Enforced Exercise Induces Rapid Exhaustion in C. elegans Dystrophin Mutants
Authors: Carls Christopher , Phillips James , Krajacic Predrag 
Published: November 2023

Duchene Muscular Dystrophy (DMD) is an incurable genetic disease and those who suffer from it typically die in their second decade. The disease is currently incurable. The objective of this study was to facilitate cost effective high throughput drug screening for DMD. In order to achieve that we developed a worm treadmill platform. Using electrotaxis, the worm treadmill reveals a unique phenotype for dys‐1 (dystrophin) mutants comparable to DMD mouse model. Briefly, the nematode, C. elegans, displays a specific locomotion response to various stimuli including food, chemicals, and electricity. C. elegans’ response to electricity has been previously well described and dubbed electrotaxis ‐ the repeatable behavior of crawling toward the negative pole when subjected to an electric field. For the first time, we used this behavior to initiate and maintain locomotion in order to perform exhaustion tests.Materials and methodsThe “worm treadmill” consists of a specially formulated gel and buffer placed in a classic DNA‐electrophoresis box, a power source, a microscope camera, and an infrared thermometer to monitor gel temperature. 50–150 hypochlorite synchronized worms were transferred to the gel, and a voltage of ≥4 v/cm was applied. The poles of the electric field were alternated as the nematodes reached the end of the gel. The distance and disposition of crawling C. elegans was continuously tracked. The goal was to run the worms to a state of exhaustion – defined as a state when at least 75% of the worms ceased locomotion toward the anode. In order to assess the effects of therapeutics on exercised worms, hypochlorite synchronized eggs were placed on plates with 0.5mg/mL Prednisone, 0.05mg/mL Fluoxetine, or a combination of the two.ResultsWild type worms ran significantly longer and farther than dys‐1 mutants. In addition, we can report that dys‐1 mutants treated with prednisone, fluoxetine, and a combination of both drugs ran statistically significantly longer and farther than untreated dys‐1 mutants. The greatest improvement was observed in the dys‐1 mutants exposed to prednisone and fluoxetine; these worms ran significantly longer than all other groups, as well as significantly farther than untreated dys‐1 mutants or dys‐1 mutants exposed to prednisone or fluoxetine alone.ConclusionOur platform provides a unique phenotype for dys‐1 mutants comparable to DMD mouse model that was previously lacking. It offers a cost effective and simple way for initial screening of novel pharmacological compounds in order to find a cure for this currently incurable disease. In addition, as expected, based on animal models and previously established prednisone effects in other C. elegans mutants, Prednisone has shown therapeutic effects. However, this was the first time this has been shown in forcefully exercised dys‐1 (dystrophin) mutants. The greatest improvement observed with a single drug treatment was with fluoxetine, indicating serotonergic transmission may represent a novel therapeutic target for DMD. The greatest overall improvement was observed with simultaneous fluoxetine and prednisone treatment. This finding provides initial data suggesting that synergistic drug effects should be strongly considered in future screenings for potential DMD therapeutics.Support or Funding InformationThis work was funded by WVSOM intramural grant