Effects of Secretoneurin and Gonadotropin-Releasing Hormone Agonist on the Spawning of Captive Greater Amberjack (Seriola dumerili)

Abstract

The greater amberjack (Seriola dumerili), a pelagic marine species with a global distribution, has considerable worldwide potential as an aquaculture species. However, difficulties have been encountered in inducing spontaneous spawning in cultured fish stocks. In this study, we analysed the key regulatory factors, secretoneurin (SN) and gonadotropin-releasing hormone (GnRH), in greater amberjack. Active peptides of SN and GnRH, SdSNa, and SdGnRH, respectively, were obtained by comparative analysis of homologous proteins from different species. Amino acid substitutions of the SdGnRH decapeptide at position 6 with a dextrorotatory (D) amino acid and at position 10 with an ethylamide group yielded a super-active agonist (SdGnRHa). The injection of SdSNa and SdGnRHa elevated luteinizing hormone, thyroid-stimulating hormone, and oxytocin levels in the sera of sexually mature fish, whereas it reduced the level of follicle-stimulating hormone. Furthermore, in response to the SdSNa and SdGnRHa injections, we detected an increase in the expression of genes associated with oocyte development and spermatogenesis. We established that the greater amberjack cultured along the southern coast of China reached sexual maturity at three years of age, and its reproductive season extended from February to April. Spawning of the cultured greater amberjack was successfully induced with a single injection of SdGnRHa/SdSN/DOM/HCG. Our findings indicate that similar to GnRHa, SNa is a potential stimulator of reproduction that can be used to artificially induce spawning in marine fish.

Keywords: agonist; cultured greater amberjack; gonadotropin-releasing hormone; secretoneurin; spawn.

Figure 1

Comparative analysis of the secretoneurin active peptide (SNa) and gonadotropin-releasing hormone (GnRH) protein sequences of multiple species. (A) SNa consists of a conserved 34 amino acids active peptide. (B) GnRH consists of a conserved decapeptide. Accession IDs of secretogranin-2 proteins are listed in Supplemental Table S1 and accession IDs of gonadotropin-releasing hormone proteins are listed in Supplemental Table S2. The * indicates the most conserved amino acids.

Figure 2

Serum hormone levels of S. dumerili after injection of SdSNa/DOM, SdGnRHa/DOM, and normal saline (n = 6). Serum levels of luteinizing hormone (A) and follicle-stimulating hormone (B) were increased and reduced, respectively, following injection with SdSNa/DOM or SdGnRHa/DOM. Serum levels of both thyroid-stimulating hormone (C) and oxytocin (D) were elevated in response to SdSNa/DOM and SdGnRHa/DOM treatment. The a, b, c indicates the indicate statistically significant differences, p < 0.05.

Figure 3

Transcriptome analysis of differentially expressed genes in the gonads of SdSNa/SdGnRHa/DOM injected S. dumerili (n = 3). (A) An MA plot showing the significantly downregulated (in green) and upregulated (in red) genes in the ovaries of injected females. (B) Genes implicated in oocyte development were upregulated after injection. (C) An MA plot showing significantly altered genes in the testes of injected males. (D) Genes implicated in spermatocyte development that were upregulated in the SdSNa/SdGnRHa/DOM injected S. dumerili. The ＊ indicates statistically significant differences, p < 0.05.

Figure 4

Micrographs of gonadal tissue sections sampled at three different growth stages from male and female S. dumerili, and the body weight, body length, and gonadosomatic index of sexually mature individuals were measured during the breeding season. (A) Oogonia and primary growth oocytes were detected in the ovaries of 2-year-old fish. (B) Spermatogonia and a few meiotic-stage spermatocytes were detected in the testes of 2-year-old males. (C) Perinucleolar oocytes as the most advanced stage in the ovaries of 3-year-old females during the non-reproductive season. (D) Although seminiferous ducts were observed at this time, no sperm was detected. (E) Vitellogenic oocytes in the ovaries of 3-year-old females in February. (F) On entering the reproductive season, the seminiferous ducts of male fish were filled with mature sperm. No significant differences were detected in the body weight (G) or body length (H) of male and female fish. The gonadosomatic index (I) of females was notably higher than that of males. Scale bar: 200 μm (A,C–F), 50 μm (B). The ＊ indicates statistically significant differences, p < 0.05.

Figure 5

Hormonal treatments were used to induce the spawning of S. dumerili. (A) Body size of 3-year-old sexually mature greater amberjack. The ovaries (B) and testes (C) were sampled to assess the developmental and mature stages. (D) The fertilized eggs were collected and weighed. (E) The development of fertilized eggs into embryos.