Changes in levels of GnRH in the brain and pituitary and GTH in the pituitary in male masu salmon,Oncorhynchus masou,from hatching to maturation

Levels of two types of gonadotropin-releasing hormone (salmon GnRH and chicken GnRH-II) in the brain and pituitary, and content of gonadotropin (GTHIβ and IIβ) in the pituitary were measured in male masu salmon from hatching to gonadal maturation for three years in order to clarify the involvement of GnRHs in precocious maturation. Underyearling precocious males were distinguishable in summer of year 1 and were marked by an increased GSI. Spermiation was observed among these individuals thereafter every autumn. Pituitary GTHIβ content in both precocious and immature males, and GTHIIβ content in precocious males showed seasonal fluctuations — high in autumn and low in winter. Pituitary GTHIIβ content was low in immature males. Pituitary sGnRH content in precocious males increased from spring to autumn during the three-year period. sGnRH concentrations in discrete brain areas showed seasonal changes — high during autumn to winter and low in summer. Concentrations in the olfactory bulbs and hypothalamus increased significantly in association with testicular maturation during year 3. sGnRH concentrations in the hypothalamus were significantly higher in precocious males than in immature males; this was possibly due to positive feedback of steroid hormones. cGnRH-II was undetectable in the pituitary and no distinct changes were observed in its concentration in the brain in relation to maturation. The phenomenon of underyearling precocious maturation is considered to be triggered before the onset of early summer. It is suggested that males which mature precociously are larger in size and contain much sGnRH in the pituitary before the outward signs of precocity appear; sGnRH may stimulate GTH II synthesis and induce precocious maturation.     

In vivo and in vitro effects of Rhizopus extract on body growth and steroid production in masu salmon, Oncorhynchus masou Brevoort

This study investigated the in vivo and in vitro effects of Rhizopus (filamentous fungus) extract (RU) in masu salmon Oncorhynchus masou Brevoort. Underyearling fish were fed with RU for 16 months. Monthly changes in body growth, gonadal maturation and serum levels of sex steroids were monitored. Gonads were also incubated at 0, 1, 10, 100 and 1000 μg RU mL⁻¹ Leibovitz's L‐15 medium for 18 h. The levels of steroids in serum and cultured medium were measured. It was determined that RU‐fed immature and mature males, when compared with control groups, showed significantly higher body growth during spring, summer and the spawning period. Similarly, immature RU‐fed females showed significantly higher fork length and body weight in autumn, spring and summer. Furthermore, RU‐fed males showed significantly higher levels of serum testosterone (T) and 11‐ketotestosterone (11‐KT) levels in the pre‐spawning season, and 17α, 20β‐dihydroxy‐4‐pregnen‐3‐one (DHP) in the spawning season. In vitro RU incubation of gonads showed a dose‐dependent and significant increase in T, 11‐KT, oestradiol‐17β and DHP release in the medium. It appears that the causes of enhanced body growth and increased steroid production herein observed in salmonids are the physiologically active substances that may be present in the mycelium of the fungus.     

Regulation of LH synthesis and release by GnRH and gonadal steroids in masu salmon

The roles of salmon GnRH (sGnRH) and gonadal steroid hormones in regulation of LH synthesis and release were examined in primary pituitary cell cultures of masu salmon (Oncorhynchus masou). Pituitaries were taken from fish at four reproductive stages: in March (initiation of sexual maturation); May (early maturation); July (pre-spawning); and September (spawning period). Amounts of LHβ subunit mRNA in the pituitary cells were determined by real-time PCR, and LH levels in the medium were determined by RIA. sGnRH and gonadal steroids including estradiol-17β (E2), testosterone (T) and 11-ketotestosterone (11-KT) were added to the cultures to examine their direct effects on LH response. sGnRH had no significant effect on LHβ mRNA levels at any stages, although a stimulatory trend was noted in March. In contrast, E2 and T considerably increased LHβ subunit mRNA levels in March and May during initial stages of maturation, and the effects were less pronounced in July and September. On the other hand, sGnRH stimulated LH release at all stages in the males and the effects were most prominent in July and September. E2 and T also stimulated LH release in July and September, but their effects were weaker than that of sGnRH. The present results indicate that sGnRH and gonadal steroids directly regulate LH synthesis and release in masu salmon pituitary cells: sGnRH mainly stimulates LH release in the late stage of sexual maturation; whereas, E2 and T are effective in stimulating LH synthesis at earlier stages of maturation.     

