Effects of Salmon Gonadotropin-Releasing Hormone on Follicle Stimulating Hormone Secretion and Subunit Gene Expression in Coho Salmon (Oncorhynchus kisutch)

Abstract

Previous work has indicated that, during the process of gametogenesis in salmon, follicle-stimulating hormone (FSH) and luteinizing hormone (LH) are differentially synthesized and released. Although substantial information is available on the regulation of LH in many fish species, relatively little is known about the regulation of FSH biosynthesis and secretion or the regulation of two types of α subunit in salmon. In this study, the effects of salmon gonadotropin-releasing hormone (sGnRH) on in vitro secretion of FSH, and α1, α2, LH β, and FSH β subunit gene expression were investigated in coho salmon (Oncorhynchus kisutch) using primary pituitary cell cultures. To quantify FSH β, LH β, α1, and α2 subunit transcript levels, a multiplex RNase protection assay (RPA) was developed. Probes for the β subunits of coho salmon FSH and LH were available from previous studies. To generate probes for the α subunit RPAs, α1 and α2 subunit cDNAs were cloned using reverse transcriptase PCR. Release of FSH and LH into cell culture medium was quantified by radioimmunoassays. The effects of sGnRH on gonadotropin release and gene expression were tested at two points during the spring (April and May) prior to spawning in the autumn; a period when plasma and pituitary FSH levels are increasing and females are in early stages of secondary oocyte growth. In both experiments, sGnRH increased steady-state mRNA levels of FSH β, α1, and α2, whereas LH β mRNA levels were not detectable. Secretion of FSH was stimulated by sGnRH in a concentration-dependent manner. Medium LH was not detectable in the first experiment (April) and was measurable only after sGnRH treatment in the second experiment (May). Control levels of medium FSH and transcripts for FSH β and α1 subunits increased approximately fourfold between April and May, whereas α2 transcript levels remained relatively constant, suggesting that the seasonal increase in FSH release may involve increased production of α1. Therefore, sGnRH has direct stimulatory effects on both secretion of FSH and FSH subunit biosynthesis, most likely due to increased transcription. However, alterations in rates of transcript degradation cannot be ruled out.