These experiments tested the hypothesis that in vitro diaphragmatic fatigue resistance is enhanced in animals treated with glucocorticoids. Female Sprague-Dawley rats (4 months old) were randomly assigned to a control (N =12) or glucocorticoid treatment group (N =12). Treatment animals were injected daily for 8 days with prednisolone (5 mg kg(-1)); control animals were injected with the same volume of the vehicle. Twenty-four hours after the last injection, the following in vitro diaphragmatic contractile properties were examined in costal diaphragm strips: maximal twitch (P(t)) half time to peak tension (1/2 TPT), half relaxation time (1/2 RT), force-frequency relationship, and the rate of fatigue development. Diaphragmatic fatigue was assessed by monitoring the decrease in force production (measured as percent of initial force) over a 60-min contractile period. The in vitro fatigue protocol incorporated a supramaximal stimulus delivered at 30 Hz every 2 s with a train duration of 250 ms (duty cycle 12.5%). Citrate synthase (CS), superoxide dismutase (SOD), and water content of the costal diaphragm were also determined. Glucocorticoid administration induced an 18.9% (P<0.05) decrease in animal body weight when compared to the control. Similar weight losses also occurred in the diaphragm with a decrease (P<0.05) in mass of 16.5% compared to the control. Furthermore, prednisolone treatment resulted in a significant reduction in the cross-sectional area (CSA) of type IIb fibres with no change in the CSA area of type I and IIa fibres. 1/2 TPT and 1/2 RT were significantly prolonged (P<0.05) in the glucocorticoid treated rats whereas the force-frequency curve was unaltered (P>0.05). Fatigue resistance was greater in the glucocorticoid group (P<0.05); the relative force production differed between groups at the end of 1 min of contractions and remained different throughout the 60-min fatigue protocol. Citrate synthase, SOD, and water content were not different between groups. These experiments support the hypothesis that costal diaphragm strips of glucocorticoid-treated rats possess a greater resistance to fatigue. We postulate that this fatigue resistance is due to glucocorticoid-induced changes muscle fibre type composition.