Gonadotropin-releasing hormone and gonadotropin in goldfish and masu salmon

Reproductive activities in vertebrates are regulated by an endocrine system, consisting of the brain-pituitary-gonad axis. In teleosts, gonadotropin-releasing hormone (GnRH) in the brain stimulates gonadotropin (GTH) release in the pituitary gland, but because of lack of the portal vessel, it is not known when and how much GnRH is released for the regulation of GTH release. There are multiple molecular types of GnRH in teleosts and several distinct populations of GnRH neurons in the brain. However, we do not know which types and populations of GnRH neurons regulate reproductive activities. Here we summarize our recent studies on GnRH and GTH in masu salmon Oncorhynchus masou and goldfish Carassius auratus. Immunocytochemistry showed the location and molecular types of GnRH neurons. Salmon (sGnRH) and chicken-II GnRH (cGnRH-II) neuronal fibers were widely distributed in the brain of both masu salmon and goldfish. Only sGnRH fibers were observed in the pituitary of masu salmon, whereas both sGnRH and cGnRH-II fibers were observed in the goldfish pituitary, indicating that species specific GnRH profiles are involved in the regulation of pituitary function in teleosts. A series of experiments in masu salmon and goldfish suggest that among GnRH neuron populations GnRH neurons in the ventral telencephalon and the hypothalamus regulate GTH release, and that GnRH of the terminal nerve origin is not essential to gonadal maturation and ovulation. The biological function of other GnRH neurons remains unkown. Two GTHs appear to be characteristic of teleost; however, regulation of reproduction by these GTHs is a question that remains to be elucidated. In salmonid species, it is proposed that GTH I stimulates early gonadal development, whereas GTH II acts in later stages. When GTH expression was examined in goldfish, both GTH I and II mRNA levels in the pituitary gland showed increases in accordance with gonadal development, unlike the sequential expression of GTH subunits in salmonids. The expression of these GTH subunit mRNAs were affected by water temperature, starvation, and steroid hormones in goldfish, but in what manner these two GTHs regulate gonadal development remains to be clarified.     

Dominance and predator avoidance in domesticated and wild masu salmon Oncorhynchus masou

ABSTRACT:   Dominance, aggression and predator avoidance were compared among farmed, sea-ranched and wild juvenile masu salmon Oncorhynchus masou in laboratory experiments. Domesticated fish (farmed and sea-ranched), which had been exposed to artificial selection, were not dominant against wild fish in pairwise contests, nor did they show greater aggressiveness. Farmed fish did show greater feeding than wild fish. Under chemically simulated predation risk, farmed fish were more willing to leave cover and feed than wild fish, indicating reduced predator avoidance in the farmed fish. Our results indicate that selection for fast growth (domestication) in masu salmon favors fish that respond to food quickly and ignore predation risk.     

Distinct expression of GnRH genes in the red seabream brain

This paper reports the molecular cloning of a cDNA encoding the precursor of seabream gonadotropin-releasing hormone (prepro-sbGnRH) and the localization of salmon GnRH (sGnRH) and seabream GnRH (sbGnRH) expressing neurons in the brain of the red seabream (Pagrus major). The cloned prepro-sbGnRH cDNA has a 285 bps open reading frame encoding a 23 amino acid signal peptide, a 10 amino acid sbGnRH, the cleavage site (Gly-Lys-Arg), and a 59 amino acid GnRH-associated peptide. The expression of sGnRH and sbGnRH peptides, and prepro-sGnRH and prepro-sbGnRH mRNA were studied using immunocytochemistry and non-radioactive in situ hybridization, respectively. We found cell bodies that reacted positively with both the sGnRH cRNA probe and anti-sGnRH serum, but not with the sbGnRH cRNA probe or anti-sbGnRH serum in the ganglion of the terminal nerve. Cell bodies that reacted positively with the sbGnRH cRNA probe, anti-sbGnRH serum, and anti-sGnRH serum, but negatively with the sGnRH cRNA probe were found in the preoptic area (POA). Immunocytochemistry showed that a distinct bundle of axons arises in the POA which projected to the pituitary gland. These results suggest that sbGnRH is the most relevant hypophysiotropic form of GnRH.     

