Immunohistochemical changes in production of pituitary hormones during artificial maturation of female Japanese eel <Emphasis Type="Italic">Anguilla japonica</Emphasis>

ABSTRACT:   Specific antibodies against follicle-stimulating hormone β subunit (FSHβ), prolactin (PRL), and somatolactin (SL) of the Japanese eel Anguilla japonica were produced. These antibodies, as well as antibodies against luteinizing hormone β subunit (LHβ) and growth hormone (GH) produced previously, were used to examine changes in the production of pituitary hormones in female eels during maturation induced by salmon pituitary homogenate (SPH) injection. Immunohistochemical observations showed a decrease in FSH production after SPH injection, suggesting that SPH inhibits FSH production. In contrast, LH production increased markedly with maturation. The number of GH producing cells decreased gradually during maturation, possibly because of inhibition by exogenous GH present in the SPH and/or endogenous insulin-like growth factor-I produced by the stimulation of salmon GH. Although changes in the number of PRL producing cells with maturation were not evident, the number of SL producing cells showed a peak at the late vitellogenic stages, and thereafter decreased to the migratory nucleus stage. These results suggest that GH and SL are involved in sexual maturation in SPH injected eels, as in other fishes.     

Synthesis and possible function of 11-ketotestosterone during oogenesis in eel (Anguilla spp.)

The control of 11-ketotestosterone (11-KT) biosynthesis and its physiological roles were examined in female Japanese eel (Anguilla japonica) and New Zealand longfinned eel (Anguilla dieffenbachii). 11-KT was detected in serum of female eels of both species. Among various tissues from Japanese eel, the ovary had the greatest capacity to synthesize 11-KT in vitro. In addition, the oocyte diameters of eels treated with 11-KT had increased significantly. Furthermore, these oocytes were found to have an increased number of oil droplets. These findings suggest that 11-KT in female eels may be mostly of ovarian origin and that this androgen appears to play an important role in controlling pre-vitellogenic oocyte growth.     

Variability in growth rates of freshwater eels (Anguilla spp.) in New Zealand

Abstract— Both species of New Zealand freshwater eels (the shortfinned eel Anguilla australis and the longfinned eel A. dieffenbachii ) are widespread and abundant, supporting important commercial and recreational fisheries. This article reviews growth studies from 35 widely distributed sites throughout New Zealand and discusses factors that influence growth rates. Length at given age is characterized by high intra- and inter-population variability; growth rates for eels >30 cm are typically slow (2–3 cm per year) and linear, with females generally growing faster than males. Water temperature affects the length of the growing season, although growth rates are not correlated with latitude. Other factors suggested as affecting growth rates are eel density, quantity and quality of food, and interactions between both eel species. Al though growth of some New Zealand eel populations is the slowest recorded for any species of Anguilla , growth in culture can be rapid, similar to that of other temperate eel species.     

Spatiotemporal variation in the distribution of eel (Anguilla spp.) populations in three New Zealand lowland streams

Abstract – Populations of shortfinned eel ( Anguilla australis ) and longfinned eel ( A. dieffenbachii ) were sampled at randomly chosen sites along the length of three New Zealand streams during summer for three consecutive years to determine their variability in size distribution between streams, reaches (within streams) and years. The longfinned eel was numerically dominant in two of the streams, whereas the shortfinned eel was more abundant in the third stream. For comparisons, the data were stratified into 100-mm size classes for eels ≤600 mm long, whereas eels >600 mm were pooled into a single class. Log-linear analysis indicated that the size distribution of both species differed significantly between streams. Moreover, the size distribution of shortfinned, but not longfinned eels, differed significantly between reaches. Neither species showed significant change in size distribution between years. In contrast to the shortfinned eel, the density of longfinned eel ≤100 mm long was consistently low for all three years, which probably indicates poor recruitment.     

