Maintaining mitochondrial NAD+ homeostasis is key for heat-induced skeletal muscle injury prevention despite presence of intracellular cation alterations.

Abstract

Mitochondrial dysfunction is implicated in heat-induced skeletal muscle injury and its underlying mechanisms remain unclear. Evidence suggests that cellular ions and molecules, including divalent cations and adenine nucleotides, are involved in the regulation of mitochondrial function. In this study, we examined Ca2+, Mg2+, and NAD+ levels in mouse C2C12 myoblasts and skeletal muscle in response to heat exposure. During heat exposure, mitochondrial Ca2+ levels increased significantly, whereas cytosolic C2+ levels remained unaltered. The mitochondrial Ca2+ levels in the skeletal muscle of heat-exposed mice were 28% higher, compared to control mice. No changes in cytosolic Ca2+ were detected between the two groups. Following heat exposure, cytosolic and mitochondrial Mg2+ levels were reduced by 47% and 23% in C2C12 myoblasts, and by 51% and 44% in mouse skeletal muscles, respectively. In addition, heat exposure decreased mitochondrial NAD+ levels by 32% and 26% in C2C12 myoblasts and mouse skeletal muscles, respectively. Treatment with the NAD+ precursor nicotinamide riboside (NR) partially prevented heat-induced depletion of NAD+. Additionally, NR significantly reduced heat-increased mitochondrial fission, mitochondrial depolarization, and apoptosis in C2C12 myoblasts and mouse skeletal muscles. No effects of NR on heat-induced changes in intracellular Ca2+ and Mg2+ levels were observed. This study provides the in vitro and in vivo evidence that acute heat stress causes alterations in mitochondrial Ca2+, Mg2+, and NAD+ homeostasis. Our results suggest mitochondrial NAD+> homeostasis as a therapeutic target for the prevention of heat-induced skeletal muscle injury.