Population structure of the Japanese eel, Anguilla japonica

Mitochondrial DNA (mtDNA) sequences that include (a) a part of the cytochrome b gene, (b) two tRNA genes, and (c) a part of the noncoding D-loop region of 31 Anguilla japonica (Japanese eel) and 1 A. marmorata collected from Taiwan, Japan, and mainland China were determined to evaluate the population structure of Japanese eel. Among 30 genotypes identified from the 31 Japanese eel mtDNAs sequenced, there are 58 variable sites, predominantly clustered at the D-loop region. The phylogenetic tree constructed by the unweighted pair-group method with arithmetic mean shows neither significant genealogical branches nor geographic clusters. Furthermore, the sequence-statistics test reveals little, if any, significant genetic differentiation. These results indicate that the 31 Japanese eels might come from a single population. Analysis of sequence variation in mtDNA by using the relationship between the number of segregating sites and the average number of nucleotide differences under the neutral mutation hypothesis reveals that neutral mutation acts as a major factor influencing the evolutionary divergence of the Japanese eel mitochondrial genome sequenced, especially in the noncoding region.      

Recruitment mechanism of the eel, Anguilla japonica, to the Japanese coast

A total of 149 Anguilla japonica elvers collected at 10 locations along the Japanese coast were aged according to otolith daily increments. The interrelationships among age, birth date, body size, pigmentation stage, sampling location and the timing of recruitment were examined in order to determine the recruitment mechanism of elvers to coastal waters.
First catches of eels were earlier in the locations at lower latitudes. Age at recruitment was roughly constant, 218 ± 29 (mean ± s.d.) days old, disregarding the localities, but a weak positive correlation was obtained between age and sampling date or timing of recruitment. Body length at recruitment was also constant, 56.3 ± 2.3 mm s.l., and showed no significant correlation with either locality or recruitment timing. Estimated birth date ranged from April to November, the mean ± s.d. being 22 July 1982 ± 42 days, suggesting a peak season spawning in summer. Birth date was closely related with both the latitude of location and sampling time. Pigmentation developed more at lower latitude. Recruitment mechanism of the Japanese eel was summarized as follows: the earlier-born individual recruits earlier at lower latitude and at younger age, but at a constant body size.     

Antidipsogenic effects of eel bradykinins in the eel Anguilla japonica

A peptide with bradykinin (BK)-like immunoreactivity was isolated from an incubate of heat-denatured eel plasma with porcine pancreatic kallikrein. The purified peptide had the following amino acid sequence: Arg-Arg-Pro-Pro-Gly-Ser-Trp-Pro-Leu-Arg. This decapeptide, named eel [Arg(0)]BK, was identical to two previously identified BK homologs from cod and trout. High conservation of the BK sequence among distant teleost species suggests an important function in this vertebrate group. Bolus intra-arterial injections of eel [Arg(0)]BK, BK, and [Arg(0)]-des-Arg(9)-BK (1-10 nmol/kg) caused significant (P < 0.05) inhibition of drinking in seawater-adapted eels. The potency of the inhibition was ranked in the following order: [Arg(0)]BK > [Arg(0)]-des-Arg(9)-BK = BK. The BK peptides also produced an immediate, transient increase followed by a sustained increase in arterial blood pressure and an initial decrease followed by an increase in heart rate. Strong tachyphylaxis occurred for the cardiovascular effect but not for the antidipsogenic effect. The order of the potency of the cardiovascular actions, [Arg(0)]BK > BK > [Arg(0)]-des-Arg(9)-BK, was different from that of the antidipsogenic action. Slow infusions of eel [Arg(0)]BK in the dose range 1-1,000 pmol x kg(-1) x min(-1) produced concentration-dependent inhibition of drinking without changes in arterial pressure, plasma osmolality, and hematocrit. At the infusion rate of >100 pmol x kg(-1) x min(-1), plasma concentrations of angiotensin II, a potent dipsogenic hormone in eels, increased, suggesting an interaction of the kallikrein-kinin and renin-angiotensin systems. In mammals, BK is dipsogenic and vasodepressor, so that our data demonstrate opposite effects on fluid and cardiovascular regulation of BK in the eel and suggest a new physiological role for the kallikrein-kinin system in teleost fish.     

