SUMMARY: In this study, ovarian development and vitellogenin levels were determined during the reproductive and non‐reproductive molt cycles in the giant freshwater prawn Macrobrachium rosenbergii in order to examine the relationship between vitellogenesis and molting. During the reproductive molt cycle, the ovary developed synchronously with the advancement of molting stage. At molting stages B–C0, the ovaries contained oogonia and oocytes at the previtellogenic and endogenous vitellogenic stages, and hemolymph vitellogenin levels were relatively low (2.4–3.0 mg/mL). At stage C1, yolk accumulation commenced in oocytes at the endogenous vitellogenic stage, after which oocytes entered the exogenous vitellogenic stage. Vitellogenin levels increased (4.9 mg/mL) at this stage. During premolt stages D0–D3, vitellogenin titers were high (4.7–7.3 mg/mL), and gonadosomatic index (GSI) increased rapidly due to the yolk accumulation in the oocytes. Following ecdysis, yolk accumulation was completed at stage A0, and oviposition occurred. Entering stage A, vitellogenin levels became low (0.6–1.3 mg/mL). In contrast, throughout the non‐reproductive molt cycle, GSI remained low, and oocytes did not develop beyond the previtellogenic or endogenous vitellogenic stage. Some atretic oocytes at the exogenous vitellogenic stage were observed in a few females. During the non‐reproductive molt cycle, vitellogenin levels did not show distinct changes in relation to the molt cycle.

To investigate the nutritional regulation of lipid metabolism in fish, molecular characterization of lipases was conducted in red sea bream Pagrus major, and the effects of fasting and refeeding on their gene expression was examined. Together with data from a previous study, a total of four lipase genes were identified and characterized as lipoprotein lipase (LPL), hepatic lipase (HL) and pancreatic lipase (PL). These four lipase genes, termed LPL1, LPL2, HL and PL, share a high degree of similarity. LPL1 and LPL2 genes were expressed in various tissues including adipose tissue, gill, heart and hepatopancreas. HL gene was exclusively expressed in hepatopancreas. PL gene expression was detected in hepatopancreas and adipose tissue. Red sea bream LPL1 and LPL2 gene expression levels in hepatopancreas were increased during 48 h of fasting and decreased after refeeding, whereas no significant change in the expression levels of LPL1 and LPL2 was observed in adipose tissue, indicating that LPL1 and LPL2 gene expression is regulated in a tissue-specific manner in response to the nutritional state of fish. HL and PL gene expression was not affected by fasting and refeeding. The results of this study suggested that LPL, HL and PL gene expression is under different regulatory mechanisms in red sea bream with respect to the tissue-specificities and their nutritional regulation.

Various kinds of hormones including insulin, triiodothyronine (T(3)) and fat-soluble vitamins have been proposed as mediators of adipocyte differentiation in mammals. To investigate the factors which are responsible for fish adipocyte differentiation, we developed a serum-free culture system of stromal-vascular cells of red sea bream adipose tissue and examined the effects of bovine insulin, T(3), and fat-soluble vitamins (all-trans retinoic acid, retinyl acetate and 1,25-dihydroxyvitamin D(3)) on the differentiation-linked expression of the lipoprotein lipase (LPL) gene. As assessed by the increase in LPL gene expression after 3 day cultivation, like in mammalian adipocytes, insulin enhanced the adipocyte differentiation in a concentration-dependent manner. During 2 week cultivation, bovine insulin promoted lipid accumulation in differentiating adipocytes concentration-dependently until the terminal differentiation. These results indicate that the differentiation of fish adipocytes is inducible by insulin alone. T(3) alone had no effect but enhanced the differentiation-linked LPL gene expression in the presence of insulin. Fat-soluble vitamins, unlike in mammalian adipocytes, did not show any significant effects. The method developed in this study should be of interest for the characterization of factors involved in fish adipocyte differentiation.

SUMMARY: Hemolymph ecdysteroid levels were traced during the molt cycle in female giant freshwater prawn, Macrobrachium rosenbergii. Levels were compared between the reproductive molt cycle, accompanied by ovarian development and oviposition following ecdysis, and the non‐reproductive molt cycle, marked by the absence of ovarian development, to examine the involvement of ecdysteroids in ovarian development. Total ecdysteroid levels determined by radioimmunoassay (RIA) showed similar fluctuation patterns between both molt cycles. Levels were low during postmolt stages A–B and gradually decreased to low basal levels at intermolt stages C0–C1. Levels then increased during premolt stages D0–D2, reached a large peak at the early D3 stage, and declined sharply prior to ecdysis. In addition, ecdysteroids in the hemolymph were analyzed using high‐performance liquid chromatography combined with RIA in order to identify individual ecdysteroid species. During both molt cycles, 20‐hydroxyecdysone, ecdysone and high polarity immunoreactive products were detected, and 20‐hydroxyecdysone was the most predominant ecdysteroid. Fluctuations in all ecdysteroid species during each type of molt cycle paralleled those of total ecdysteroid levels, and did not show significant differences between the reproductive and non‐reproductive molt cycles. These results suggest that ecdysteroids are not involved in ovarian development but in the molting process only.

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Shrimp culture is a significant world-wide industry, with current production levels reaching over 3 million tons per year. The expansion of the industry has given rise to the problems of environmental deterioration due to intensive-scale culture, and the outbreak of disease. While many of these issues are now being sufficiently addressed, the establishment of sustainable seed production technology is an area that should be given continued attention. In this regard, it remains difficult to control reproduction under hatchery conditions for a large number of commercially-important species. At present, an understanding of reproductive mechanisms in Crustacea is not complete, although in recent years, a great deal of knowledge has accumulated on vitellogenin structure, processing, and synthetic site in a number of economically-important species. This monograph will cover the current status of research on vitellogenin in decapod crustaceans, especially prawns and shrimp, and discuss mechanisms of vitellogenin synthetic control, both demonstrated and postulated. The monograph will also present current knowledge of crustacean vitellogenin receptors, and cover related facets of reproductive development, such as mechansisms of cortical rod formation and the utilization of vitellin during embryogenesis. Finally, future directions for this research and potential applications to aquaculture will be discussed.

In the present study, the effects of bilateral eyestalk ablation on molt interval, ovarian development, and hemolymph levels of ecdysteroid and vitellogenin were investigated in the giant freshwater prawn, Macrobrachium rosenbergii in order to examine the roles of the eyestalk hormones. In destalked males and females, ecdysteroid levels increased rapidly, and molt intervals were significantly shortened in comparison with control non‐destalked prawns. All destalked females exhibited a reproductive molt cycle accompanied by ovarian development and oviposition even under continuous hemolymph sampling. In contrast, in control females, the percentages of animals undergoing a reproductive molt cycle were 69 and 51% with and without sampling, respectively. The duration of ovarian development was shortened in the destalked females. These results indicate the inhibitory roles of the eyestalk hormones on molting and vitellogenesis. On the other hand, after eyestalk ablation, male prawns molted more than twice, and destalked females showed a correlation of vitellogenin level to molt stage similar to control reproductive females. These results indicate that the molt cycle and the correlation between vitellogenesis and molting are maintained in the absence of eyestalk hormone regulation.