Mice expressing T4826I-RYR1 are viable but exhibit sex- and genotype-dependent susceptibility to malignant hyperthermia and muscle damage

Department of Veterinary Molecular Biosciences, University of California, Davis, California, USA.
The FASEB Journal (Impact Factor: 5.04). 11/2011; 26(3):1311-22. DOI: 10.1096/fj.11-197582
Source: PubMed


Mutation T4825I in the type 1 ryanodine receptor (RYR1(T4825I/+)) confers human malignant hyperthermia susceptibility (MHS). We report a knock-in mouse line that expresses the isogenetic mutation T4826I. Heterozygous RYR1(T4826I/+) (Het) or homozygous RYR1(T4826I/T4826I) (Hom) mice are fully viable under typical rearing conditions but exhibit genotype- and sex-dependent susceptibility to environmental conditions that trigger MH. Hom mice maintain higher core temperatures than WT in the home cage, have chronically elevated myoplasmic[Ca(2+)](rest), and present muscle damage in soleus with a strong sex bias. Mice subjected to heat stress in an enclosed 37°C chamber fail to trigger MH regardless of genotype, whereas heat stress at 41°C invariably triggers fulminant MH in Hom, but not Het, mice within 20 min. WT and Het female mice fail to maintain euthermic body temperature when placed atop a bed whose surface is 37°C during halothane anesthesia (1.75%) and have no hyperthermic response, whereas 100% Hom mice of either sex and 17% of the Het males develop fulminant MH. WT mice placed on a 41°C bed maintain body temperature while being administered halothane, and 40% of the Het females and 100% of the Het males develop fulminant MH within 40 min. Myopathic alterations in soleus were apparent by 12 mo, including abnormally distributed and enlarged mitochondria, deeply infolded sarcolemma, and frequent Z-line streaming regions, which were more severe in males. These data demonstrate that an MHS mutation within the S4-S5 cytoplasmic linker of RYR1 confers genotype- and sex-dependent susceptibility to pharmacological and environmental stressors that trigger fulminant MH and promote myopathy.

