Physical exertion exacerbates decline in the musculature of an animal model of Duchenne muscular dystrophy

Abstract

Duchenne muscular dystrophy (DMD) is a genetic disorder caused by loss of the protein dystrophin. In humans, DMD has early onset, causes developmental delays, muscle necrosis, loss of ambulation, and death. Current animal models have been challenged by their inability to model the early onset and severity of the disease. It remains unresolved whether increased sarcoplasmic calcium observed in dystrophic muscles follows or leads the mechanical insults caused by the muscle’s disrupted contractile machinery. This knowledge has important implications for patients, as potential physiotherapeutic treatments may either help or exacerbate symptoms, depending on how dystrophic muscles differ from healthy ones. Recently we showed how burrowing dystrophic (dys-1) C. elegans recapitulate many salient phenotypes of DMD, including loss of mobility and muscle necrosis. Here, we report that dys-1 worms display early pathogenesis, including dysregulated sarcoplasmic calcium and increased lethality. Sarcoplasmic calcium dysregulation in dys-1 worms precedes overt structural phenotypes (e.g., mitochondrial, and contractile machinery damage) and can be mitigated by reducing calmodulin expression. To learn how dystrophic musculature responds to altered physical activity, we cultivated dys-1 animals in environments requiring high intensity or high frequency of muscle exertion during locomotion. We find that several muscular parameters (e.g., size) improve with increased activity. However, longevity in dystrophic animals was negatively associated with muscular exertion, regardless of effort duration. The high degree of phenotypic conservation between dystrophic worms and humans provides a unique opportunity to gain insight into the pathology of the disease as well as the initial assessment of potential treatment strategies.

Pluronic gel-based burrowing assay for rapid assessment of neuromuscular health inC. elegans

“…Under standard assay conditions of 26 % w/w, we observed that animals burrow with a mean velocity of 1.14 mm/min and undulatory frequency of 0.07 Hz. In contrast, at 3 % w/v agar, animal velocity was found to be 2 mm/min [19] and at 9 % w/v agar the undulatory frequency was reported to be 0.2 Hz [16]. Additionally, 3D behavioral studies in 3 % w/v gelatin report a velocity of 5 mm/min [17].…”

“…In this section, we harness these capabilities to demonstrate that burrowing can be used in diverse applications. None of these demonstrative applications and the results thereof have been shown by prior burrowing studies [16][17][18][19][20]. Since our intent is to demonstrate the power of the assay platform we designed, we consider our results in each of these applications as laying foundation for future in-depth investigations.…”

“…Even though burrowing of nematodes in soil-like environments has been previously studied [48][49][50], only recently has it been used for evaluation of neuromuscular health in C. elegans [16,18,19,22,51]. These recent investigations employed agar as a medium for burrowing.…”

“…Very few studies have exploited burrowing ability as a way to evaluate neuromuscular health in C. elegans. Since the original method was conceived [18], it has been applied to score muscular dystrophy mutants [16], and to probe calcium dysregulation and longevity outcomes due to physical exertion in the burrowing environment in these mutants [19]. Additionally, a burrowing assay has been used to assess mutants with defects in mitochondrial fission and fusion proteins [22].…”

“…Although existing methods have revealed important aspects of neuromuscular control of animal behavior [14], assay culture conditions are typically not the best representation of C. elegans natural habitat, in which animals burrow in three-dimensions in soil, rotten fruit, and fluid drops [15]. Indeed, burrowing environments to mimic this natural habitat have been designed to better reflect true behavioral conditions [16][17][18][19][20].…”