My laboratory focuses on two central questions: 1) What are the structural domains and essential amino acids that control critical functional properties of these two muscle Ca²⁺ channels and 2) How are these Ca²⁺ channel functions and EC coupling altered in diseased human skeletal muscle. My laboratory utilizes tissue culture, molecular biology (site-directed and chimeric mutagenesis), whole-cell and single channel patch clamp, and fluorescence measurements of intracellular Ca²⁺ to begin to address these questions.

Currently, one project is taking advantage of a transgenic line of mice in which the gene encoding the protein that forms the skeletal muscle SR Ca²⁺ release channel (RyR1) has been disrupted through gene targeting (RyR1-knockout or dyspedic mice). Skeletal myotubes obtained from mice homozygous for this mutation contain plasma membrane L-channels, but lack the SR Ca²⁺ release channel. These mice are being used to understand the cellular abnormalities that are seen in malignant hyperthermia (MH) and central core disease (CCD), two related human skeletal muscle diseases. Both MH and CCD are caused by different point mutations (MH=8, CCD=5) in the RyR1 protein which result in a disturbance in skeletal muscle Ca²⁺ homeostasis. However, the precise nature of this disturbance in intracellular Ca²⁺ regulation has yet to be elucidated. We are beginning to characterize the physiologic effects of expressing the mutant MH and CCD RyR1s in RyR1-deficient dyspedic myotubes. Our experiments will begin to unravel the mystery of how these RyR1 mutations alter muscle Ca²⁺ homeostasis, and subsequently, result in the clinical manifestations associated with each disease.

Dirksen, R.T. and Avila, G.  (2004)  Distinct alterations in Ca2+ handling caused by malignant hyperthermia and central core disease mutations in the ryanodine receptor.  Biophys. J. 87:3193-3204.

Du, G.G., Avila, G., Sharma, P., Khanna, V.K., Dirksen, R.T., and MacLennan, D.H.  (2004)  Role of the sequence surrounding predicted transmembrane helix M4 in membrane association and function of the Ca2+ release channel of skeletal muscle sarcoplasmic reticulum (ryanodine receptor isoform 1).  J. Biol. Chem. 279:37566-37574.

Lueck, J.D., Goonasekera, S., and Dirksen, R.T.  (2004)  Ryanodinopathies: Muscle disorders linked to mutations in ryanodine receptors.  Basic and Applied Myology, 14:277-283.

Lyfenko A., Goonasekera, S., and Dirksen, R.T.  (2004)  Dynamic alterations in myoplasmic Ca2+ in malignant hyperthermia and central core disease.  Biochem. Biophys. Res. Commun. 322:1256-1266.

Weiss, R.E., O’Connell, K.M.S., Flucher, B.E., Allen, P.D., Grabner, M., and Dirksen, R.T.  (2004)  Altered bi-directional coupling by the R1086H malignant hyperthermia mutation in the skeletal muscle dihydropyridine receptor.  Am. J. Physiol. 287:C1094-C1102.

Avila, G. and Dirksen, R.T.  (2005)  Rapamycin and FK506 reduce skeletal muscle voltage sensor expression and function. Cell Calcium 38:35-44.

Beutner, G,. Sharma, V.K., Olbinski, B., Lin, L., Valdivia, H.H., Dirksen, R.T., and Sheu, S-S.  (2005)  Type 1 ryanodine receptor in cardiac mitochondria: Transducer of excitation-metabolism coupling.  Biochim. Biophys. Acta 1717:1-10.

Dirksen, R.T., and Avila, G.  (2005)  Pathophysiology of muscle disorders linked to mutations in the skeletal muscle ryanodine receptor.  In Ryanodine Receptors:  Structure, Function and Dysfunction in Clinical Disease, eds., Wehrens, X.H. and Marks, A.R., Kluwer Academic Publishers, New York, New York, 229-242.

Goonasekera, S.A., Chen, S.R.W, and Dirksen, R.T.  (2005)  Reconstitution of local calcium signaling between cardiac dihydropyridine and ryanodine receptors: Novel insights into regulation by FKBP12.6.  Am. J. Physiol. 289:C1476-C1484.

Kimura, T., Nakamori, M., Lueck, J.D., Pouliquin, P., Aoike, F., Fujimura, H., Takahashi, M.P., Dulhunty, A.F., and Sakoda, S.  (2005)  Altered mRNA splicing of the skeletal muscle ryanodine receptor and sarcoplasmic/endoplasmic reticulum Ca2+ ATPase in myotonic dystrophy.  Hum. Mol. Genet. 14:2189-2000.