Correspondence to: Robert T. Dirksen, Department of Pharmacology and Physiology, University of Rochester School of Medicine and Dentistry, 601 Elmwood Avenue, Rochester, NY 14642. Fax:(716) 273-2652 E-mail:robert_dirksen{at}URMC.rochester.edu.
![]() |
Abstract |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Central core disease (CCD) is a human myopathy that involves a dysregulation in muscle Ca2+ homeostasis caused by mutations in the gene encoding the skeletal muscle ryanodine receptor (RyR1), the protein that comprises the calcium release channel of the SR. Although genetic studies have clearly demonstrated linkage between mutations in RyR1 and CCD, the impact of these mutations on release channel function and excitation-contraction coupling in skeletal muscle is unknown. Toward this goal, we have engineered the different CCD mutations found in the NH2-terminal region of RyR1 into a rabbit RyR1 cDNA (R164C, I404M, Y523S, R2163H, and R2435H) and characterized the functional effects of these mutations after expression in myotubes derived from RyR1-knockout (dyspedic) mice. Resting Ca2+ levels were elevated in dyspedic myotubes expressing four of these mutants (Y523S > R2163H > R2435H R164C > I404M RyR1). A similar rank order was also found for the degree of SR Ca2+ depletion assessed using maximal concentrations of caffeine (10 mM) or cyclopiazonic acid (CPA, 30 µM). Although all of the CCD mutants fully restored L-current density, voltage-gated SR Ca2+ release was smaller and activated at more negative potentials for myotubes expressing the NH2-terminal CCD mutations. The shift in the voltage dependence of SR Ca2+ release correlated strongly with changes in resting Ca2+, SR Ca2+ store depletion, and peak voltagegated release, indicating that increased release channel activity at negative membrane potentials promotes SR Ca2+ leak. Coexpression of wild-type and Y523S RyR1 proteins in dyspedic myotubes resulted in release channels that exhibited an intermediate degree of SR Ca2+ leak. These results demonstrate that the NH2-terminal CCD mutants enhance release channel sensitivity to activation by voltage in a manner that leads to increased SR Ca2+ leak, store depletion, and a reduction in voltage-gated Ca2+ release. Two fundamentally distinct cellular mechanisms (leaky channels and EC uncoupling) are proposed to explain how altered release channel function caused by different mutations in RyR1 could result in muscle weakness in CCD.
Key Words: excitation-contraction coupling, calcium channels, muscle disease, calcium homeostasis
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Rapid elevations of intracellular Ca2+ in skeletal muscle are controlled by a unique, bidirectional signaling interaction between L-type Ca2+ channels (L-channels* or DHPRs) in the sarcolemma and Ca2+ release channels (ryanodine receptors or RyR1s) of the SR. During skeletal muscle excitation-contraction (EC) coupling, action potentials in the sarcolemma induce conformational changes in dihydropyridine receptors (DHPRs) that rapidly trigger the opening of nearby SR Ca2+ release channels (orthograde coupling;
CCD, first described by
CCD and MH are localized to human chromosome 19q13.1, which includes the locus of RyR1 (
However, the physiological effects of the CCD mutations in RyR1 on EC coupling and intracellular Ca2+ homeostasis in intact skeletal muscle have yet to be systematically studied. As a step toward this goal, we recently evaluated the functional properties of a CCD mutation in MH/CCD region 3 (I4897T) after expression in skeletal myotubes derived from "RyR1-knockout" (dyspedic) mice (
Five amino acid residues (R163, I403, Y522, R2163, and R2435) account for the six different CCD mutations in the NH2-terminal or cytoplasmic "foot" region of the human RyR1 protein (MH/CCD regions 1 and 2;
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Preparation and cDNA Injections of Dyspedic Myotubes
Myotubes were prepared from primary cultures of dyspedic muscle as described previously (
|
|
|
|
|
Intracellular Ca2+ Measurements in Intact Myotubes
Measurements of resting Ca2+ were obtained in intact, Indo-1 AMloaded myotubes (Molecular Probes) as described previously (150 nM resulted in contractions that could introduce movement artifacts into the calibration (
10-min wash intervals were similar. Simple gravity perfusion (<30s for complete solution exchange) was used to monitor the slower changes in intracellular Ca2+ induced by application of 30 µM CPA (
Simultaneous Measurements of Macroscopic Ca2+ Currents and Ca2+ Transients
The whole-cell patch-clamp technique was used to simultaneously measure voltage-gated L-currents and Ca2+ transients in expressing myotubes (
![]() |
(1) |
where Vrev is the extrapolated reversal potential of the Ca2+ current, Vm is the membrane potential during the test pulse, Gmax is the maximum L-channel conductance, VG is the voltage for half activation of Gmax, and kG is a slope factor. Relative changes in cytosolic Ca2+ in patch-clamp experiments were recorded in Fluo-3-dialyzed myotubes. Fluorescence traces were analogue-filtered (
= 0.5 ms) before digitization (10 kHz), and subsequently expressed as
F/F. Amplitudes at the end of each test pulse were plotted as a function of the membrane potential, and fitted according to:
![]() |
(2) |
where (F/F)max is the calculated maximal fluorescence change during the test pulse, VF
is the midpoint potential, and kF is a slope factor.