Importance of instream cover for young masu salmon, Oncorhynchus masou, in autumn and winter

Densities of wild masu salmon, Oncorhynchus masou Brevoort, were investigated before (late-September) and after (mid-November) autumnal habitat shifts in a small river in Hokkaido, northern Japan. Abundance of instream cover habitat formed by coarse woody debris, submerged vegetation and undercut banks was quantified. Density of 1+ and older masu salmon was significantly correlated with cover abundance in autumn, although cover was not significant for density of 0+ fish, indicating that clear-cut reaches were utilized by 0+ masu salmon in autumn. In early winter, densities of 0+ and 1+ and older masu salmon were both correlated with cover availability. These results suggest that the amount of cover habitat is an important regulator for densities of juvenile masu salmon during winter.     

Effects of cortisol and angiotensin II on the number and size of juxtaglomerular cells in masu salmon, Oncorhynchus masou

The authors previously reported that the number and size of juxtaglomerular cells (JGCs) in the kidney increased during smoltification in masu salmon, Oncorhynchus masou. In the present study, the effects of cortisol and/or angiotensin (Ang) II ([Asn¹, Val⁵]-Ang II) on the JGC number and size in masu salmon were examined to elucidate hormonal regulation of the changes in the JGC number and size during smoltification. These hormones were injected intraperitoneally every 2 days for a total of 6 injections. There was a significant increase in the JGC number and size with time following the start of the experiment in cortisol- and cortisol + Ang II-treated groups and no significant change in control and Ang II-treated groups. On both days 5 and 11, the JGC number and size in the cortisol-treated group were significantly large compared to those of control and Ang II-treated groups, respectively. The JGC number and size in the cortisol + Ang II-treated group were significantly large compared to those of control on both days 5 and 11, and those of the Ang II-treated group only on day 11, respectively. On the other hand, there was no significant difference in the JGC number and size between the Ang II-treated and control groups and between the cortisol- and cortisol + Ang II-treated groups during the experiment, respectively. The means of the JGC number and size in cortisol-treated group on day 11 were close to those previously reported in smolt. These results suggest that cortisol induces an increase in JGC number and size during smoltification in masu salmon. This revised version was published online in August 2006 with corrections to the Cover Date.     

The effect of cortisol administration on blood plasma immunoglobulin M (IgM) concentrations in masu salmon (Oncorhynchus masou)

Immunoglobulin M (IgM) is known as a main factor in the humoral immune system of teleosts. In the present study, the effect of cortisol on plasma IgM concentrations was investigated using a specific antibody to IgM in masu salmon (Oncorhynchus masou). Cortisol was orally administered each day for 2 weeks at a dose of 1 mg g⁻¹ in the diet, and for the following week the fish were fed a non-treated diet. Blood plasma samples were collected at 0, 1, 2 and 3 weeks after the initiation of treatment. Oral administration of cortisol elevated plasma cortisol concentrations to about 40 ng/ml for 2 weeks after administration and slightly reduced plasma IgM concentration; the suppression was statistically significant one week after the period of hormone administration. However, treatment with cortisol did not affect plasma concentrations of total protein or α1-protein, one of the major serum proteins, during the experimental period. These results indicate that cortisol specifically suppresses plasma IgM concentrations.     

