Ca2+ Release in Muscle Fibers Expressing R4892W and G4896V Type 1 Ryanodine Receptor Disease Mutants

The large and rapidly increasing number of potentially pathological mutants in the type 1 ryanodine receptor (RyR1) prompts the need to characterize their effects on voltage-activated sarcoplasmic reticulum (SR) Ca2+ release in skeletal muscle. Here we evaluated the function of the R4892W and G4896V RyR1 mutants, both associated with central core disease (CCD) in humans, in myotubes and in adult muscle fibers. For both mutants expressed in RyR1-null (dyspedic) myotubes, voltage-gated Ca2+ release was absent following homotypic expression and only partially restored following heterotypic expression with wild-type (WT) RyR1. In muscle fibers from adult WT mice, both mutants were expressed in restricted regions of the fibers with a pattern consistent with triadic localization. Voltage-clamp-activated confocal Ca2+ signals showed that fiber regions endowed with G4896V-RyR1s exhibited an,30 % reduction in the peak rate of SR Ca2+ release, with no significant change in SR Ca2+ content. Immunostaining revealed no associated change in the expression of either a1S subunit (Cav1.1) of the dihydropyridine receptor (DHPR) or type 1 sarco(endo)plasmic reticulum Ca2+ ATPase (SERCA1), indicating that the reduced Ca2+ release resulted from defective RyR1 function. Interestingly, in spite of robust localized junctional expression, the R4892W mutant did not affect SR Ca2+ release in adult muscle fibers, consistent with a low functional penetrance of this particular CCD-associated mutant