Recording Solutions
Cytosolic Ca2+ levels were monitored in myotubes bathed in normal rodent Ringer containing the following (in mM): 145 NaCl, 5 KCl, 2 CaCl2, 1 MgCl2, and 10 HEPES, pH 7.4. For measurements of macroscopic Ca2+ currents and intracellular Ca2+ transients, the pipette solution contained the following (in mM): 145 cesium aspartate, 10 CsCl, 0.1 Cs2-EGTA, 1.2 MgCl2, 5 Mg-ATP, 0.2 K5-Fluo-3 (Molecular Probes), and 10 HEPES, pH 7.4. In these experiments, the external solution contained the following (in mM): 145 TEA-Cl, 10 CaCl2, 0.003 TTX, and 10 HEPES, pH 7.4. Except where noted, all chemical reagents were obtained from Sigma-Aldrich.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Effects of CCD Mutations on Resting Ca2+ and SR Ca2+ Content
Compared with wild-type RyR1, HEK-293 cells expressing NH2-terminal CCD mutations in RyR1 exhibited elevated resting Ca2+ levels and a reduction in the Ca2+ content of the ER ( RyR1. For clarity, the data for each of the different constructs in Fig 1 Fig 2 Fig 3 Fig 4 are presented using this rank order. Under control conditions, some of the myotubes expressing either wild-type RyR1 or the different NH2-terminal CCD mutants exhibited spontaneous intracellular Ca2+ transients that varied in frequency, amplitude, and duration (Fig 1 A). The percentages of myotubes exhibiting spontaneous Ca2+ oscillations during the first
60 s of recording were the following: 33% (14/43), 25% (5/20), 54% (15/28), 65% (13/20), 54% (15/28), and 10% (2/20), for RyR1-, I404M-, R164C-, R2435H-, R2163H-, and Y523S-expressing myotubes, respectively. In contrast, none of the I4897T-expressing myotubes (0/16) or uninjected dyspedic myotubes (0/33) studied exhibited spontaneous Ca2+ oscillations. In general, resting Ca2+ levels were elevated and Ca2+ oscillations were smaller and of higher frequency in the NH2-terminal CCD-expressing myotubes (Fig 1 A and 2 A). However, since the frequency and magnitude of Ca2+ oscillations could be influenced by differences in spontaneous electrical activity, the patterns of these Ca2+ oscillations for the different constructs were not characterized in greater detail. Interestingly, a similar pattern of Ca2+ oscillations were also observed at the holding potential (-80 mV) in whole-cell patch-clamp experiments (data not shown), suggesting that increased spontaneous release does not solely arise from elevated intracellular Ca2+ levels or membrane depolarization.