Temperature effects on the incorporation of strontium in otolith of Japanese eel Anguilla japonica

The effects of temperature on somatic and otolith growth and the incorporation of strontium in otolith of the Japanese eel, were studied in laboratory-reared and field-caught eels. The somatic and otolith growth rates of the eel increased significantly with temperature and were estimated as approximately 0·096 mm t.l, (P<0·01) and 0·36 μm in otolith diameter per degree-day (0·01相似文献   

A novel progestogen receptor subtype in the Japanese eel, Anguilla japonica.

A cDNA encoding a second type of a progestogen receptor (ePR2) was isolated from the same library as we had previously cloned a functional PR (ePR1) in eel testis. The amino acid sequence of the ePR2 shows low homology with ePR1 (34%), but both PRs showed progestogen-dependent transactivation in transfection experiment. Tissue distribution of ePR2 mRNA was clearly different from that of ePR1. Protein interaction between two PRs was demonstrated in vitro by a glutathione S-transferase pull-down assay. These results indicate that ePR2 is also a functional PR. This is the first isolation of two different functional PR molecules from a vertebrate.     

Effect of lunar periodicity on the locomotor activity of silver-stage Japanese eel,Anguilla japonica

Lunar periodicity has been thought to influence the onset of the spawning migration of anguillid eels. In this study, we measured daily locomotor activity of 8 silver-phase Japanese eels Anguilla japonica in outdoor tanks to examine the effect of lunar periodicity on their activity and the following seaward migration. The activity of silver eels was highest around the new moon during the early part of the experiment, which is the ordinary season of seaward migration in Japan. The observed patterns of activity may reflect the importance of the lunar cycle for the onset of the spawning migration in anguillid eels.     

Temperature influence on the spawning performance of artificially-matured Japanese eel, Anguilla japonica, in captivity

We studied the influence of temperature on the spawning performance of artificially matured Japanese eels, Anguilla japonica, in captivity. We used routine hormone injections to bring females and males to maturity in separate aquaria. We recorded the behavior of three pairs of such hormone-treated matured eels in an aquarium (2 replicates) at four temperatures: 14, 18, 22, and 27°C, respectively. They became active and frequently left the bottom swimming in the water column, and spawning events occurred. Females released eggs in the water column around the activity peaks. Males preceded females in reaching activity peaks (presumably the timing of sperm ejection and egg release), possibly resulting in the low fertilization we observed in this experiment. Males and females returned back to the aquarium bottoms and became quiet after spawning. On several occasions, male-female or female-female pairs were observed to ‘cruise together’ in the water column for several to tens of seconds prior to egg releasing, but no courtship behavior indicative of spawning such as pairing and chasing was observed in the eels in our study. Our results suggest that 18–22°C might be the thermal preference for spawning for Japanese eels, which approximates the temperature range of the 500 m deep water layer around the Mariana Islands seamount area, the presumed spawning site for the Japanese eel.

Daily growth increments in the larval otolith of the Japanese eel,Anguilla japonica

Early formation of otolith was studied on artificially hatched larvae of the Japanese eel,Anguilla japonica. Newly hatched larvae had a pair of sagittae which were flat and subelliptical with 8.3 μm in mean diameter. The diameter of the sagitta increased linearly with age. No growth increments were observed in the sagitta at hatching, while larvae which were 2, 4 and 6 days old had on average 2.1, 3.6 and 6.0 increments, respectively. The number of the increments (Y) and the age in days after hatching (X) showed a close linear relationship (Y=0.96X+ 0.06, r = 0.913, n = 40), suggesting daily deposition of sagittal increments. In 95 % of the field-caught elvers of this species, a distinct dark ring (check) with the diameter of 6–12 μm was found around the nucleus of the sagitta. This seems to be a “hatch check” deposited at hatching, since its diameter roughly agreed with that of the sagitta in the newly-hatched larvae. Possibly, the number of the increments outside the hatch check represents the age of the fish in days.     