In vivo gonadotropic effects of recombinant Japanese eel follicle-stimulating hormone

Recombinant Japanese eel follicle-stimulating hormone (rjeFSH) produced by methylotrophic yeast was subjected to in vivo bioassay to assess its gonadotropic activities and availability to artificially induce maturation of this species. Intramuscular injections of rjeFSH into male eels at doses of 0.1 and 1.0 U/g body weight were repeated 3 times during 12 days. The rjeFSH significantly increased plasma 11-ketotestosterone levels, and induced both testicular growth and spermatogenesis in a dose-dependent manner. These results demonstrate that rjeFSH is effective in promoting male eel gamatogenesis in vivo. Instead of scarce native eel FSH, abundant recombinant eel FSH is now ready for future application to maturation induction of Japanese eel.     

Lipid content and fatty acid composition of muscle,liver, ovary and eggs of captive-reared and wild silver Japanese eel <Emphasis Type="Italic">Anguilla japonica</Emphasis> during artificial maturation

ABSTRACT:   Changes in lipid content and fatty acid composition of muscle, liver and ovary of captive-reared and wild silver Japanese eel Anguilla japonica were examined during artificial maturation induced by salmon pituitary homogenate (SPH) injections. Although the relative levels of n-3 and n-6 highly unsaturated fatty acids (HUFA) in liver and ovary were higher than in muscle in both captive and wild silver eels before SPH injection, these tended to decrease with maturation. The relative levels of n-6 HUFA in muscle, liver, ovary and eggs of wild silver eels were remarkably higher than those in captive eels. Therefore, we attempted to alter the ratio of n-6 HUFA in eggs by feeding eels a diet supplemented with linoleic acid-rich plant oil. Although the percentage of n-6 polyunsaturated fatty acids to total fatty acids in eggs of eels fed the supplemented diet was similar to wild silver eels, the percentage of n-6 HUFA remained remarkably lower than in wild silver eels. Hence, it appears that the supplemented diet affected the fatty acid composition of eggs, but did not result in much conversion of linoleic acid to its higher homologs in eels.     

鳗鲡繁殖生物学研究进展

鳗鲡是优质养殖鱼类,但鳗鲡苗种完全依靠天然捕捞。近年来鳗鲡苗种的短缺限制了鳗鲡养殖业的进一步发展,开展鳗鲡人工繁殖技术研究势在必行。文章从鳗鲡的催熟与催产、产后鳗鲡的某些繁殖生物学以及鳗鲡胚胎和仔鱼发育等方面简要回顾了鳗鲡繁殖生物学的研究概况,并着重介绍了日本的最新研究进展,日本已将人工苗培育至20多厘米的成鱼,实现了实验室内鳗鲡由卵到成鱼的全人工养殖。文章最后指出人工育苗中存在的问题,并提出今后应从内因和外因两方面研究,才能真正实现鳗鲡苗种规模化生产。     

Assessing sexual maturity of feminized Japanese eel Anguilla japonica by measuring eye size

For efficient production of Japanese eel Anguilla japonica eggs, knowledge of the status of the sexual maturity of potential broodstock females is important because this status directly influences the time required to produce mature eggs by successive hormone doses. Here we apply an eye index (relative eye size) to evaluate the gonadal status of feminized A. japonica, which were induced by administration of estradiol-17β. Examination of gonad somatic and eye indexes of 267 feminized eels, cultivated for 12–56 months, revealed a significant correlation between these two indexes. Artificial maturation of 35 feminized eels showed that the number of hormone injections administered before ovulation was significantly lower as the eye index score increased, indicating availability of this noninvasive method of predicting sexual maturity of feminized eel. There was no correlation between eye index and fertilization rate, hatching rate, or larval survival rate, suggesting that sexual maturity before hormonal treatment does not affect egg quality.     

