Counteracting Ethanol Stress: EPS Improves Energy and Muscle Regeneration in C2C12 Myotubes

Alcohol-induced skeletal muscle dysfunction is characterized by impaired mitochondrial bioenergetics, oxidative stress, and reduced regenerative capacity. Chronic or repeated exposure to ethanol disrupts myoblast differentiation, contributing to deficits in muscle repair and functional recovery. Exercise-mimetic electrical pulse stimulation (EPS) has emerged as a promising approach to enhance mitochondrial metabolism and improve myogenic potential; however, its capacity to counteract ethanol-induced bioenergetic and differentiation impairments remains unclear. Our goal was to evaluate if EPS would restore ATP production, reduce oxidative stress, or improve myotube formation in C2C12 cells exposed to ethanol. C2C12 myoblasts were differentiated for 7 days in the presence or absence of ethanol (50 mM). Cells received an EPS protocol that mimics resistance exercise (25 Hz, 1-s train every 5 s for 1 hour daily) throughout the differentiation period. Bioenergetic function was assessed via ATP quantification using the Promega's luminescence ATP detection assay. Hydrogen peroxide release was measured using Amplex Red fluorescence. Differentiation indices, including myotube density and fusion index, were quantified following HEMA 3 staining. A student t-test analysis with appropriate post-hoc was performed to evaluate statistical significance. Results showed that there was a decreased in fusion index in the presence of ethanol compared to control (p< 0.05), and that EPS showed an increased in fusion index alone compared to control (p< 0.01) as well as in the presence of ethanol (p< 0.01). Similarly, there was a main effect of ethanol to decrease the number of myotubes per field (p< 0.001) and a main effect of EPS to increase it (p< 0.05). Preliminary findings show that EPS showed a trending increase in ATP production compared to ethanol alone (p=0.08). ROS production was increased by ethanol (p< 0.05) and this was ameliorated by EPS (p=0.08). These results suggest that chronic EPS during differentiation enhances mitochondrial bioenergetics, reduces oxidative stress, and amelioratesethanol-induced deficits in myogenic differentiation. Ongoing studies will explore molecular mechanisms underlying EPS-mediated metabolic and regenerative improvements, including effects on mitochondrial dynamics, redox signaling, and myogenic regulatory factors. This research was supported by F32AA031901 (KMRG), P60AA009803 (PEM), K01AA024494 (LS), and R21AA030869 (LS). This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.