Development of an in vitro culture system for producing eel larvae from immature ovarian follicles in Japanese eel <Emphasis Type="Italic">Anguilla japonica</Emphasis>

To standardize conditions during the final maturation and ovulation of ovarian follicles from Japanese eel, we have developed a culture system for the production of fertilizable eggs from post-vitellogenic ovarian follicles in vitro. Post-vitellogenic ovarian follicles were incubated in culture medium supplemented with 17α,20β-dihydroxy-4-pregnen-3-one (DHP) with or without bovine serum albumin (BSA) to assess the effects of protein concentration. Eggs that ovulated during incubation were fertilized, and the remaining follicles were incubated in prostaglandin F_2α (PGF_2α) for a further 3 or 6 h before fertilization. Male eels were injected repeatedly with human chorionic gonadotropin. The quality of eggs obtained under the different culture conditions was evaluated after artificial fertilization in terms of hatching success. Hatching rates tended to decrease with increasing concentrations of BSA in the incubation medium in a dose-dependent manner. The addition of PGF_2α drastically increased the number of eggs that ovulated, but the rate of hatching was greatly decreased compared with eggs obtained earlier by DHP incubation alone. The larvae obtained from artificially fertilized eggs produced in vitro survived for 14 days without feeding. We conclude that in vitro culture systems thus have a great potential for the acquisition of good quality eggs under tightly controlled artificial conditions, culminating in the production of eel larvae.     

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.     

Relationship between coastal water properties and adult return of chum salmon (Oncorhynchus keta) along the Sanriku coast,Japan

We investigated the relationship between adult (age‐4) return rates for chum salmon (Oncorhynchus keta) originating from the Iwate Prefecture, Japan, and coastal environmental conditions during their early ocean life in coastal residency. We analyzed distributions of water properties via intensive hydrographic observations using a conductivity–temperature–depth profiler. Both the return rates and water properties vary strongly over interannual time scales. We found that the time when the return rate decreased drastically corresponded well to the time when the frequency of warm waters in the coastal residency increased.     

Immunolocalization of steroidogenic enzymes (P450scc, P450c17, P450arom) in gonad of Japanese eel

Antibodies against P450scc, P450c17 and P450arom were generated using recombinant proteins. In eel testis, P450scc and P450c17 were immunolocalized as clusters in Leydig cells. In vitellogenic eel ovary, P450scc and P450c17 immunoreactive cells were localized as clusters in the outer layer of the ovarian follicle. In contrast, P450arom seemed to be immunolocalized in the innermost follicle layer.     

Effect of temperature decrease on oocyte development,sex steroids,and gonadotropin β-subunit mRNA expression levels in female Japanese eel <Emphasis Type="Italic">Anguilla japonica</Emphasis>

To improve understanding of the mechanism of early ovarian development in eels, the effects of water temperature decrease on oocyte development, plasma levels of sex steroids [estradiol 17β (E2), testosterone (T), 11-ketotestosterone (11-KT)], and gonadotropin β-subunit [follicle-stimulating hormone (FSHβ), luteinizing hormone (LHβ)] messenger RNA (mRNA) expression levels were investigated. A total of 27 female Japanese eels Anguilla japonica were divided into initial, control, and test (water temperature decrease) groups. Starting on 22 September 2009, eels in the test group were reared in a tank with gradual temperature decrease from 25°C to 15°C over 39 days, while the control group was maintained at 25°C. The test group accumulated more oil droplets in their oocytes than did the other groups. Levels of sex steroids, especially 11-KT, were higher in the test group. In contrast, FSHβ and LHβ mRNA expression levels were lower in the test group. These results suggest that water temperature decrease only induced an early stage of ovarian development that was partly affected by an 11-KT increase. For further maturation, other environmental factors related to induction of gonadotropin increase appear to be needed.     

Soil seed banks in a mature Hinoki (<Emphasis Type="Italic">Chamaecyparis obtusa</Emphasis> Endl.) plantation and initial process of secondary succession after clearcutting in southwestern Japan

To profile secondary succession after clearcutting a conifer plantation, we examined the composition of the soil seed bank and the process of vegetation recovery after clearcutting a mature Hinoki (Chamaecyparis obtusa) plantation in southwest Japan. We set up two study plots and collected 25 and 26 soil samples (each 20 cm × 30 cm × 5 cm depth), respectively, in April 1999, before clearcutting. Using the seedling emergence method, we detected 500–839 buried viable seeds m⁻² in 42–44 species. Soil seed banks in both plots contained typical pioneer species but few late-successional species. After clearfelling the plantation in June 1999, we censused seedling establishment for 4 years and the composition of the vegetation for 6 years. More than half of the seedlings emerged in the second year, suggesting that the timing of clearcutting is possibly critical to seedling establishment. Nineteen plant species disappeared after clearcutting, 40 species were retained and 43 species newly emerged after clearcutting. Most of the newly emerged species were pioneer species which originated in the soil seed bank. Seven of the pioneer species, as typified by Rubus crataegifolius, showed clonal growth from the third year after clearcutting. Japanese serow browsed the seedlings of major pioneer species, possibly affecting the process of secondary succession. Late-successional species such as Abies firma declined in coverage or disappeared after clearcutting. Our results suggest that the soil seed bank makes a major contribution to vegetation recovery after clearcutting, but possibly not to restoration of the late-successional species.     

Relationship between abundance and localization of maternal messenger RNA and egg quality in artificially matured Japanese eel Anguilla japonica

Fisheries Science - Artificial maturation and ovulation of Japanese eel do not always result in the production of good-quality eggs. The molecular basis for compromised egg quality in artificially...     

A soil seed bank in a mature conifer plantation and establishment of seedlings after clear-cutting in southwest Japan

Compositions and spatial distribution patterns of buried viable seeds, and demographic transition from buried viable seeds to seedlings after clear-cutting were examined in a 75-year-old conifer plantation in southwest Japan. A part of the plantation was cleared in December 1997 and study plots were set up on a cleared site and a preserved site. Soil samples (a total of 105 l for the preserved plot and 90 l for the cleared plot) were collected from surface soils in spring 1998 and were incubated in a greenhouse for 120 days. There were an estimated 1064 buried viable seeds per square meter from 67 species in the preserved plot and 1754 seeds/m² from 53 species in the cleared plot. The soil seed bank was from mostly low density species with some high density species, and contained many pioneer species that were absent in the floor vegetation. Buried viable seeds were clumped together, especially in gravity-dispersal species. Although plenty of buried viable seeds were pooled in the soil, few seedlings emerged on the forest floor. In contrast, a number of seedlings, including many pioneer species, emerged in the cleared plots, showing a positive correlation between seed weight and germination rate. It is assumed that dominance of the seedlings established after clear-cutting depends on germination rates and/or growth rates of the seedlings rather than the initial density of buried viable seeds. These results suggest strong contribution of the soil seed bank to the establishment of the secondary vegetation after disturbance in a mature conifer plantation.