日本鳗鲡产卵后生存及其繁育

为证明日本鳗鲡(Anguilla japonica)生活史最后一步一产后鳗的命运,本研究模拟产后的日本鳗鲡继续在海水中养殖,观察其存活率及繁育情况.结果表明,产后鳗在海水中停食约18 d后,体能得到恢复,部分亲鱼开始出现摄食,1个月左右全部恢复摄食,经244 d养殖,雌、雄鳗体质量增加,存活率达94.6%.随后,给产后鳗注射外源性促性腺激素(鲤鱼脑垂体匀浆CPE和人绒毛膜促性腺激素HCG)后激发其退化的性腺(卵巢和精巢)重新发育(与当年银鳗作对照).通过性腺组织切片观察产后鳗和对照鳗性腺发育成熟的全过程及其差异,发现产后鳗起初性腺发育比当年银鳗差,但经多次注射激素后,产后鳗性腺成熟与当年银鳗同步,证明产后鳗生殖细胞对激素的敏感件高.应用17α,20β-双羟孕酮和促性腺激素释放激素类似物(GnRH-A3)使催熟的产后鳗和对照鳗均产卵和排精,并孵化出仔鱼,从而有力地证明,鳗鲡产后虽体质弱,但待体能恢复后能够继续生存和繁殖.本研究旨在探讨利用产后鳗作为今后鳗鲡人工繁殖亲鳗的可行性.     

Estimation of gonadal status of Japanese female eel <Emphasis Type="Italic">Anguilla japonica</Emphasis> by a single injection of salmon pituitary extract

ABSTRACT:   Cultivated eels often either lack a part of the ovary (abnormal) or do not have any gonadal tissue (completely sterile). These abnormalities may cause a failure of induction of sexual maturation of female eels. In order to effectively induce sexual maturation and obtain good quality eggs, female eels having normal ovaries should be selected before induction of maturation. The present study examined effects of injection of salmon pituitary extract (SPE) on serum estradiol-17β levels to estimate gonadal status of sexually immature Japanese eels. At the initial stage (just prior to SPE injection), serum estradiol-17β levels were low and no significant difference was observed among eels possessing the different gonad types (normal, abnormal, and sterile), suggesting that gonadal status cannot be estimated by the measurement of serum estradiol-17β levels. A single injection of SPE significantly increased serum estradiol-17β levels in eels possessing normal and abnormal ovaries, but not in sterile eels. Serum estradiol-17β levels significantly increased from 24 h and were high on day 3. These data indicate that eels possessing ovaries can be distinguished from sterile eels by measurement of serum estradiol-17β between 1 and 3 days after a single injection of SPE.     

Effects of temperature on vitellogenesis in Japanese eel Anguilla japonica

ABSTRACT:   Cultured immature female Japanese eels acclimated to sea water at either 10 or 20°C were treated weekly with salmon gonadotropin fraction (sGTH) in order to investigate the effects of water temperature on artificial induction of ovarian maturation. In eels maintained at 20°C, ovulation was induced in 11 of 18 fish during the experimental period of 13 weeks, whereas at 10°C all fish showed a low gonadosomatic index (GSI) at the end of the experiment. Plasma vitellogenin levels were higher in eels kept at 20°C than in eels at 10°C throughout the experiment. However, no significant differences were observed in the plasma testosterone and estradiol-17β levels between groups. Eels pretreated with sGTHs at 10°C for 13 weeks were separated to two different temperature (10 and 20°C) groups, and received the same weekly sGTH injections. In eels transferred to 20°C, plasma vitellogenin levels, GSI and oocyte diameter were increased, but these values were maintained at low levels in eels that remained at 10°C. These results clearly indicate that water temperature is an important factor regulating vitellogenesis in the Japanese eel.     

Developmental changes in pituitary–thyroid axis, and formation of gonads in leptocephali and glass eels of Anguilla spp.

SUMMARY: Pituitary, thyroid gland and gonads in leptocephali of Japanese eel Anguilla japonica (19.8–32.6 mm in total length), A. obscura (45.0 mm), and A. bicolor pacifica (49.5 mm) and those in glass eels of the Japanese eel were histologically and immunohistochemically examined in order to observe the developmental changes of these endocrine organs in the Anguillidae. The pituitary, consisting of adenohypophysis and neurohypophysis in Japanese eel leptocephali over 22.5 mm, did not contain thyroid stimulating hormone (TSH) immunoreactive cells. Such cells were, however, detectable in the more developed pituitaries of leptocephali of A. obscura and A. bicolor pacifica and in those of glass eels. Conversely, thyroxine (T₄)-immunoreactive thyroid follicles could be detected in all specimens, both leptocephalic and glass eel. Only in glass eels, gonads were found in the body cavity, and these gonads harbored one or two primordial germ cells (PGC) per cross-section. Our results indicate that thyroid hormones (TH) production started prior to TSH production, and that TSH and TH are both secreted during the metamorphosis from leptocephalus to glass eel. Therefore, it is plausible that the TSH–TH axis is involved in the metamorphosis from leptocephalus to glass eel, but not in the early growth from preleptocephalus to leptocephalus.     