The trend for the different CCD mutations in RyR1 to cause an elevation in resting Ca2+ as well as smaller amplitude Ca2+ oscillations is consistent with the notion that the CCD mutations lead to the expression of leaky SR Ca2+ release channels. Since leaky SR Ca2+ release channels may result in SR Ca2+ store depletion we evaluated SR Ca2+ content after application of a maximal concentration of caffeine (10 mM; Fig 1 A). Dyspedic myotubes expressing four of the five different NH2-terminal CCD mutant RyR1 proteins (R164C, R2435H, R2163H, and Y523S) exhibited a significantly smaller response to caffeine compared with that of wild-type RyR1 (P < 0.05; Fig 1). Interestingly, repetitive Ca2+ oscillations were typically observed throughout the caffeine application in 90% (17/19) of RyR1-expressing myotubes. However, repetitive caffeine-induced Ca2+ oscillations were less frequent: 43% (6/14), 31% (4/13), 43% (3/7), 0% (0/11), and 0% (0/8) for I404M, R164C, R2435H, R2163H, and Y523S, respectively. A reduction in repetitive Ca2+ oscillations could arise from differences in the caffeine-induced plateau Ca2+ level or reductions in either SR Ca2+ content and/or release channel inhibition by high Ca2+ (
The data presented in Fig 1 suggest that expression of NH2-terminal CCD mutant RyR1 proteins in dyspedic myotubes results in a significant depletion of SR Ca2+ stores. This idea was further tested by evaluating SR Ca2+ content using a method to release Ca2+ by a mechanism that is independent of RyR1 activation (Fig 2). For these experiments, luminal SR Ca2+ stores were assessed after treatment with CPA, an agent that increases cytosolic Ca2+ by reversibly inhibiting SR Ca2+-ATPase pumps, and thereby prevents the reuptake of Ca2+ lost through passive leak pathways. Application of CPA (Fig 2, 30 µM, black bars) induced similar increases in cytosolic Ca2+ in intact dyspedic myotubes either expressing RyR1 or I404M. However, dyspedic myotubes expressing the other NH2-terminal CCD mutants exhibited significantly smaller elevations in cytosolic Ca2+ after application of CPA, which is consistent with these mutants reducing luminal SR Ca2+ levels. Although caffeine fails to activate Ca2+ release in I4897T-expressing myotubes (Fig 1 C), these myotubes exhibit a resting Ca2+ level and CPA-sensitive Ca2+ store comparable to that of RyR1-expressing myotubes (Fig 1 A and 2 B;
Effects on Retrograde and Orthograde Signals of EC Coupling
EC coupling in skeletal muscle involves a unique bidirectional signaling interaction between RyR1s and DHPRs. Accordingly, while DHPRs trigger the activation of RyR1 proteins in response to a sarcolemmal depolarization (orthograde coupling), the functional activity of the DHPR (retrograde coupling) is in turn strongly influenced by the presence of RyR1 (
Although the NH2-terminal CCD mutations in RyR1 did not alter retrograde coupling, significant effects on orthograde coupling were observed (Fig 4). Fig 4 A illustrates representative voltage-gated Ca2+ transients recorded under whole-cell voltage clamp for dyspedic myotubes expressing wild-type RyR1 and each of the different CCD mutant RyR1 proteins. In general, voltage-gated Ca2+ transients attributable to the NH2-terminal CCD mutant release channels were smaller and activated at more negative potentials than Ca2+ transients arising from wild-type RyR1. This can best be appreciated by comparing voltage-gated Ca2+ transients arising from RyR1 (Fig 4 A, upper left) and Y523S (Fig 4 A, lower right). Even though the Y523S transients illustrated in Fig 4 A were the largest we recorded (n = 11), the maximal transient (at +70 mV) for this construct was still more than three times smaller than that of wild-type RyR1. In addition, the threshold for activation of Y523S-expressing myotubes was 40 mV more hyperpolarized than that of RyR1-expressing myotubes. The other NH2-terminal CCD mutations in RyR1 also resulted in reductions in maximal voltage-gated Ca2+ release and a negative shift in the voltage dependence of release, although to a lesser extent that Y523S. Thus, although the largest maximal voltage-gated Ca2+ transients were recorded from wild-type RyR1-expressing myotubes, the NH2-terminal CCD mutants actually exhibited larger Ca2+ transients at threshold potentials. For example, the Ca2+ transient amplitudes at -10 mV were the following (in
F/F): 0.04 ± 0.02, 0.11 ± 0.05, 0.72 ± 0.19, 0.33 ± 0.10, 1.18 ± 0.45, and 0.27 ± 0.15, for RyR1, I404M, R164C, R2435H, R2163H, and Y523S, respectively. To provide a more quantitative description of this observation, Boltzmann fits to the voltage dependence of SR Ca2+ release for each construct were used to determine the maximal change in fluorescence (
F/Fmax), the half-maximal activation voltage (VF
), and voltage sensitivity (kF; Fig 4 B). This analysis revealed that each of the NH2-terminal CCD VF
mutations in RyR1 caused a significant negative shift in without changing kF (Table 1). The various degrees of hyperpolarizing shift in the activation of voltage-gated SR Ca2+ release caused by the different NH2-terminal CCD mutations in RyR1 are best appreciated by comparing the normalized voltage dependence of release for each mutant to that of wild-type RyR1 (Fig 4 C).
