The objective of the present repeat-measures study was to determine whether plasma serum levels of testosterone, cortisol, osteocalcin or type I collagen C-telopeptide (CT) are acutely affected following an electro-myostimulation (EMS) bout, and their relation to bone mineral density and muscle mass. Ten men with recent (8 weeks) thoracic spinal cord injury (SCI) (ASIA A) and 10 age-matched able-bodied (AB) men performed one EMS bout on the quadriceps femoris muscle. Blood samples were drawn at basal condition, immediately after EMS, and 15 min, 30 min, 24 h and 48 h post-EMS. Muscle cross-sectional area was measured by magnetic resonance imaging. Bone mineral density (BMD) was determined by dual-energy X-ray absorptiometry. In the SCI group, a significant decrease in testosterone, cortisol and CT together with a significant increase in testosterone/cortisol ratio and osteocalcin/CT ratio was observed after EMS. For the AB subjects, only testosterone and CT decreased significantly following EMS. Muscle size was only related to testosterone/cortisol ratio in the SCI sample (R = 0.659, p < 0.05), whereas BMD did not show any relation to any biomarker. Acute EMS in recent spinal cord injured men seems to induce positive effects on bone turnover biomarkers, and anabolic and catabolic hormones.