Endoplasmic reticulum (ER) stress is linked to insulin resistance and several muscle diseases, including myopathies. ER defects are also implicated in skeletal muscle dysfunction associated with aging. Physical contact between the ER and mitochondria is essential for the local transport of materials and effective signal transduction between these organelles. Nicotinamide phosphoribosyltransferase (NAMPT) regulates mitochondrial biogenesis and maintains skeletal muscle function and integrity. Our study aims to investigate the effect of the NAMPT activator P7C3-A20 on tunicamycin-induced ER stress using an in-vitro myoblast cell model. We found that tunicamycin treatment decreased the expression of several mitochondrial proteins, such as NAMPT and the mitophagy regulator PINK1 in C2C12 murine myoblast cells. It also reduced the phosphorylation of Drp1, a master regulator of mitochondrial fission, in C2C12 cells treated with tunicamycin. JC-1 imaging revealed that treatment with P7C3-A20 for 6 h at concentrations of 1, 5, and 10 μM increased mitochondrial potential in C2C12 cells. Moreover, P7C3-A20 ameliorated tunicamycin-induced cell death in a concentration-dependent manner. In conclusion, the NAMPT activator P7C3-A20 can mitigate tunicamycin-induced cell damage in C2C12 murine myoblasts. Activation of NAMPT is a potential novel therapeutic approach for muscle diseases associated with ER stress, such as sarcopenia.