|
Since individuals with CCD are typically heterozygous for the causative RyR1 mutation, we also investigated the effects of coexpression of RyR1 and Y523S in dyspedic myotubes (Fig 5). We evaluated the impact of RyR1/Y523S coexpression since this mutation resulted in the most severe disruption in Ca2+ homeostasis (Fig 1 and Fig 2) and voltage-gated SR Ca2+ release (Fig 4). Not surprisingly, RyR1/Y523S-expressing dyspedic myotubes displayed L-currents with a similar magnitude, kinetics, and voltage dependence as those recorded from homozygous RyR1-expressing myotubes (Fig 5a and Fig B, and Table 1). However, RyR1/Y523S-expressing myotubes exhibited the following: only a moderate elevation in resting Ca2+ (for RyR1, 49 ± 7.8 nM, n = 43; for RyR1/Y523S, 94 ± 11 nM, n = 23); and voltage-gated Ca2+ transients that were intermediate in magnitude and voltage dependence to those arising from homozygous expression of either construct (Fig 5C and Fig D). The effects on voltage-gated Ca2+ release are qualitatively similar to those observed after coexpression of RyR1/I4897T ( Ca2+ and SR Ca2+ content (
Correlation between Effects on Ca2+ Dynamics and Release Channel Sensitivity to Activation by Voltage (VF)
To test whether changes in release channel sensitivity to voltage (i.e., VF) might account for the observed changes in resting Ca2+, SR Ca2+ content, and maximal voltage-gated Ca2+ release, a linear correlation analysis was conducted for the data obtained from dyspedic myotubes expressing RyR1 and each of the NH2-terminal CCD mutations (Fig 6). The elevation in resting Ca2+ correlated strongly with the degree of shift in VF
(Fig 6 A, r = 0.97, P < 0.05). In addition, myotube responsiveness to maximal concentrations of both caffeine (Fig 6 B, r = 0.97, P < 0.05) and CPA (Fig 6 C, r = 0.80, P = 0.05), two different means of assessing SR Ca2+ content, also correlated well with VF
. Finally, the degree of reduction in maximal voltage-gated SR Ca2+ release also correlated with release channel sensitivity to activation by voltage (Fig 6 D, r = 0.86, P < 0.05) across the different RyR1 constructs. Although these correlational analyses fall short of establishing causation, the results suggest that increased release channel sensitivity to activation by voltage contributes to the enhanced SR Ca2+ leak, Ca2+ store depletion, and a reduction in Ca2+ released during EC coupling in dyspedic myotubes expressing the different NH2-terminal CCD mutations in RyR1.
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Functional Impact of NH2-terminal CCD Mutations on RyR1 Activity in Dyspedic Myotubes
Our experiments are the first to characterize the relative effects of the different NH2-terminal CCD mutations in RyR1 on release channel activity operating within a skeletal muscle environment. The results indicate that dyspedic myotubes expressing the different NH2-terminal CCD mutations in RyR1 exhibit different degrees of SR Ca2+ leak as judged by the presence of parallel, but opposing, changes in resting Ca2+ and SR Ca2+ content. None of the NH2-terminal CCD mutants markedly altered L-current magnitude, kinetics, or voltage dependence (Fig 3). However, the NH2-terminal CCD mutations reduced maximal voltage-gated Ca2+ release and caused a negative shift in VF (Fig 4), indicating that the CCD mutations in RyR1 selectively alter the orthograde signal of skeletal muscle EC coupling. The degree of reduction in voltage-gated release correlated well with the shift in VF
across the different NH2-terminal CCD mutations in RyR1. These data are the first to suggest that increased release channel activity at negative membrane potentials results in an elevation in resting Ca2+ and a partial reduction in SR Ca2+ content. In spite of these correlations, we should point out that our experiments have not definitively demonstrated that the observed increased sensitivity to activation by voltage (i.e., negative shift in VF
) observed for the NH2-terminal CCD mutants is directly responsible for an increase in SR Ca2+ leak.
