We examined the effects of progesterone and some synthetic progestins and other steroids on the physical properties of the progesterone receptor of guinea pig uterine cytosol and on the binding of the receptor by nuclei. Progestational potency seemed to correlate with the ability to keep the receptor in the 7S form and to prevent dissociation into smaller subunits. The rate of activation prior to nuclear binding was slower with steroids of increasing progestational activity. Therefore activation in vitro may be unrelated to biological activity. Concentration of the cytosol led to a decrease in the equilibrium association constant. The extent of the decrease was less with progesterone than with its metabolite, 5 alpha-pregnanedione. When cytosol and nuclei were incubated in the absence of ligand measureable progesterone receptor was bound by the nuclei. The uncomplexed nuclear receptor bound [3H]-progesterone of [3H]-R5020 rapidly at 0 degrees, but progesterone-receptor complexes exchanged [3H]-progestin slowly at 0 degrees. Progesterone increased the amount of nuclear receptor at concentrations of 10(-9) and 10(-8)M, but decreased binding at higher concentrations. 5 alpha-Pregnanedione had the same effect as progesterone, but other metabolites of progesterone that had little affinity for the 7S progesterone receptor in cytosol had no effect on nuclear binding at any concentration. Glucocorticoids, testosterone and estradiol-17 beta increased the nuclear binding of the progesterone receptor when present at concentration of 10(-8)M and greater.