Functional Characterization of Three GnRH Isoforms in Small Yellow Croaker Larimichthys polyactis Maintained in Captivity: Special Emphasis on Reproductive Dysfunction

SIMPLE SUMMARY: Small yellow croaker is a popular marine fish in southeast Asian countries. Due to a decline in production in the wild, marine aquaculture has been initiated in Korea. Seed production is performed using captive-reared broodstock, which is known to undergo reproductive dysfunction. Reproductive dysfunction is closely linked to endocrinological dysfunction in the brain-pituitary-gonad (BPG) axis. Gonadotropin-releasing hormone (GnRH) acts as a central regulator in the BPG-axis. To determine the possible involvement of GnRHs in reproductive dysfunction of small yellow croaker reared in captivity, three GnRH isoforms were cloned and functional characterization has been performed. The expression of GnRH1 in the brain was significantly higher at the ripen stage in both sexes during gonadal development. Gonadotropin subunits (GPα, FSHβ, LHβ) were simultaneously increased in the pituitary at the ripen stage in both sexes. Interestingly, females showed significantly lower expression of GnRH1 and GtHs than males. Both in vivo and in vitro administration of GnRH1 showed that GtHs were increased significantly in the pituitary at high concentration. However, sex-steroids (E2 and MT) significantly inhibited the GnRH1 expression in the brain in a dose-dependent manner. Altogether, GnRH1 plays a key role in gonadal maturation, and a low level of GnRH1 and GtHs might be responsible for reproductive dysfunction in the female small yellow croaker. ABSTRACT: Fish reproduction is regulated by the brain–pituitary–gonad (BPG) axis where the gonadotropin-releasing hormone (GnRH) plays a central role. Seed production of small yellow croaker (Larimichthys polyactis) is performed using captive-reared broodstock known to undergo reproductive dysfunction, which is connected to endocrinological dysfunction. To determine the endocrinological mechanism of GnRHs in the BPG axis of small yellow croaker, full-length sequences of three GnRH isoforms encoding sbGnRH (GnRH1), cGnRH-II (GnRH2), and sGnRH (GnRH3) were cloned and characterized from brain tissue. qRT-PCR, in vivo, and in vitro experiments were performed for functional characterization. The mRNA expression of GnRH1 in the brain and gonadotropin subunits (GPα, FSHβ, and LHβ) in the pituitary were significantly higher at the ripen stage during gonadal development and GnRH1 at spawning stage during spawning events. Expression of both GnRH1 and GtH subunits was significantly lower in females than males. GtH subunits were induced at higher concentrations of GnRH1 in vivo and in vitro. Sex-steroids significantly inhibited the GnRH1 expression in vitro in a dose-dependent manner. Taken together, results indicated that GnRH1 plays a key role in gonadal maturation and sex-steroids induced negative feedback in the regulation of GnRH. A lower level of GnRH1 and GtHs might be responsible for reproductive dysfunction in a female small yellow croaker.