Two Distinct Cellular Mechanisms for Muscle Weakness in CCD
Since our results obtained with the NH2-terminal CCD mutants are in contrast to those obtained after expression of I4897T ( observed in porcine myotubes (
. Our finding of variable shifts in VF
(ranging from 5 to 31 mV) caused by the NH2-terminal CCD mutations in RyR1 could, therefore, arise from variable reductions in the value of K.
|
We propose two fundamentally distinct cellular mechanisms (leaky channels and EC uncoupling) to explain how altered SR Ca2+ release channel activity caused by different mutations in RyR1 may result in muscle weakness in CCD (Fig 7). The three basic schemes presented in Fig 7 are based on our results obtained from expression of RyR1, Y523S, and I4898T in dyspedic myotubes and are based on the 5-state kinetic model of DHPR-RyR1 coupled gating ( for Ca2+ release without significantly altering the voltage dependence of L-channel opening. Thus, our finding of a selective shift in VF
with expression of Y523S mutation in RyR1 (as well as R2163H, RyR1/Y523S, R164C, R2435H, and I404M) in dyspedic myotubes can also be explained by a reduction in K. The increased sensitivity of Y523S-containing release channels to activation at negative voltages should result in a partial depletion of the SR Ca2+ store. If store content is reduced enough, SR Ca2+ release elicited by a single action potential would be smaller than usual, thus, resulting in muscle weakness. Coexpression of Y523S with RyR1 resulted in an intermediate shift in VF
, a more moderate reduction in peak voltage-gated SR Ca2+ release, and a resting Ca2+ level that was more than RyR1 but less than Y523S. Thus, similar to our previous findings regarding I4897T, the Y523S mutation in RyR1 exerts a dominant negative action on the activity of the resulting SR Ca2+ release channels, which is consistent with the autosomal dominant pattern of inheritance of this human myopathy (
We have previously reported that expression in dyspedic myotubes of release channels harboring the I4898T CCD mutation do not function as leaky SR Ca2+ release channels, but rather reflect a functional uncoupling of sarcolemmal excitation from SR Ca2+ release (EC uncoupling) ( values of RyR1- and RyR1/I4897T-expressing dyspedic myotubes did not differ significantly (
, and thus, likely involves a fundamentally distinct mechanism than that of the NH2-terminal CCD mutations. One possibility is that the lack of voltage-gated Ca2+ release after homozygous expression of I4897T could arise from a selective disruption in orthograde coupling, such that the transition governed by K becomes essentially nonpermissible (i.e., K approaches infinity; not depicted in Fig 7). A reduction in the maximal voltage-gated Ca2+ transient occurring in the absence of a change in VF
after RyR1/I4897T coexpression could arise from certain heteromeric release channel combinations (e.g., consisting predominantly of RyR1) that undergo the transition governed by K normally, whereas other combinations (e.g., consisting predominantly of I4897T) almost never undergo this transition. Alternatively, the I4897T mutation may drastically alter release channel permeation and/or gating in such a way that Ca2+ flux through activated channels is severely compromised (Fig 7 C, shaded release channels). Thus, release channels comprised of I4897T subunits may undergo the voltage-independent activating transition governed by K normally, but Ca2+ may only conduct poorly through activated I4897T release channels. Recent findings that the I4897T residue contributes to a conserved RyR pore-forming sequence (
Comparison of Results with Previous Reports
Due to the presence of the porcine model for MH, the functional effects of the R615C MH mutation in RyR1 has been the most thoroughly characterized of the different disease mutations in RyR1 (for reviews see without markedly altering the voltage dependence of L-channel activation.