Changes in Serum Steroid Hormones and Vitellogenin Levels in Cultured Female European Eels Anguilla anguilla during Artificially Induced Ovarian Development

Changes in serum concentrations of estradiol-17β, testosterone, 17α,20βdihydroxy-4-pregnen-3-one, and vitellogenin were investigated during ovarian development induced by injection of a salmon pituitary extract in cultured European eel Anguilla anguilla . Vitellogenesis was induced with a weekly dose of 50 mg pituitary extract/kg body weight. In eels receiving that dose, gonadsomatic indices ranged from 20–43% after the 10th–11th weekly injection. Body weights were relatively stable during vitellogenesis, but increased dramatically during final maturation. Serum estradiol-17β levels increased slightly during vitellogenesis and peaked at an average of 6.95 ng/mL at final maturation. The profile of serum vitellogenin followed that of estradiol-17β which increased markedly from an average of 0.36 to 20.72 mg/mL. Control levels of vitellogenin were undetectable throughout the study. Average serum levels of testosterone increased to a peak of 17.74 ng/mL in the early stage of vitellogenesis, followed by a sharp drop to initial levels (3.86 ng/mL) in the late stage of vitellogenesis, and then increased again to an average of 8.84 ng/mL at the final maturation stage. Serum 17α,20β-dihydroxy-4-pregnen-3-one was not detected (<50 pg/mL) throughout the experiment. Profiles of serum estradiol-17β, testosterone, and vitellogenin observed during ovarian development appear different from those found in salmonids and other teleosts. This study indicates, however, that cultured European eels are a useful model for study of gonadal maturation in the eel.     

Serum steroid profiles in artificially maturing female Japanese eel,Anguilla japonica

To investigate whether steroid profiles in salmon pituitary homogenate (SPH)-induced artificially maturing Japanese eel, Anguilla japonica, resemble those in other, naturally maturing fishes, the daily changes in 11 steroids were analyzed for a 70-day period (average time needed to reach the maturational phase). Concentrations of most steroids were low and changed on a weekly basis, with maximum values 2–5 days after an SPH injection. Thus, pregnenolone, 17α-hydroxypregnenolone, dehydroepiandrosterone, progesterone, 17α-hydroxyprogesterone, 17α,20 β-dihydroxy-4-pregnen-3-one, androstenedione and estrone levels were barely or not detectable in serum throughout the experimental period, which is largely in keeping with what is known about oogenesis-related steroids in other fishes. In contrast, serum testosterone (T) levels were high, but fluctuated considerably with each SPH injection (about 0.3–8.3 ng/ml). The serum estradiol-17β (E₂) levels increased after SPH injections and gradually rose throughout the experiment, peaking at the end of the experimental period (about 0.2–7.8 ng/ml). Serum levels of 11-ketotestosterone (11-KT) before SPH treatment were higher (approximately 2 ng/ml) than those of the other steroid hormones (less than 0.5 ng/ml). 11-KT levels increased gradually over the experimental period, and, like E₂, levels peaked towards the end of the experimental period (about 15 ng/ml). The observed patterns for T, E₂ and 11-KT are unlike those in other fishes. Furthermore, the consistent elevations in levels of 11-KT, both before and after SPH treatment, are suggestive of an important role for this steroid in controlling oocyte growth.     

