Estrogen receptor-alpha (ERalpha) in the gonads of the rainbow trout (Oncorhynchus mykiss).

摘要

Estradiol-17β (E2) is an important hormone in both male and female reproduction. As many of the biological actions of E2 are mediated through estrogen receptors (ERs), an important step in understanding E2 function is to identify its target sites in cells. Little is known about ER localization or E2 functioning in the gonads of non-mammalian vertebrates. The first objective of this research was to identify ER-alpha (ERα) protein localization in the testis of the oviparous fish, the rainbow trout (Oncorhynchus mykiss), during the annual reproductive cycle. ERα protein was found to be present within the interstitial cells only, throughout the reproductive cycle.; Secondly, the cellular localization pattern of ERα protein was examined in the ovary of the rainbow trout throughout the reproductive cycle. In the ovary, ERα protein was localized in the interstitial cells (i.e. fibroblasts, blood vessels) and oocytes with a ovarian follicular diameter >200 μm. Furthermore, ERα protein was also present in the theca cells associated with ovarian follicular diameters >2500 μm.; Thirdly, in order to investigate a possible direct function of E2 in the immature rainbow trout testis, a floating organ culture was utilized. E2 caused a significant increase in testicular interstitial cell proliferation. This effect appeared to be mediated by an intracellular ER, as the proliferation could be nullified using the ER antagonist ICI 182,780, and no effect was observed using an E2-bovine serum albumin (E2-BSA) conjugate.; Finally, preliminary experiments were conducted to investigate the feasibility of utilizing the floating organ set up to culture whole immature testis and ovary and generate transgenic rainbow trout germ cells. Sexually immature testis and ovary were successfully cultured for 5 days. Furthermore, using the floating organ culture system, testicular fragments were transfected with a green fluorescent protein (GFP) reporter gene plasmid. GFP was successfully detected in both somatic and germ cells. These experiments suggest the potential of using this floating organ culture in combination with a recently developed transplantation protocol, to eventually generate mature transgenic rainbow trout sperm. This technology may lead to the creation of an ER knockout rainbow trout.