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Available from: Sam Goth, Oct 22, 2015
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    • "Importantly, strain specificity of gene-targeted mutations has been seen in such processes as glucocorticoid synthesis [57] and transport [58-61], as well as programmed cell death [52,62], all known to involve the endoplasmic reticulum and/or mitochondria. However, while some authors began to suspect the strain background when mouse model results did not match the results on human patients with the analogous mutation [63], numerous other authors did not question the results obtained only on C57BL/6J [64-72], and some even defended the superiority of this strain and considered the results obtained on other strains as problematic [73]a. "
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    ABSTRACT: Creation of lethal and synthetic lethal mutations in an experimental organism is a cornerstone of genetic dissection of gene function, and is related to the concept of an essential gene. Common inbred mouse strains carry background mutations, which can act as genetic modifiers, interfering with the assignment of gene essentiality. The inbred strain C57BL/6J, commonly known as “Black Six”, stands out, as it carries a spontaneous homozygous deletion in the nicotinamide nucleotide transhydrogenase (Nnt) gene [GenBank: AH009385.2], resulting in impairment of steroidogenic mitochondria of the adrenal gland, and a multitude of indirect modifier effects, coming from alteration of glucocorticoid-regulated processes. Over time, the popular strain has been used, by means of gene targeting technology, to assign “essential” and “redundant” qualifiers to numerous genes, thus creating an internally consistent “parallel universe” of knowledge. It is unrealistic to suggest phasing-out of this strain, given the scope of shared resources built around it, however, continuing on the road of “strain-unawareness” will result in profound waste of effort, particularly where translational research is concerned. The review analyzes the historical roots of this phenomenon and proposes that building of “parallel universes” should be urgently made visible to a critical reader by obligatory use of unambiguous and persistent tags in publications and databases, such as hypertext links, pointing to a vendor’s strain description web page, or to a digital object identifier (d.o.i.) of the original publication, so that any research done exclusively in C57BL/6J, could be easily identified. Reviewers This article was reviewed by Dr. Neil Smalheiser and Dr. Miguel Andrade-Navarro.
    Biology Direct 07/2014; 9(1):18. DOI:10.1186/1745-6150-9-18 · 4.66 Impact Factor
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    • "Samsó et al. 2005). New insight into the disease processes has been obtained from studies of the YS mice (Durham et al. 2008; Andronache et al. 2009; Boncompagni et al. 2009; Lanner et al. 2012) and other mouse models of malignant hyperthermia (Estéve et al. 2010; Feng et al. 2011; Yuen et al. 2012) and CCD (Zvaritch et al. 2007). Here we apply recently developed techniques to further evaluate the fundamental changes in Ca 2+ handling that are caused by the Y524S mutation. "
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    ABSTRACT: Malignant hyperthermia (MH) is linked to mutations in the type 1 ryanodine receptor, RyR1, the Ca(2+) channel of the sarcoplasmic reticulum (SR) of skeletal muscle. The Y522S MH mutation was studied for its complex presentation, which includes structurally and functionally altered cell "cores". Imaging cytosolic and intra-SR [Ca(2+)] in muscle cells of heterozygous YS mice we determined Ca(2+) release flux activated by clamp depolarization, permeability of the SR membrane- P, ratio of flux and [Ca(2+)] gradient- and SR Ca(2+) buffering power-B. In YS cells resting [Ca(2+)]SR was 45% of the value in normal littermates (WT). P was more than doubled, so that initial flux was normal. Measuring [Ca(2+)]SR(t) revealed dynamic changes in B(t). The alterations were similar to those caused by cytosolic BAPTA, which promotes release by hampering Ca(2+)-dependent inactivation (CDI). The [Ca(2+)] transients showed abnormal "breaks", decaying phases after an initial rise, traced to a collapse in flux and P. Similar breaks occurred in WT myofibers with calsequestrin reduced by siRNA; calsequestrin content, however, was normal in YS muscle. Thus, the Y522S mutation causes greater openness of the RyR1, lowers resting [Ca(2+)]SR and alters SR Ca(2+) buffering in a way that copies the functional instability observed upon reduction of calsequestrin content. The similarities with the effects of BAPTA suggest that the mutation, occurring near the cytosolic vestibule of the channel, reduces CDI as one of its primary effects. The unstable SR buffering, mimicked by silencing of calsequestrin, may help precipitate the loss of Ca(2+) control that defines a fulminant MH event.
    The Journal of Physiology 06/2013; 591(18). DOI:10.1113/jphysiol.2013.259572 · 5.04 Impact Factor
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    • "), and sudden death (Laitinen et al. 2004). Recent studies demonstrate that specific RYR mutations con­ fer sex– and gene–dose­dependent susceptibil­ ity to pharmacological (halogenated anesthetic) and environmental (heat) stressors that trigger malignant hyperthermia and muscle damage in otherwise asymptomatic individuals (Barrientos et al. 2012; Yuen et al. 2012). Importantly, PCB­95 is significantly more potent and effica­ cious in disrupting cation regulation of mutant R615C­RYR1 compared with wild type RyR1 (Ta and Pessah 2007). "
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    ABSTRACT: Aroclor 1254 (A1254) interferes with normal dendritic growth and plasticity in the developing rodent brain, but the mechanism(s) mediating this effect have yet to be established. Non-dioxin-like (NDL) polychlorinated biphenyls (PCBs) enhance the activity of ryanodine receptor (RyR) calcium ion (Ca(2+)) channels, which play a central role in regulating the spatiotemporal dynamics of intracellular Ca(2+) signaling. Ca(2+) signaling is a predominant factor in shaping dendritic arbors, but whether PCB potentiation of RyR activity influences dendritic growth is not known. We determined whether RyR activity is required for PCB effects on dendritic growth. Golgi analysis of hippocampi from weanling rats confirmed that developmental exposure via the maternal diet to NDL PCB-95 (2,2',3,5'6-pentachlorobiphenyl), a potent RyR potentiator, phenocopies the dendrite-promoting effects of A1254. Dendritic growth in dissociated cultures of primary hippocampal neurons and in hippocampal slice cultures is similarly enhanced by PCB-95 but not by PCB-66 (2,3,4',4-tetrachlorobiphenyl), a congener with negligible effects on RyR activity. The dendrite-promoting effects of PCB-95 are evident at concentrations as low as 2 pM and are inhibited by either pharmacologic blockade or siRNA knockdown of RyRs. Our findings demonstrate that environmentally relevant levels of NDL PCBs modulate neuronal connectivity via RyR-dependent effects on dendritic arborization. In addition, these findings identify RyR channel dysregulation as a novel mechanism contributing to dysmorphic dendritogenesis associated with heritable and environmentally triggered neurodevelopmental disorders.
    Environmental Health Perspectives 04/2012; 120(7):997-1002. DOI:10.1289/ehp.1104832 · 7.98 Impact Factor
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