Heterologous expression of MH and CCD mutations in nonmuscle cells has been an alternate approach used to compare the functional properties of mutant release channels (
Effects of CCD Mutations in RyR1 on Resting Ca2+
The influence of the MH/CCD mutations in RyR1 on resting intracellular Ca2+ levels is still one of the most controversial and hotly debated issues in the field (for review see
The precise mechanism(s) underlying how leaky SR Ca2+ release channels lead to long-term elevations in resting Ca2+ is still unclear. If SR Ca2+ leak exceeds calcium uptake and extrusion, a new dynamic equilibrium of calcium mobilization could be established that leads to a higher resting calcium level. However, since the predominant Ca2+ extrusion processes of the plasma membrane (Ca2+ pumps and Na/Ca2+ exchangers) are stimulated by elevations in resting Ca2+, then over longer periods of time these transport mechanisms may be sufficient to compensate for the Ca2+ leak, ultimately resulting in a return of Ca2+ toward normal levels. Under such a scenario, the long-term effect of SR leak would manifest as a similar resting Ca2+ level in the face of SR Ca2+ store depletion. The caveat to this would be the presence of restricted intracellular domains in which Ca2+ diffusion to the plasma membrane is limited via some currently unknown mechanism (e.g., a central core;
Implications for the Pathophysiology of CCD
Although diagnosis of CCD is determined through identification of amorphous central areas in type 1 muscle fibers that lack mitochondria and oxidative enzyme activity, the underlying processes involved in the formation of central cores and their biochemical composition is still a mystery. Global resting Ca2+ levels are elevated after transient expression of CCD mutations in RyR1 in HEK-293 cells (
![]() |
Footnotes |
---|
* Abbreviations used in this paper: CCD, central core disease; CPA, cyclopiazonic acid; DHPR, dihydropyridine receptor; EC, excitation-contraction; L-channel, L-type Ca2+ channel; MH, malignant hyperthermia.
![]() |
Acknowledgements |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
We would like to thank Drs. Kurt G. Beam and Paul D. Allen for providing us access to the dyspedic mice used in this study, as well as for their advice and continued support. We would also like to thank Drs. W. Melzer, S.-S. Sheu, and Ms. K.M.S. O'Connell for helpful discussions and comments on the manuscript, and Linda Groom for excellent technical assistance.
This work was supported by a grant from the National Institutes of Health (AR44657 to R.T. Dirksen), a Neuromuscular Disease Research grant from the Muscular Dystrophy Association (to R.T. Dirksen), and a Consejo Nacional de Ciencia y Tecnologia postdoctoral fellowship (to G. Avila).
Submitted: 22 May 2001
Revised: 25 July 2001
Accepted: 26 July 2001
![]() |
References |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Avila, G., O'Brien, J.J., and Dirksen, R.T. 2001a. Excitation-contraction uncoupling by a human central core disease mutation in the ryanodine receptor. Proc. Natl. Acad. Sci. USA. 7:4215-4220.
Avila, G., O'Connell, K.M.S., Groom, L., and Dirksen, R.T. 2001b. Ca2+ release through ryanodine receptors regulates skeletal muscle L-type Ca2+ channel expression. J. Biol. Chem. 276:17732-17738
Balshaw, D., Gao, L., and Meissner, G. 1999. Luminal loop of the ryanodine receptor: a pore-forming segment? Proc. Natl. Acad. Sci. USA. 7:3345-3347.
Barone, V., Massa, O., Intravaia, E., Bracco, A., Di Martino, A., Tegazzin, V., Cozzolino, S., and Sorrentino, V. 1999. Mutation screening of the RYR1 gene and identification of two novel mutations in Italian malignant hyperthermia families. J. Med. Genet. 2:115-118.
Beutner, G., Sharma, V.K., Giovannucci, D.R., Yule, D.I., and Sheu, S.-S. 2001. Identification of a ryanodine receptor in rat heart mitochondria. J. Biol. Chem. 276:21482-21488
Buck, E.D., Nguyen, H.T., Pessah, I.N., and Allen, P.D. 1997. Dyspedic mouse skeletal muscle expresses major elements of the triadic junction but lacks detectable ryanodine receptor protein and function. J. Biol. Chem. 272:7360-7367
Dietze, B., Henke, J., Eichinger, H.M., Lehmann-Horn, F., and Melzer, W. 2000. Malignant hyperthermia mutation Arg615Cys in the porcine ryanodine receptor alters voltage dependence of Ca2+ release. J. Physiol. 526:507-514
Dubowitz, V., and Pearse, A.G.E. 1960. Oxidative enzymes and phosphorylase in central-core disease of muscle. Lancet 2:23-24[Medline].
