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Fifteen middle-aged to older, overweight cats attending a first-opinion clinic were investigated to rule out hyperadrenocorticism as a cause of their weight problem, using two different protocols for the adrenocortlcotropic hormone (ACTH) stimulation test. The cats received intravenous synthetic ACTH (tetracosactrin) at an initial dose of 125 μg; a second test was performed between two and three weeks later, using a dose of 250 vg intravenously. The mean basal serum cortisol concentration was 203 nmol/litre (range 81 to 354 nmol/litre). The highest mean serum cortisol concentration occurred at 60 minutes following the 125 μg dose and at 120 minutes following the 250 μg dose. There was, however, no statistically significant difference between these peak cortisol concentrations attained using either dose of tetracosactrin. A significantly higher mean serum cortisol concentration was attained after the higher dose at the 180 minutes time point, indicating a more prolonged response when compared with the lower dose. The cats were followed up for one year after the initial investigations and none were found to develop hyperadrenocorticism during this time.