The purpose of this study was to determine if the SPRINT exercise protocol provided protection to MHC I and IIa myonuclear and satellite cell number in the vastus lateralis and soleus during 70 days of bedrest. Twenty-five subjects (24 M, 1F) were recruited, screened, and randomized into one of three groups: bedrest only (BR; n=8, 37±3 yrs), bedrest with resistance and aerobic exercise (BRE; n=9, 34±2 yrs), and bedrest with resistance and aerobic exercise concurrent with low dose of testosterone injections (BRE+T; n=8, 33±3 yrs). Muscle biopsies from the vastus lateralis and soleus were sectioned, immunofluorescently stained, and analyzed for myonuclear and satellite cell number, along with fiber size and fiber type composition. Myonuclear number in MHC I and IIa fibers of the vastus lateralis and soleus were unchanged (P>0.05) from pre- to post-bedrest within BR, BRE, and BRE+T. However, there was a main time effect (P<0.05) for a decrease in myonuclear domain for vastus lateralis MHC I fibers (-8%), as well as soleus MHC I (-18%) and IIa (-18%) fibers. Also, there was a trend for a main effect for time (P=0.059) for an increase (+37%) in satellite cell number for vastus lateralis MHC IIa fibers, which was primarily driven by the exercise groups (BR: -4%, BRE: +36%, BRE+T: +67%). The main effect of time that was observed in the myonuclear domain was mirrored by the main time effect in fiber atrophy in vastus lateralis MHC I fibers (-10%), as well as soleus MHC I (-21%) and IIa (-23%) fibers. Unloading did not reduce MHC I fiber proportion in both the vastus lateralis and soleus (P>0.05). BR trended to decrease vastus lateralis MHC IIa fiber composition (P=0.065) while BRE and BRE+T increased the vastus lateralis MHC IIa. Additionally, BRE decreased total hybrid proportions in the vastus lateralis (P<0.05). In the soleus there was a main time effect in the increase of total hybrids (P<0.05). Prolonged unloading did not alter myonuclei and satellite cells in both the vastus lateralis and soleus, but SPRINT may enhance satellite cell number in the vastus lateralis MHC IIa fibers. The decline in myonuclear domain was driven by the muscle fiber atrophy in the vastus lateralis MHC I fibers, and in soleus MHC I and IIa fibers. The results suggest that the SPRINT exercise is effective at maintaining fiber size, and consequently preserving myonuclear domain in both the vastus lateralis and soleus. The current data along with prior findings within this cohort support SPRINT as an effective countermeasure for protecting myocellular characteristics and function during prolong unloading.