Fill, M., Coronado, R., Mickelson, J.R., Vilven, J., Ma, J.J., Jacobson, B.A., and Louis, C.F. 1990. Abnormal ryanodine receptor channels in malignant hyperthermia. Biophys. J. 3:471-475.
Fujii, J., Otsu, K., Zorzato, F., de Leon, S., Khanna, V.K., Weiler, J.E., O'Brien, P.J., and MacLennan, D.H. 1991. Identification of a mutation in porcine ryanodine receptor associated with malignant hyperthermia. Science. 5018:448-451.
Gallant, E.M., and Lentz, L.R. 1992. Excitation-contraction coupling in pigs heterozygous for malignant hyperthermia. Am. J. Physiol. 262:C422-C426
Gallant, E.M., and Jordan, R.C. 1996. Porcine malignant hyperthermia: genotype and contractile threshold of immature muscles. Muscle Nerve. 1:68-73.
Gallant, E.M., Balog, E.M., and Beam, K.G. 1996. Slow calcium current is not reduced in malignant hyperthermic porcine myotubes. Muscle Nerve. 4:450-455.
Gao, L., Balshaw, D., Xu, L., Tripathy, A., Xin, C., and Meissner, G. 2000. Evidence for a role of the lumenal M3-M4 loop in skeletal muscle Ca2+ release channel (ryanodine receptor) activity and conductance. Biophys. J 2:828-840.
Ginty, D.D. 1997. Calcium regulation of gene expression: isn't that spatial? Neuron. 2:183-186.
Grabner, M., Dirksen, R.T., Suda, N., and Beam, K.G. 1999. The II-III loop of the skeletal muscle dihydropyridine receptor is responsible for the bi-directional coupling with the ryanodine receptor. J. Biol. Chem. 31:21913-21919.
Gunter, T.E., Gunter, K.K., Sheu, S.-S., and Gavin, C.E. 1994. Mitochondrial calcium transport: physiological and pathological relevance. Am. J. Physiol. 267:C313-C339
Hayashi, K., Miller, R.G., and Brownell, A.K. 1989. Central core disease: ultrastructure of the sarcoplasmic reticulum and t-tubules. Muscle Nerve. 2:95-102.
Isaacs, H., Heffron, J.J., and Badenhorst, M. 1975. Central core disease. A correlated genetic, histochemical, ultramicroscopic, and biochemical study. J. Neurol. Neurosurg. Psychiatry. 12:1177-1186.
Jurkat-Rott, K., McCarthy, T., and Lehmann-Horn, F. 2000. Genetics and pathogenesis of malignant hyperthermia. Muscle Nerve. 1:4-17.
Kausch, K., Lehmann-Horn, F., Janka, M., Wieringa, B., Grimm, T., and Muller, C.R. 1991. Evidence for linkage of the central core disease locus to the proximal long arm of human chromosome 19. Genomics. 3:765-769.
Loke, J., and MacLennan, D.H. 1998. Malignant hyperthermia and central core disease: disorders of Ca2+ release channels. Am. J. Med. 5:470-486.
Lorenzon, N.M., and Beam, K.G. 2000. Calcium channelopathies. Kidney Intl. 3:794-802.
Lynch, P.J., Tong, J., Lehane, M., Mallet, A., Giblin, L., Heffron, J.J., Vaughan, P., Zafra, G., MacLennan, D.H., and McCarthy, T.V. 1999. A mutation in the transmembrane/luminal domain of the ryanodine receptor is associated with abnormal Ca2+ release channel function and severe central core disease. Proc. Natl. Acad. Sci. USA. 7:4164-4169.
MacKenzie, A.E., Korneluk, R.G., Zorzato, F., Fujii, J., Phillips, M., Iles, D., Wieringa, B., Leblond, S., Bailly, J., and Willard, H.F. 1990. The human ryanodine receptor gene: its mapping to 19q13.1, placement in a chromosome 19 linkage group, and exclusion as the gene causing myotonic dystrophy. Am. J. Hum. Gen. 6:1082-1089.
MacLennan, D.H. 1992. The genetic basis of malignant hyperthermia. Trends Pharmacol. Sci. 8:330-334.
