal muscle rigidity, and rhabdomyolysis (Denborough et al 1962;Ellis et al 1988;Pamukcoglu 1988;Britt et al 1991;Ryan and Tedeschi 1997) and is life threatening if not immediately treated with dantrolene, currently the only clinically approved treatment for MH (Ward et al 1986;Zhao et al 2001;Paul-Pletzer et al 2002). The incidence of MH is 1 in 15,000 anesthetized children and 1 in 50,000 to 100,000 anesthetized adults (MacLennan 1992;Strazis and Fox 1993;Rosenberg et al 2007).…”

Section: Role Of Ryanodine Receptors In Human Diseasesmentioning

confidence: 99%

Ryanodine Receptors: Structure, Expression, Molecular Details, and Function in Calcium Release

Lanner¹,

Georgiou²,

Joshi³

et al. 2010

Cold Spring Harbor Perspectives in Biology

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Ryanodine receptors (RyRs) are located in the sarcoplasmic/endoplasmic reticulum membrane and are responsible for the release of Ca 2þ from intracellular stores during excitation-contraction coupling in both cardiac and skeletal muscle. RyRs are the largest known ion channels (.2MDa) and exist as three mammalian isoforms (RyR 1 -3), all of which are homotetrameric proteins that interact with and are regulated by phosphorylation, redox modifications, and a variety of small proteins and ions. Most RyR channel modulators interact with the large cytoplasmic domain whereas the carboxy-terminal portion of the protein forms the ion-conducting pore. Mutations in RyR2 are associated with human disorders such as catecholaminergic polymorphic ventricular tachycardia whereas mutations in RyR1 underlie diseases such as central core disease and malignant hyperthermia. This chapter examines the current concepts of the structure, function and regulation of RyRs and assesses the current state of understanding of their roles in associated disorders.

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“…Functional studies on muscle fibers suggested the RyR as the potential target for the action of dantrolene (25,26), and respective binding sites were mapped to the protein (27)(28)(29). However, no action of dantrolene was found in most studies on single RyRs incorporated into lipid bilayers (26,30,31).…”

mentioning

confidence: 99%

Dantrolene requires Mg ²⁺ to arrest malignant hyperthermia

Choi

Koenig

Launikonis

2017

Proc. Natl. Acad. Sci. U.S.A.

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Malignant hyperthermia (MH) is a clinical syndrome of skeletal muscle that presents as a hypermetabolic response to volatile anesthetic gases, where susceptible persons may develop lethally high body temperatures. Genetic predisposition mainly arises from mutations on the skeletal muscle ryanodine receptor (RyR). Dantrolene is administered to alleviate MH symptoms, but its mechanism of action and its influence on the Ca 2+ transients elicited by MH triggers are unknown. Here, we show that Ca 2+ release in the absence of Mg 2+ is unaffected by the presence of dantrolene but that dantrolene becomes increasingly effective as cytoplasmic-free [Mg 2+ ] (free [Mg 2+ ] cyto ) passes mM levels. Furthermore, we found in human muscle susceptible to MH that dantrolene was ineffective at reducing halothane-induced repetitive Ca 2+ waves in the presence of resting levels of free [Mg 2+ ] cyto (1 mM). However, an increase of free [Mg 2+ ] cyto to 1.5 mM could increase the period between Ca 2+ waves. These results reconcile previous contradictory reports in muscle fibers and isolated RyRs, where Mg 2+ is present or absent, respectively, and define the mechanism of action of dantrolene is to increase the Mg 2+ affinity of the RyR (or "stabilize" the resting state of the channel) and suggest that the accumulation of the metabolite Mg 2+ from MgATP hydrolysis is required to make dantrolene administration effective in arresting an MH episode. malignant hyperthermia | dantrolene | ryanodine receptor | magnesium | skeletal muscle fiber R yanodine receptors (RyRs) are essential regulators of cytoplasmic Ca 2+ in muscle, heart, and brain (1-3). Congenital or acquired mishandling of Ca 2+ by RyRs is associated with organ dysfunction, myopathy, and the increased risk of sudden death (1, 4-7). Consequently, the search for drugs to modulate RyR function is an area of intense research (8-10). An example of a successful drug controlling the Ca 2+ mishandling of the RyR is the muscle relaxant dantrolene (11). It is primarily used to treat malignant hyperthermia (MH), a life-threatening condition in which genetically predisposed individuals adversely react to the exposure of volatile anesthetics (5, 12). Since its approval, the drug has cut the mortality rate associated with MH from more than 80% to below 2% (11). Despite this success, the exact mechanism of how dantrolene antagonizes MH episodes and depresses overactive Ca 2+ release during MH episodes remains unknown.Susceptibility to MH most commonly arises from mutations in the RyR1 gene, the Ca 2+ release channel of skeletal muscle. Mg 2+ is present in the muscle at ∼1 mM and exerts an inhibitory action over the RyR1, which needs to be overcome for normal voltagecontrolled Ca 2+ release (13-16). RyR1 variants have a lowered affinity for Mg 2+ (13,17), making them more prone to opening and resulting in increased sensitivity to RyR agonist. The ensuing abnormal Ca 2+ release in RyR variants (18-24) during an MH event leads to excessive heat production as the muscle attempts to clear the...

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Identification of a Dantrolene-binding Sequence on the Skeletal Muscle Ryanodine Receptor

Cited by 145 publications

References 46 publications

Azumolene Inhibits a Component of Store-operated Calcium Entry Coupled to the Skeletal Muscle Ryanodine Receptor

Azumolene Inhibits a Component of Store-operated Calcium Entry Coupled to the Skeletal Muscle Ryanodine Receptor

Ryanodine Receptors: Structure, Expression, Molecular Details, and Function in Calcium Release

Dantrolene requires Mg ²⁺ to arrest malignant hyperthermia

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Identification of a Dantrolene-binding Sequence on the Skeletal Muscle Ryanodine Receptor

Cited by 145 publications

References 46 publications

Azumolene Inhibits a Component of Store-operated Calcium Entry Coupled to the Skeletal Muscle Ryanodine Receptor

Azumolene Inhibits a Component of Store-operated Calcium Entry Coupled to the Skeletal Muscle Ryanodine Receptor

Ryanodine Receptors: Structure, Expression, Molecular Details, and Function in Calcium Release

Dantrolene requires Mg 2+ to arrest malignant hyperthermia

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Dantrolene requires Mg ²⁺ to arrest malignant hyperthermia