Insulin-like growth factor I of Japanese eel, Anguilla japonica: cDNA cloning, tissue distribution, and expression after treatment with growth hormone and seawater acclimation

In an attempt to understand growth regulation in the Japanese eel, Anguilla japonica, we cloned insulin-like growth factor-I (IGF-I) cDNAs and examined their mRNA expression in several tissues. Two eel IGF-I (eIGF-I) cDNAs encoding preprohormones, eIGF-I-Ea1and eIGF-I-Ea2, were cloned from the liver by polymerase chain reaction (PCR). The preproIGF-Is were identical in signal peptide and mature IGF-I, but different in the E domain—eIGF-I-Ea2 mRNA was 36 bp longer than eIGF-I-Ea1 mRNA. Eel IGF-I was 83–94% identical with that of teleosts, 71% identical with that of dogfish, 87% identical with that of bullfrog and chicken, and 83% identical with that of humans. In both males and females the highest eIGF-I-Ea1 mRNA levels were observed in the liver, with detectable levels also found in the gills, heart, stomach, spleen, kidney, intestine, swim-bladder, muscle, and gonads. eIGF-I-Ea1 mRNA levels in the liver were higher in females than in males whereas in the intestine they were lower than in males. eIGF-I-Ea2 mRNA was detected in all the tissues examined and at similar levels in males and females. In this experiment higher eIGF-I-Ea1 mRNA levels were observed in the liver of larger glass eels than in those of smaller fish. eIGF-I-Ea2 mRNA levels were also higher in larger eels, although they were lower than IGF-I-Ea1 mRNA levels. Both eIGF-I mRNA levels in liver were positively correlated with the body size of the␣glass eels. Intraperitoneal injection of recombinant eel GH (reGH), 0.25 μg g⁻¹ body weight, into glass eels resulted in a significant increase in both eIGF-I mRNAs in the liver 1 day after injection compared with control fish, but no elevation was observed 2 days after injection. Incubation of liver slices with reGH at concentrations of 10, 100, and 1,000 ng mL⁻¹ for 24 h resulted in a significant concentration-dependent increase in the levels of both eIGF-I mRNAs. Higher levels of eIGF-I-Ea1 and Ea2 mRNA were observed in the gills ofseawater-reared eels than in those of freshwater-reared fish, but no differenceswere observed in the whole kidney. These results suggest that IGF-I is involved in the regulation of somatic growth and also in adaptation of the Japanese eel to seawater.     

Induction of spermatogenesis in Japanese eel by recombinant goldfish gonadotropins

We previously demonstrated the biological activities of single-chain recombinant gonadotropins (scGTHs) of goldfish Carrassius auratus follicle-stimulating hormone (scFSH) and luteinizing hormone (scLH), produced by a baculovirus–silkworm larvae system, by using in vivo bioassays with some fishes including Japanese eel Anguilla japonica. Among the bioassays, we succeeded in induction of spermatogenesis of sexually immature male Japanese eels by both scFSH and scLH, especially resulting in the occurrence of spermatozoa in scLH-administered males. However, those recombinant hormones did not induce enlargement of testes. In order to further confirm the potency of recombinant GTHs for use in aquaculture species, we administered scFSH and scLH to males of Japanese eel at higher dosage and frequency (eight times with 2–5 days interval) than those of the previous study (five or six times with 7 days intervals), including combination of scFSH and scLH administration (scFSH–scLH). Gonadosomatic indices (GSI) of scLH- and scFSH–scLH-administered males were larger than those of initial control males and of control males that were injected with saline. Enlargement of testes was also confirmed by measurement of testicular lobe size in scFSH-, scLH-, and scFSH–scLH-administered males. By histological observation, occurrence of spermatozoa was confirmed in scLH- and scFSH–scLH-administered eels. Although milt production was not induced, higher dosage and frequency of scGTH administration was effective in promoting testicular development of immature eels. Thus, single-chain fish GTHs produced by the baculovirus–silkworm larvae system could be a useful tool for promotion of gonadal maturation in aquaculture fishes.     

Longevity of longfinned eels Anguilla dieffenbachii in a New Zealand high country lake

A sample of 146 longfinned eels (Anguilla dieffenbachii), ranging in length from 231–1203 mm, was collected from Lake Rotoiti, Nelson Lakes National Park, New Zealand. Otoliths (n= 114) of eels were difficult to read, mainly due to the considerable age (maximum age 106 years). Growth rate was slow and linear, averaging 9 mm. year^?1. These are the slowest growth rate and maximum age from any study of Anguilla spp. to date. Lakes within National Parks provide the largest commercially unfished waters within New Zealand and hence provide reserve breeding stocks of longfinned eels; however, the average generation time of 93 years for female eels from Lake Rotoiti implies that stock management should be conservative.