Hypoxia is known to reduce isometric contractile properties of isolated rat diaphragm bundles. Its effect on isotonic contractile properties (i.e. force-velocity relationship and power output) has not been studied. We hypothesized that hypoxia reduces velocity of shortening and consequently power output of the unfatigued muscle, and shortens endurance time during isotonic contractions. Force- velocity relationship, power output, and fatigue resistance of rat diaphragm muscle bundles were measured during hypoxia (PO2: 6.6 ± 0.2 kPa) and compared with hyperoxia (PO2: 91.8 ± 0.7 kPa). Force was clamped from 1 to 100% of maximal tetanic force (Po). Fatigue during isotonic contractions was induced by repeated stimulation every 2 s at a clamp level of 33% of Po. Hypoxia did not affect isometric force generation compared with hyperoxia, nor contraction or relaxation time. In contrast, maximum shortening velocity decreased significantly (hypoxia: 4.2 ± 0.3, hyperoxia: 6.0 ± 0.2 Lo/s, P < 0.05). The force-velocity curve shifted downwards (P < 0.05). Hypoxia lowered power output at each load compared with hyperoxia (P < 0.05). The isotonic endurance time was shorter during hypoxia compared with hyperoxia (80 ± 2 vs. 130 ± 3 s, P < 0.05). These data show that hypoxia depresses isotonic contractile properties and power output, and reduces endurance time during repeated isotonic contractions.