A. R. Midgley

Development of Ovarian Responsiveness: Follicle Maturation and Luteinization

ABSTRACT Although granulosa cells may be isolated by simple expression from Graafian follicles, the viability and functional activities of the resulting cells have been markedly enhanced by incubating ovaries sequentially in EGTA and hypertonic sucrose prior to expression. It is believed that this treatment gently dissociates interconnecting gap junctions and thereby reduces the extent of cellular disruption that usually results from expression. By incubating the cells with [35S]-methionine at various times after injecting human chorionic gonadotropin (hCG), and then subjecting cellular homogenates to two dimensional electrophoresis, it was found that hCG-induced luteinization is accompanied by changes in synthesis of specific proteins, and that these changes occur prior to the onset of morphologic signs of luteinization. To obtain information on the fate of hCG bound to the granulosa cells, we labeled the non-identical subunits with different radioisotopes of iodine, and demonstrated that the dual labeled hormone was biologically active. The particulate fraction of granulosa cells showed a preferential retention of the beta subunit-associated radioactivity that was not seen in any other target or non-target control tissues. We conclude that a portion of the beta subunit of hCG is selectively retained by luteinizing granulosa cells. Whether or not this retention is causally related to the onset of luteinization remains to be determined.