1 Canrenone, the major active metabolite of spironolactone, decreased [3H]-progesterone binding to isolated uterine cytosolic progesterone receptors. The inhibition was concentration-dependent. 2 Canrenone did not alter [3H]-oestradiol binding to isolated uterine cytosolic oestrogen receptors. 3 Canrenone inhibition of progesterone binding to isolated cytosolic receptors was strictly competitive: Kd (apparent dissociation constant for progesterone binding) was increased in a concentration-dependent manner by canrenone, whereas Bmax (maximal number of progesterone binding sites/mg cytosolic protein) was unaltered. There was marked cooperativity in progesterone binding at high canrenone and low progesterone concentrations. The implication is that canrenone alters the subunit interaction of the receptor protein. 4 Kd for progesterone was 3.2 X 10(-9)M. Ki (the inhibition constant for canrenone with respect to progesterone binding) was 300 X 10(-9)M. Reports in the literature suggest that, following spironolactone administration, canrenone may rise to concentrations sufficiently high to inhibit progesterone binding. This action may contribute to the effect of spironolactone in inducing menstrual disturbances in female patients.