ABSTRACT
Throughout a vertebrate organism's lifespan, skeletal muscle mass and function progressively decline. This age-related condition is termed sarcopenia. In humans, sarcopenia is associated with risk of falling, cardiovascular disease, and all-cause mortality. As the world population ages, projected to reach 2 billion older adults worldwide in 2050, the economic burden on the healthcare system is also projected to increase considerably. Currently, there are no pharmacological treatments for sarcopenia, and given the long-term nature of aging studies, high-throughput chemical screens are impractical in mammalian models. Zebrafish is a promising, up-and-coming vertebrate model in the field of sarcopenia that could fill this gap. Here, we developed a surface electrical impedance myography (sEIM) platform to assess skeletal muscle health, quantitatively and noninvasively, in adult zebrafish (young, aged, and genetic mutant animals). In aged zebrafish (~85% lifespan) as compared to young zebrafish (~20% lifespan), sEIM parameters (2 kHz phase angle, 2 kHz reactance, and 2 kHz resistance) robustly detected muscle atrophy (p < 0.000001, q = 0.000002; p = 0.000004, q = 0.000006; p = 0.000867, q = 0.000683, respectively). Moreover, these same measurements exhibited strong correlations with an established morphometric parameter of muscle atrophy (myofiber cross-sectional area), as determined by histological-based morphometric analysis (r = 0.831, p = 2 × 10-12; r = 0.6959, p = 2 × 10-8; and r = 0.7220; p = 4 × 10-9, respectively). Finally, the genetic deletion of gpr27, an orphan G-protein coupled receptor (GPCR), exacerbated the atrophy of skeletal muscle in aged animals, as evidenced by both sEIM and histology. In conclusion, the data here show that surface EIM techniques can effectively discriminate between healthy young and sarcopenic aged muscle as well as the advanced atrophied muscle in the gpr27 KO animals. Moreover, these studies show how EIM values correlate with cell size across the animals, making it potentially possible to utilize sEIM as a "virtual biopsy" in zebrafish to noninvasively assess myofiber atrophy, a valuable measure for muscle and gerontology research.
ABSTRACT
Age-related deficits in skeletal muscle function, termed sarcopenia, are due to loss of muscle mass and changes in the intrinsic mechanisms underlying contraction. Sarcopenia is associated with falls, functional decline, and mortality. Electrical impedance myography (EIM)-a minimally invasive, rapid electrophysiological tool-can be applied to animals and humans to monitor muscle health, thereby serving as a biomarker in both preclinical and clinical studies. EIM has been successfully employed in several species; however, the application of EIM to the assessment of zebrafish-a model organism amenable to high-throughput experimentation-has not been reported. Here, we demonstrated differences in EIM measures between the skeletal muscles of young (6 months of age) and aged (33 months of age) zebrafish. For example, EIM phase angle and reactance at 2 kHz showed significantly decreased phase angle (5.3 ± 2.1 versus 10.7 ± 1.5°; p = 0.001) and reactance (89.0 ± 3.9 versus 172.2 ± 54.8 ohms; p = 0.007) in aged versus young animals. Total muscle area, in addition to other morphometric features, was also strongly correlated to EIM 2 kHz phase angle across both groups (r = 0.7133, p = 0.01). Moreover, there was a strong correlation between 2 kHz phase angle and established metrics of zebrafish swimming performance, including turn angle, angular velocity, and lateral motion (r = 0.7253, r = 0.7308, r = 0.7857, respectively, p < 0.01 for all). In addition, the technique was shown to have high reproducibility between repeated measurements with a mean percentage difference of 5.34 ± 1.17% for phase angle. These relationships were also confirmed in a separate replication cohort. Together, these findings establish EIM as a fast, sensitive method for quantifying zebrafish muscle function and quality. Moreover, identifying the abnormalities in the bioelectrical properties of sarcopenic zebrafish provides new opportunities to evaluate potential therapeutics for age-related neuromuscular disorders and to interrogate the disease mechanisms of muscle degeneration.
