A nuclear progestogen receptor was identified in the ovary of the spotted seatrout, Cynoscion nebulosus. A single class of high-affinity, low-capacity cytoplasmic binding sites for 17 alpha,20 beta-dihydroxy-4-pregnen-3-one (17 alpha,20 beta-P) was characterized by saturation and Scatchard analyses (KD = 1.89 +/- 0.61 nM, Bmax = 1.80 +/- 0.63 pmol/g ovary, n = 4), as well as by one-point assay (Bmax = 1.41 +/- 0.26 pmol/g ovary, n = 12). Analysis of the binding kinetics indicated a fairly rapid association rate (T1/2 = 72 +/- 10.2 min) and a slightly slower dissociation rate (T1/2 = 99 +/- 9.4 min). Competition studies revealed that several steroids exhibited the same range of affinity (17 alpha, 20 beta-P > 17 alpha,20 beta,21-trihydroxy-4-pregnen-3-one (20 beta-S) > 11-deoxycorticosterone > progesterone) while others displayed an order of magnitude less affinity (17 alpha-hydroxy-4-pregnen-3-one > pregnenolone > 11-deoxycortisol > testosterone). No displacement was found with 1000-fold excess estradiol-17 beta or cortisol. Binding activity was also present within the testis, but not in the brain, gill, muscle, or plasma. Nuclear binding was detected by DNA-cellulose column chromatography and was inhibited by the addition of molybdate, a characteristic of nuclear steroid receptors.(ABSTRACT TRUNCATED AT 250 WORDS)