Serum estradiol-17beta and testosterone levels during silvering in wild Japanese eel Anguilla japonica

To understand the changes of serum levels of sex steroids in the wild Japanese eel Anguilla japonica during silvering process, eels collected from the Kaoping River of Taiwan from August 2000 through June 2001 were examined. The maturational stages of female eels before and during silvering were divided into four stages: juvenile, sub-adult, pre-silver and silver stages based on skin coloration and oocyte diameter. Male eels were investigated only in the silver stage. Radioimmunoassays were employed to measure serum levels of estradiol-17β (E₂) and testosterone (T). The mean liver mass of the female eels increased significantly during silvering, but the mean hepatosomatic index remained constant. In contrast, mean ovarian mass and gonadosomatic index increased significantly during silvering. Serum concentrations of E₂ in females increased significantly during silvering (P<0.05), while E₂ was undetectable in silver males. The mean serum T concentrations increased significantly in females (P<0.05) during silvering, with lowest mean values in the juvenile stage and highest mean value in the silver stage. The mean serum T level in the silver males was significantly lower than in silver females (P<0.05). In conclusion, both serum E₂ and T concentrations increased with ovarian development of wild Japanese eels during silvering, while serum E₂ was undetectable in the silver male eels. The findings support the idea that androgen, but not estrogen, plays a major role in silvering process of the eels in both sexes.     

Effects of changes in environmental calcium on prolactin secretion in Japanese eel,Anguilla japonica

Effects of changes in environmental Ca²⁺ on the secretion of prolactin, a possible hypercalcemic hormone, were examined both in vivo and in vitro in the Japanese ecl, Anguilla japonica. Transfer of seawater- or freshwater-adapted fish to fresh water, fresh water containing 10 mmol Ca²⁺ · 1^-1 sea water, Ca²⁺-free sea water, or deionized water was accompanied by significant changes in plasma Ca²⁺ levels after 7 days, except for the fish transferred from fresh water to fresh water and from sea water to sea water. Changes in external Ca²⁺ concentrations did not affect plasma prolactin levels, although plasma prolactin levels as well as pituitary prolactin contents were significantly greater in fish in a hypotonic environment than those in a hypertonic environment, regardless of the external Ca²⁺ concentration. Hypercalcemia, induced by removal of the corpuscles of Stannius, did not alter plasma prolactin levles. Incubation of the pituitary in the medium with different Ca²⁺ concentrations (up to 2.9 mmol·l^-1) did not affect the basal release of prolactin, except at an extremely low Ca²⁺ concentration (less than 0.1 mmol·l^-1) where prolactin release was inhibited. Addition of Ca²⁺ ionophore (A23187) to the medium led to a marked and significant increase in prolactin release, indicating that an increase in intracellular Ca²⁺ stimulates prolactin release. However, the effect was not specific to prolactin cells; a similar increase was seen in growth hormone release. These results indicate that changes in environmental Ca²⁺ concentration may not be the primary factor influencing prolactin secretion in the eel; changes in environmental osmolality or Na⁺ levels seem to be more critical for the regulation of prolactin secretion.Abbreviations CSX stanniectomy - DMSO dimethylsulphoxide - DW deionized water - FW fresh water - GH growth hormone - PRL prolactin - SW sea water     

