Hypothalamic cytosol receptor binding of gonadal steroids was studied in both male and female intact and castrate rats. Assessment of specific binding by the protamine precipitation technique indicated a capacity of 1.34–2.03 × 10−14 moles estradiol bound/mg protein among the animal models studied. Although testosterone and 5α-dihydrotestosterone (17β-hydroxy-5αandrostane-3-one) also showed a small amount of binding to cytosol, further experimentation showed that this binding did not obey the characteristic saturability phenomena of high affinity receptor systems as did the estradiol binding system. Quantification of the estradiol receptor interaction was estimated as 1.48 and 1.55 × 10−14 moles/mg protein for male and female, respectively, by gel filtration, with elution of the complex in a peak just following the void volume. A specific 8S-sedimenting estradiol binding component was demonstrated on sucrose density gradient centrifugation. Patterns from male and female cytosol samples were very similar and no androgen binding component could be demonstrated in either sex. Binding analyses at intervals during the incubation periods employed showed that no loss of receptor was occurring due to instability. The nature and extent of the highaffinity receptor binding in the animal models studied was determined by construction of estradiol binding curves. Analysis of these curves showed equivalent binding in the range of 1.3 to 2.2 × 10−14 moles/mg protein with association constants of 4.0 to 6.7×1010 M−1. Thus, steroid feedback control over cyclicity of gonadotropin secretion cannot be attributed to physical differences in nature or amount of cytoplasmic gonadal steroid receptor in the hypothalamus. (Endocrinology94: 785, 1974)