The overall goal of this project is to define the physiological basis for enhanced excitability and abnormal responses to anesthetics inherent in muscles of malignant hyperthermia susceptible (MHS) humans and pigs. 1) First we will test the hypothesis that myotubes cultured from neonatal MHS muscles exhibit abnormalities characteristic of MHS muscles. Fluorescence imaging of the calcium indicator fura will be used to determine whether myoplasmic calcium of MHS myotubes is abnormally elevated after exposure to halothane (which triggers MH episodes). The strength-duration relationship of MHS and normal myotubes will be determined using whole cell patch-clamp techniques to establish that the contractile threshold of MHS myotubes is lower than normal. 2) Excitation-contraction (EC) coupling in MHS skeletal muscles appears to be altered at the level of the transverse-tubules (TT). We will test whether MHS myotubes are functionally defective at the TT level by recording voltage-dependent TT signals associated with EC coupling, i.e., the slow activating, long lasting calcium current (I-s) and intramembrane charge-movement. Whole cell patch-clamp techniques will be used to characterize I-s and charge-movement as to voltage-dependence of activation and inactivation and sensitivity to calcium channel antagonists to determine whether any of these parameters are altered in MHS myotubes. Dantrolene and azumolene, EC coupling inhibitors which prevent MH episodes, will be studied to determine whether they alter I-s and/or charge-movement. 3) Finally, we will test the hypothesis that the known sarcoplasmic reticulum (SR) abnormality of MHS muscles alters function at the TT level. Muscles heterozygous for the MH gene, which codes for the SR calcium channel exhibit some of the same EC-coupling abnormalities as homozygous MHS muscles. Myotubes cultured from piglets heterozygous for the MH gene will be used to determine whether the presence of a subpopulation of abnormal SR calcium channels results in altered I-s and charge movement in the TTs. SR influence on TT function will be further explored by recording whole cell I-s and charge-movement of normal myotubes in the presence of one of three drugs which act at the SR calcium channel: Ryanodine which binds to SR calcium channels and holds them in a low conductance state, and halothane or caffeine both of which act at the SR calcium channel to enhance calcium release. In sum, these studies will address whether, in addition to the well documented SR calcium channel defect, there is also an abnormality in I-s and/or charge movement in MHS muscles and whether abnormal SR calcium channels can influence events at the TT level. Elucidation of the EC coupling defect in MHS muscles will be of importance for understanding the basis for initiation of NM and contribute to clarifying an as yet undefined portion of the EC coupling process of skeletal muscles.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR041270-03
Application #
2080587
Study Section
Surgery, Anesthesiology and Trauma Study Section (SAT)
Project Start
1992-08-10
Project End
1996-07-31
Budget Start
1994-08-01
Budget End
1995-07-31
Support Year
3
Fiscal Year
1994
Total Cost
Indirect Cost
Name
University of Minnesota Twin Cities
Department
Veterinary Sciences
Type
Schools of Veterinary Medicine
DUNS #
168559177
City
Minneapolis
State
MN
Country
United States
Zip Code
55455
Lentz, L R; Valberg, S J; Balog, E M et al. (1999) Abnormal regulation of muscle contraction in horses with recurrent exertional rhabdomyolysis. Am J Vet Res 60:992-9
Balog, E M; Gallant, E M (1999) Modulation of the sarcolemmal L-type current by alteration in SR Ca2+ release. Am J Physiol 276:C128-35
Prakash, Y S; van der Heijden, H F; Gallant, E M et al. (1999) Effect of beta-adrenoceptor activation on [Ca2+]i regulation in murine skeletal myotubes. Am J Physiol 276:C1038-45
Enzmann, N R; Balog, E M; Gallant, E M (1998) Malignant hyperthermia: effects of sarcoplasmic reticulum Ca2+ pump inhibition. Muscle Nerve 21:361-6
Gallant, E M; Jordan, R C (1996) Porcine malignant hyperthermia: genotype and contractile threshold of immature muscles. Muscle Nerve 19:68-73
Gallant, E M; Balog, E M; Beam, K G (1996) Slow calcium current is not reduced in malignant hyperthermic porcine myotubes. Muscle Nerve 19:450-5
Gallant, E M; Lentz, L R; Taylor, S R (1995) Modulation of caffeine contractures in mammalian skeletal muscles by variation of extracellular potassium. J Cell Physiol 165:254-60