LOCALIZATION OF INHIBIN AND ACTIVIN-BINDING SITES IN THE TESTIS DURING DEVELOPMENT BY IN-SITU LIGAND-BINDING

Inhibin and activin are related proteins thought to be potential parac rine regulators of testicular development and maintenance of spermatog enesis. Messenger RNA and proteins immunologically related to both fac tors have been identified in the adult testis. However, the role(s) of these factors in paracrine regulation of testicular function is poorl y understood. To identify potential targets for inhibin and activin in immature and adult testis, we used in situ binding of [I-125]-labeled ligands to localize and describe the distribution of binding sites fo r inhibin and activin in testes of 15-, 18-, 21-, 30-, 45-, and 60-day -old rats. Nonspecific binding was defined as that occurring in the pr esence of a 1000-fold excess of unlabeled recombinant human (rh) inhib in or activin. [I-125]-Inhibin was found to bind to interstitial cells throughout development. Inhibin binding was shown to co-localize with cells that showed positive staining for 3beta-hydroxysteroid dehydrog enase (3beta-HSD). Competition studies demonstrated that this binding was indeed specific for inhibin. In contrast, [I-125]-activin showed t wo distinct patterns of binding. First, [I-125]-activin was shown to b ind in a non-stage-dependent manner to cells located in the basal comp artment of the seminiferous tubules in testis obtained from animals of all ages studied. Binding of [I-125]-activin in the periphery of the tubule could be inhibited entirely by coincubation with excess unlabel ed activin and partially with excess unlabeled inhibin. The ability of inhibin to compete with activin for binding appeared to be more prono unced in younger animals. In 45- and 60-day-old animals, a second stag e-dependent component of [I-125]-activin binding was also apparent. Th is binding was localized to spermatids found in stage VII-VIII tubules and was inhibited by the presence of excess activin, but not inhibin. These results indicate that inhibin can bind specifically to testicul ar interstitial cells throughout development and may be an important r egulator of Leydig cell testosterone production or interstitial cell f unction. In contrast, activin appears to bind in a specific and stage- dependent manner to receptors or high-affinity binding proteins on spe rmatids as well as to sites on the periphery of all seminiferous tubul es. These results support the hypothesis that both activin and inhibin may act at several levels to regulate proliferation or differentiatio n of germ and Sertoli cell function as well as to modulate interstitia l cell activity.