Abstract
In this thesis, we set out to clone Pgrs from three fish species belonging to different orders, namely the zebrafish (Cypriniformes), the Atlantic salmon (Salmoniformes) and the Atlantic cod (Gadiformes). Cloning of the full-length cDNAs was the prerequisite for expressing these receptors in a host cell line, which allowed functional
... read more
characterization of the receptors. These studies indicated that 17alpha,20beta-dihydroxy-4-pregnen-3-one (DHP), a typical progestin in fish, is the most effective native ligand for the teleost Pgrs cloned in this thesis. Moreover, the cloned pgrs are predominantly expressed in testis. Taken together, these first results suggest that Pgrs have important function on the testicular level. Furthermore, the cellular localization of the pgr transcripts was investigated in testis using in situ hybridization. The results indicated pgr mRNA specific signals were observed in Sertoli cells from all three fish species. In zebrafish, pgr mRNA was also detected in Leydig cells. Using zebrafish, our results indicated that the role for Pgr expressed in Leydig cells is to modulate intratesticular levels of other steroid hormones (e.g. cortisol or 11-ketotestosterone) by increasing 11betaHsd activity. Using salmon, quantification of pgr mRNA in testis in relation to plasma DHP levels and germ cell development suggested that the salmon Pgr may be involved in the regulation of early spermatogenesis, in particular the differentiation of late type B spermatogonia and the entry into meiosis. In line with studies in Atlantic salmon, the quantification of pgr mRNA in cod testis during the onset of spermatogenesis indicated that the cod Pgr may be involved in the regulation of the late mitotic and meiotic phases of spermatogenesis. Moreover, the receptor was strongly up-regulated in cod testis tissue approaching the spawning condition, suggesting roles for the Pgr during the final stages of the reproductive cycle, such as has been described previously for other species (e.g. sperm hydration and spermatozoa mobility). Subsequently, using an in vivo experimental model (oestrogen-mediated down-regulation of androgen production to interrupt spermatogenesis), and a recently developed, ex vivo primary zebrafish testis tissue culture system, we could show that DHP treatment induced the proliferation of early spermatogonia as well as their differentiation into late spermatogonia and spermatocytes. Further, transcripts of Sertoli cell-derived growth factor genes were up-regulated after DHP treatment in vivo, but only insulin-like growth factor 1b receptor (igf1rb) mRNA levels showed a significant increase under DHP treatment in vivo and ex vivo. The present thesis expanded the knowledge on the roles of progestins and its nuclear receptor in fish spermatogenesis. Moreover, our results were taken to develop a model in which we suggest that the mitotic phase of spermatogenesis, under the overall support by gonadotropins, is driven by a potential steroid-Igf signalling axis, which may operate in concert with other paracrine factors in the fish testis to regulate spermatogenesis.
show less