The relative binding affinities of testosterone (T), 19-nortestosterone (N) and their 5 alpha-reduced derivatives: 5 alpha-dihydrotestosterone (DHT) and 5 alpha-dihydro-19-nortestosterone (DHN) to the androgen receptor of the rat seminal vesicle was studied using competition experiments. In cell-free extracts incubated at +10 degrees C for 18 h the relative binding affinities of these steroids (DHT greater than T = DHN = N) proved to be specific for the androgen receptor, in the sense that only prostatic extracts gave a similar result while three other androgen binding proteins (human sex steroid binding globulin, rat epididymal androgen binding protein and an antibody raised against T) exhibited quite different binding specificities. In minced seminal vesicles incubated at 37 degrees C for 1 h the binding affinities showed marked differences (DHT greater than N greater than T greater than or equal to DHN) and similar patterns were observed with both the cytoplasmic and the nu clear receptors. Our findings suggest that (I) the simultaneous presence of a 4-ene double bond and 19-methyl group in T does not favor the tight binding of T to the androgen receptor; therefore, either saturation of this double bond or elimination of the 19-methyl group leads to increased binding and (II) while 5 alpha-reduction of T increases the affinity of this steroid to the receptor, that of N does not influence or rather tends to decrease the binding affinity. The opposite changes observed in the binding affinities of T and N after their 5 alpha-reduction may account for the lower androgenicity of N. On the other hand, the relative myotropic activity in vivo of these steroids is apparently determined by the ratio of their affinities (N/T approximately 3 at 37 degrees C) to the androgen receptor.