MacLennan, D.H., and Phillips, M.S. 1995. The role of the skeletal muscle ryanodine receptor (RYR1) gene in malignant hyperthermia and central core disease. Soc. Gen. Physiol. Series 50:89-100[Medline].
Manning, B.M., Quane, K.A., Ording, H., Urwyler, A., Tegazzin, V., Lehane, M., O'Halloran, J., Hartung, E., Giblin, L.M., and Lynch, P.J. et al. 1998. Identification of novel mutations in the ryanodine-receptor gene (RYR1) in malignant hyperthermia: genotype-phenotype correlation. Am. J. Hum. Genet. 3:599-609.
McCarthy, T.V., Quane, K.A., and Lynch, P.J. 2000. Ryanodine receptor mutations in malignant hyperthermia and central core disease. Hum. Mutat. 5:410-417.
Melzer, W., Herrmann-Frank, A., and Luttgau, H.C. 1995. The role of Ca2+ ions in excitation-contraction coupling of skeletal muscle fibres. Biochim. Biophys. Acta. 1:59-116.
Mickelson, J.R., and Louis, C.F. 1996. Malignant hyperthermia: excitation-contraction coupling, Ca2+ release channel, and cell Ca2+ regulation defects. Physiol. Rev. 2:537-592.
Monnier, N., Romero, N.B., Lerale, J., Nivoche, Y., Qi, D., MacLennan, D.H., Fardeau, M., and Lunardi, J. 2000. An autosomal dominant congenital myopathy with cores and rods is associated with a neomutation in the RYR1 gene encoding the skeletal muscle ryanodine receptor. Hum. Mol. Genet. 18:2599-2608.
Nakai, J., Dirksen, R.T., Nguyen, H.T., Pessah, I.N., Beam, K.G., and Allen, P.D. 1996. Enhanced dihydropyridine receptor channel activity in the presence of ryanodine receptor. Nature. 6569:72-75.
Quane, K.A., Healy, J.M., Keating, K.E., Manning, B.M., Couch, F.J., Palmucci, L.M., Doriguzzi, C., Fagerlund, T.H., Berg, K., and Ording, H. 1993. Mutations in the ryanodine receptor gene in central core disease and malignant hyperthermia. Nat. Genet. 1:51-55.
Quane, K.A., Keating, K.E., Healy, J.M., Manning, B.M., Krivosic-Horber, R., Krivosic, I., Monnier, N., Lunardi, J., and McCarthy, T.V. 1994. Mutation screening of the RYR1 gene in malignant hyperthermia: detection of a novel Tyr to Ser mutation in a pedigree with associated central cores. Genomics. 1:236-239.
Shy, G.M., and Magee, K.R. 1956. A new congenital non-progressive myopathy. Brain 79:610-621.
Tong, J., Oyamada, H., Demaurex, N., Grinstein, S., McCarthy, T.V., and MacLennan, D.H. 1997. Caffeine and halothane sensitivity of intracellular Ca2+ release is altered by 15 calcium release channel (ryanodine receptor) mutations associated with malignant hyperthermia and/or central core disease. J. Biol. Chem. 42:26332-26339.
Tong, J., McCarthy, T.V., and MacLennan, D.H. 1999. Measurement of resting cytosolic Ca2+ concentrations and Ca2+ store size in HEK-293 cells transfected with malignant hyperthermia or central core disease mutant Ca2+ release channels. J. Biol. Chem. 2:693-702.
Treves, S., Larini, F., Menegazzi, P., Steinberg, T.H., Koval, M., Vilsen, B., Andersen, J.P., and Zorzato, F. 1994. Alteration of intracellular Ca2+ transients in COS-7 cells transfected with the cDNA encoding skeletal-muscle ryanodine receptor carrying a mutation associated with malignant hyperthermia. Biochem. J. 3:661-665.
Zhang, Y., Chen, H.S., Khanna, V.K., de Leon, S., Phillips, M.S., Schappert, K., Britt, B.A., Browell, A.K., and MacLennan, D.H. 1993. A mutation in the human ryanodine receptor gene associated with central core disease. Nat. Genet. 1:46-50.
Zhao, M., Li, P., Li, X., Zhang, L., Winkfein, R.J., and Chen, S.R. 1999. Molecular identification of the ryanodine receptor pore-forming segment. J. Biol. Chem. 37:25971-25974.