Porcine Pituitary Receptor Analysis Of Inhibin From Porcine Follicular Fluid

In these studies, techniques such as hormone micro-bioassay, cell culture, Radioirnmunoassay, radioreceptor assay were used to identify the receptor on the pituitary of sows and rat of inhibin which was purified from porcine follicular fluid in the pituitary of sows. The main content is as follows:1. Identification of the purity and the bioactivity of the porcine follicular fluid inhibin preparation.The aim of this experiment was to identify the purity of inhibin preparation of the porcine follicular fluid. To determine if the molecular weight of the inhibin preparation is consistent with that of the putative inhibin subunit and to identify the bioactivity of inhibin, the preparation were applied to SDS-PAGE. Results showed that the molecular weights of the two subunit of the preparation are 17-18 kDa and 14-15 kDa, respectively. Theses molecular weight are similar to that of the putative inhibin subunit a (18kDa) and P (14kDa) ; Two bands in the gel suggested that the preparation is of relatively pure.Pituitary cells isolated from pituitary of immature sows and rat were cultured in the serum free medium. Inhibin preparation were added to the medium and cultured for 48 hours, specific RIA was used to detected the content of FSH secreted by the pituitary cell. Results showed that this inhibin preparation significantly suppressed the secretion of FSH in a dose-dependent manner. This result suggested that the inhibin preparation had inhibin bioactivity.2. Studies on the inhibin receptorSow and rat were used as animal model to investigate inhibin receptor in sow and rat pituitary cells. The concentration of cellular memberane protein was measured by Bradford method and the final concentration was modulated above Img/ml. Radioligand assay results showed that specific binding site for the iodinated inhibin preparation existed on the pituitary cell of sows with a KD of 7.6 X 10~8Mol/L and a Rmax (the largest binding amount) of 50300fmol/g protein. The radioactivity in R^ tube of the rat group is lowerthan that of the non-specific tubes. Therefore, no binding site existed on the pituitary cells of the rat.In conclusion, the porcine follicular fluid inhibin preparation prepared in our laboratory are pure and and biologically active. Using iodinated inhibin preparation as ligand, Radioligand assay showed that there is inhibin binding site on the pituitary cell of the porcine, but not on the pituitary cell of the rat, which might suggeste that the porcine follicular fluid inhibin could bind to its binding site hi a species-specific manner.