Sarcolemmal Biomechanics and Excitability in Malformed Muscle Fibers of Dystrophic Mice

Abstract

Background: Duchenne muscular dystrophy (DMD), the most common and severe dystrophy, is caused by the absence of dystrophin. Muscle weakness and fragility (i.e. increased susceptibility to damage) are presumably due to structural weakness of the myofiber cytoskeleton, but recent studies suggest that malformed/split myofibers in dystrophic muscle may also play a role. We have previously studied the biomechanical properties of the sarcolemma in single myofibers isolated mechanically from extensor digitorum longus (EDL) muscles in wild-type (WT) and dystrophic (mdx, mouse model for DMD) mice. PURPOSE: We use similar biomechanical methods on enzymatically-dissociated myofibers (both normal and malformed) from the flexor digitorum brevis muscle (FDB) of WT and mdx mice.Methods: FDB muscles were enzymatically-dissociated and plated on specialized coverslips. Suction pressures (P) applied through a pipette to the membrane generated a bleb, which increased in height with increasing P. Larger increases in P ruptured the connections between the sarcolemma and myofibrils (speration P, or SP) and eventually caused the sarcolemma to burst. We also examined excitability using high-speed confocal microscopy and the voltage-sensitive indicator di-8-butyl-amino-naphthyl-ethylene-pyridinium-propyl-sulfonate to assess the action potential (AP).Results: The mechanical results from dissociated FDB myofibers match findings from dissected EDL myofibers, but SP was up to 14-fold higher in the FDB than EDL. SP was 27% lower in mdx myofibers and 50% less in branches of split mdx fibers compared to the trunk. AP amplitude was not altered in between groups, but this work is ongoing.Conclusions: Data indicate a reduction in muscle stiffness, increased sarcolemmal deformability and instability in mdx muscle. This approach corroborates the labor-intensive data obtained from single fiber dissection and allows a facile high throughput model. Findings suggest mechanical differences due to altered morphology, despite comparable excitability.