Seminiferous tubules from 20-day-old rats were isolated by mechanical dissection and the conditions to produce optimal inhibin secretion over a 5-day period of culture were established. Inhibin was measured by a specific heterologous radioimmunoassay and by an in vitro bioassay using rat pituitary cells in culture. The tubule production of biologically and immunologically active inhibin was stimulated by ovine follitropin (FSH); however, the ratio of biological to immunological (B : I) activity fell significantly with increasing dose. A similar stimulation of both B and I inhibin activity with a corresponding decrease in B : I ratio with increased FSH dose was also observed using isolated immature Sertoli cells in culture. Fractionation of seminiferous tubule and Sertoli cell culture media by reverse-phase high-performance liquid chromatography (HPLC) revealed the presence of two peaks (I and II) with inhibin biological and immunological activity both of which increased following FSH stimulation. However, while the B : I ratio for peak I remained unchanged following FSH stimulation, the B : I ratio for peak II significantly fell. The molecular weights of peak I and II immunoactivity, determined following fractionation on preparative polyacrylamide gel electrophoresis were 30 kDa and 27 kDa respectively. The 30 kDa peak, based on its inhibin in vitro biological and immunological activity, molecular weight and retention position on HPLC most likely represent 30-32 kDa inhibin. The 27 kDa material remains to be identified.
- Sertoli cell