Extremity injuries with hemorrhage have been a significant cause of death in civilian medicine and on the battlefield. The use of a tourniquet as an intervention is necessary for treatment to an injured limb; however, the tourniquet and subsequent release results in serious acute ischemia-reperfusion (IR) injury in the skeletal muscle and neuromuscular junction (NMJ). Much evidence demonstrates that inflammation is an important factor to cause acute IR injury. To find effective therapeutic interventions for tourniquet-induced acute IR injuries, our current study investigated effect of dexamethasone, an anti-inflammatory drug, on tourniquet-induced acute IR injury in mouse hindlimb. In C57/BL6 mice, a tourniquet was placed on unilateral hindlimb (left hindlimb) at the hip joint for 3 h, and then released for 24 h to induce IR. Three hours of tourniquet and 24 h of release (24-h IR) caused gastrocnemius muscle injuries including rupture of the muscle sarcolemma and necrosis (42.8 ± 2.3% for infarct size of the gastrocnemius muscle). In the NMJ, motor nerve terminals disappeared, and endplate potentials were undetectable in 24-h IR mice. There was no gastrocnemius muscle contraction in 24-h IR mice. Western blot data showed that inflammatory cytokines (TNFα and IL-1β) were increased in the gastrocnemius muscle after 24-h IR. Treatment with dexamethasone at the beginning of reperfusion (1 mg/kg, i.p.) significantly inhibited expression of TNFα and IL-1β, reduced rupture of the muscle sarcolemma and infarct size (24.8 ± 2.0%), and improved direct muscle stimulation-induced gastrocnemius muscle contraction in 24-h IR mice. However, this anti-inflammatory drug did not improve NMJ morphology and function, and sciatic nerve-stimulated skeletal muscle contraction in 24-h IR mice. The data suggest that one-time treatment with dexamethasone at the beginning of reperfusion only reduced structural and functional impairments of the skeletal muscle but not the NMJ through inhibiting inflammatory cytokines.