"Blood-contacting neurons" in the brain of the Japanese eel Anguilla japonica

To discriminate "blood-contacting neurons" within the brain of the eel, Evans blue (EB) was injected intraperitoneally. After five days, six brain areas were externally stained blue with the dye; the saccus dorsalis (SD), the epiphysis (E), the area postrema (AP), the posterior part of the magnocellular preoptic nucleus (PM), the pituitary (Pit), and the saccus vasculosus (SV). Among the EB-positive area, some cells in the PM, the anterior tuberal nucleus (NAT) and the AP were discriminated as the "blood-contacting neurons" histologically, whereas EB-positive neurons were not detected in the SD, the E, the Pit and the SV regions. In the PM, most EB-positive neurons (90 %) were immunoreactive to vasotocin (AVT) antibody, indicating that these neurons are vasotocinergic. The remaining EB-positive neurons (10 %) were not immunoreactive to ANG II and tyrosine hydroxylase (TH) antibodies. Although some neurons in the PM were immunoreactive to ANG II antibody, they were EB-negative. In contrast, almost all EB-positive neurons in the AP showed TH-like immunoreactivity (-lir), indicating that these neurons utilize catecholamine(s) as a neurotransmitter. The EB-positive neurons in the NAT were not immunoreactive to AVT, ANG II and TH antibodies, whereas some neurons without EB-staining showed ANG II-lir. Possible roles of these neurons in regulating drinking behavior in eels are discussed.     

Ecological aspects of the Japanese eel, Anguilla japonica, collected from coastal areas of Japan

The ecological characteristics of 597 yellow and silver-stage Japanese eels, Anguilla japonica, were examined and compared among collection sites located at three different latitudes of Japan (Amakusa Islands, Mikawa Bay, and Sanriku Coast) to provide basic data on this unusual catadromous fish species. Eels were sexed and their total length, body weight, age, and growth rate based on otolith analysis was compared among sexes, stages, and collection sites. The overall sex ratio favored females (94%), but the sex ratio differed among the three locations. The frequency of females was highest in the coastal waters at Sanriku in the north (100%), next highest at Mikawa Bay in central Japan (95%), and lowest in the Amakusa Islands in the south (70%). Silver eel males ranged from 41.2-66.3 cm in length and 4-10 years in age, and silver eel females from 44.3-97.2 cm in length and 5-17 years in age. Female eels generally grew faster (8.7+/-2.2 cm/year) than males (6.4+/-2.6 cm/year), and the growth rate slowed in the older eels. The growth rate of A. japonica at all three sites was much faster than that of other temperate anguillid species (< 4 cm/year), and their age at maturation was younger than that of other temperate species (approximately 7 to > 50 years), suggesting this species has important ecological differences from other similar species.     

Relative abundance, sex ratio and population structure of the Japanese eel Anguilla japonica in the Tanshui River system of northern Taiwan

From the Gong-Shy-Tyan (GST) Stream and the Tanshui River, Taiwan, eels ranged from 5.53 cm t.l . (elvers) to 72.5 cm t.l . Anguilla japonica accounted for 93-99%, A. marmorata 1-7% and A. bicolor pacifica less than 1% of all eels caught. Mean eel lengths increased from 10 cm t.l . at the downstream sites to 50 cm t.l . at the upstream sites. Females made up 92.8% of the sex-determined eels. Population density, averaged approximately 0.14 eel m⁻² (2.42 g m⁻²) and 0.25 eel m⁻² (0.92 g m⁻²) in downstream sites of the GST and Tanshui River, respectively, and decreased substantially with upstream distance. Eels were rarely found in the heavily polluted and dam constructed areas in the midstream site of the Tanshui River.     

PCNA protein expression during spermatogenesis of the Japanese eel (Anguilla japonica)

Spermatogenesis can be initiated by a single injection of human chorionic gonadotropin (hCG) into the cultivated Japanese eel, which produces only spermatogonia in the testis. To isolate the genes responsible for regulating spermatogenesis, we performed a differential mRNA display using poly (A)+ RNA extracted from the testes at different time points after hCG injection. Among several cDNA clones, the expression of which was initiated before the onset of meiosis, one clone has high homology with the proliferating cell nuclear antigen (PCNA). In this study, we investigated the protein expression of eel PCNA and found for the first time in any species that two forms (32-kDa and 36-kDa) of PCNA are present in the testis. Although the 36-kDa form existed in both the testis and spleen, the 32-kDa form was specifically expressed in the testis. In contrast to the appearance of 36-kDa PCNA 1 day after the hCG treatment, the 32-kDa PCNA appeared only 9 days after the hCG treatment, at which time active spermatogonial proliferation occurred in the testis. Both the 32- and 36-kDa forms were recognized by antibodies raised against different epitopes of PCNA, and their N-terminal amino acid sequences were identical. The 36-kDa form, but not the 32-kDa form, was recognized by antibodies against phosphoamino acids. These results suggest that the two PCNA proteins are the same molecule with different chemical modifications, including phosphorylation. We discuss the roles of these two forms of PCNA in the spermatogenesis of the Japanese eel.     

