We tested the hypotheses that: (1) Ca2+ handling and force production would be irreversibly altered in skeletal muscle during steady-state contractions when subjected to severe, prolonged hypoxia and subsequent reoxygenation; and (2) application of the cardio-protective drug trimetazidine would attenuate these alterations. Single, living skeletal muscle fibres from Xenopus laevis were injected with the Ca2+ indicator fura 2, and incubated for 1 h prior to stimulation in 100 µM TMZ-Ringer solution (TMZ; n = 6) or standard Ringer solution (CON; n = 6). Force and relative free cytosolic Ca2+ concentration ([Ca2+]c) were measured during continuous tetanic contractions produced every 5 s as fibres were sequentially perfused in the following manner: 3 min high extracellular PO2 (159 mmHg), 15 min hypoxic perfusion (3-5 mmHg) then 3 min high PO2. Hypoxia caused a decrease in force and peak [Ca2+]c in both the TMZ and CON fibres, with no significant (P < 0.05) difference between groups. However, basal [Ca2+]c was significantly lower during hypoxia in the TMZ group vs. the CON group. While reoxygenation generated only modest recovery of relative force and peak [Ca2+]c in both groups, basal [Ca2+]c remained significantly less in the TMZ group. These results demonstrated that in contracting, single skeletal muscle fibres, TMZ prevented increases in basal [Ca2+]c generated during a severe hypoxic insult and subsequent reoxygenation, yet failed to protect the cell from the deleterious effects of prolonged hypoxia followed by reoxygenation. Experimental Physiology (2003) 88.3, 415-421.