Loss of calpain3b in Zebrafish, a Model of Limb-Girdle Muscular Dystrophy, Increases Susceptibility to Muscle Defects Due to Elevated Muscle Activity.

Limb-Girdle Muscular Dystrophy Type R1 (LGMDR1; formerly LGMD2A), characterized by progressive hip and shoulder muscle weakness, is caused by mutations in . In zebrafish, mediates Def-dependent degradation of p53 in the liver and intestines. We show that is expressed in the muscle. To model LGMDR1 in zebrafish, we generated three deletion mutants in and a positive-control mutant (Duchenne muscular dystrophy). Two partial deletion mutants showed transcript-level reduction, whereas the RNA-less mutant lacked mRNA. All homozygous mutants were developmentally-normal adult-viable animals. Mutants in were homozygous-lethal. Bathing wild-type and mutants in 0.8% methylcellulose (MC) for 3 days beginning 2 days post-fertilization resulted in significantly pronounced (20-30%) birefringence-detectable muscle abnormalities in mutant embryos. Evans Blue staining for sarcolemma integrity loss was strongly positive in homozygotes, negative in wild-type embryos, and negative in MC-treated mutants, suggesting membrane instability is not a primary muscle pathology determinant. Increased birefringence-detected muscle abnormalities in mutants compared to wild-type animals were observed following induced hypertonia by exposure to cholinesterase inhibitor, azinphos-methyl, reinforcing the MC results. These mutant fish represent a novel tractable model for studying the mechanisms underlying muscle repair and remodeling, and as a preclinical tool for whole-animal therapeutics and behavioral screening in LGMDR1.