Follicle-stimulating hormone induces spermatogenesis mediated by androgen production in Japanese eel, Anguilla japonica

Follicle-stimulating hormone (FSH) plays important roles in spermatogenesis. However, the biologic activity of FSH can vary in different vertebrate classes, and the definitive function of FSH has not been established. In this study, we investigated the functions of FSH on spermatogenesis using an in vitro culture system for Japanese eel testis. The eel Fsh receptor was expressed in testis tissue during the whole process of spermatogenesis, mainly by Leydig cells that produce steroid hormones and by Sertoli cells surrounding type A spermatogonia and early type B spermatogonia. In an in vitro organ culture, recombinant eel Fsh (r-eFsh) induced complete spermatogenesis from the proliferation of spermatogonia to spermiogenesis during 36 days of culture; also, spermatozoa were observed in the testicular fragments. Spermatogenesis induced by r-eFsh was inhibited by trilostane, a specific inhibitor of 3beta-hydroxysteroid dehydrogenase. However, trilostane did not inhibit spermatogenesis induced by 11-ketotestosterone. These results clearly show that the main function of FSH in eel is to induce spermatogenesis via stimulating androgen production.     

Dynamics of reproductive hormones during downstream migration in females of the Japanese eel, Anguilla japonica

The profiles of sex steroids (estradiol-17β, testosterone and 11-ketotestosterone) and the mRNA levels of gonadotropins (luteinizing hormone and follicle-stimulating hormone) were investigated before and after downstream migration in females of the Japanese eel species Anguilla japonica, which were collected in the brackish Hamana Lake and its inlet freshwater rivers. Eels were separated into three groups using otolith microchemistry: 'migrants' that grew in the inlet rivers and then made a downstream migration to Hamana Lake mainly in October and November; 'non-migrant' yellow eels caught in rivers during the same season; and 'residents,' which were yellow eels caught in rivers in August. Sex steroid levels, especially those of testosterone and 11-ketotestosterone, were higher in migrants than in non-migrants and residents. Real-time quantitative PCR analysis indicated that mRNA levels of luteinizing hormone (LH) β-subunits were significantly higher in migrants than in other groups, whereas those of follicle-stimulating hormone β-subunits did not show significant changes during downstream migration. The high levels of these hormones during downstream migration raise the question about if they also play a role in motivating the migratory behavior of eels.     

11-ketotestosterone synchronously induces oocyte development and silvering-related changes in the Japanese eel, Anguilla japonica

To evaluate the effects of sex steroids on silvering in the Japanese eel, Anguilla japonica, the development of oocytes, eye size, digestive tract, and swim bladder were studied in relation to observations of the profiles of plasma levels of sex steroids (estradiol 17β, E2; testosterone, T; 11-ketotestosterone; 11-KT) during silvering for each sex and by administrating 11-KT to yellow eels. All steroids examined in the study increased in female eels after silvering had begun, whereas in males, only 11-KT increased significantly, and no statistical differences were found in plasma levels of E2 and T between eels in both developmental stages. 11-KT appeared to induce the early stage of oocyte growth, enlargement of the eyes, degeneration of the digestive tract and the development of the swim bladder. This suggested that 11-KT synchronously accelerates early development of the ovaries and the morphological changes, possibly in adaption to oceanic migration, and that 11-KT is one of the most important factors in early stages of development in the Japanese eel, as it appears to be in other anguillid eels.