Estimation of heritability and genetic correlation of number of abdominal and caudal vertebrae in masu salmon

ABSTRACT:   Ten sibship masu salmon Oncorhynchus masou masou families were produced to estimate the heritability of the number of abdominal vertebrae (AV) and caudal vertebrae (CV), and the genetic correlation between the two traits were calculated. The AV and CV of both the parents and their offspring were counted separately, and the heritability was estimated by performing parent–offspring regression analysis. The mean vertebral number of the offspring differed among families although they were reared under identical water temperature conditions and had the same parental vertebral number type in crossbreeding. The heritabilities were estimated as 0.65 for AV and 0.84 for CV. The genetic correlation between the number of AV and CV was calculated as −0.92. These results suggest that AV and CV are governed by genetic components even when the fish are reared under identical environmental conditions, and the total vertebral number is expected to converge within a certain range. It is necessary to analyze separately the variations in the number of AV and CV when investigating the vertebral number in masu salmon.     

Transitions of urine flow rate and kidney-somatic index in mature masu salmon

ABSTRACT:   The present study aimed to investigate functional changes of the kidney in mature masu salmon. The urine flow rate (UFR), kidney-somatic index (KSI), and epithelial cell height of second proximal convoluted segment of renal tubules were examined in mature and immature masu salmon Oncorhynchus masou . Two methods of calculating UFR were made as either the units per body weight, or body weight without gonads. The former calculation using body weight may be used for comparison between data obtained from immature fish, and the latter method was suitable when comparing mature sexes and assessing the transition from immature to finally mature stages. In both calculations of UFR, KSI, and epithelial cell height of the second proximal segment, the values for mature fish were significantly higher than for immature fish. These results first suggest that functional transition of the kidney occurs in maturing fish.     

The combined effects of temperature and GnRHa treatment on the final stages of sexual maturation in Atlantic salmon (Salmo salar L.) females

Atlantic salmon (Salmo salar L.) females (2 SW), maturing for the first time, were reared under one of three temperature regimes (high: 14.3 ± 0.5°C; natural: 10.6 ± 1.0°C; and cold: 6.9 ± 1.0°C) in combination with one of two experimental treatments; an injection of GnRH analogue (GnRHa) contained in biodegradable microspheres, or a sham injection (microspheres only). The six experimental groups were then reared under simulated natural photoperiod for 4 weeks. Blood samples were drawn for analysis of plasma steroid levels and the fish were inspected for ovulation weekly. Batches of stripped eggs were incubated in triplicate incubators in raceways until the eyed stage. Treatment with GnRHa resulted in a substantial advancement and synchronization of ovulation at all temperatures, while exposure to cold water also appeared to advance ovulation slightly. While 75% (warm and cold) to 90% (natural) of GnRHa fish ovulated during the 4-week trial, only 30% of sham-treated females exposed to cold water, and none of the sham-treated fish held at higher temperatures, ovulated during this period. Survival rates of embryos to the eyed-stage were significantly higher for broodstock exposed to cold water. Plasma levels of testosterone (T), 17β-oestradiol (E₂), and 17α,20β-dihydroxy-4-pregnen-3-one (17,20βP) were all significantly affected by treatment with GnRHa and, to a lesser extent, temperature. The efficiency of GnRHa in counteracting the negative effects of high temperature on ovulation and the associated changes in circulating sex steroids suggest that temperature inhibition operates at least in part at the brain or pituitary.     

Microsatellite variation reveals fine-scale genetic structure of masu salmon, Oncorhynchus masou, within the Atsuta River

Abstract –  To describe in detail the population genetic structure of masu salmon Oncorhynchus masou , seven anadromous populations in the Atsuta River were investigated by analysis of eight polymorphic microsatellite loci. Exact test and genetic differentiation ( F _ST) revealed that significant differentiation was observed at the within-river scale. Principal components analysis revealed that the spatial pattern of genetic composition was related to geographical location of each population. Furthermore, a significant correlation was observed between genetic differentiation and geographical distance, indicating that dispersal is less likely to occur between distant populations. The results of this study imply that masu salmon have potential to show clear genetic structure at the microgeographical scale (21 km) due to precise homing behaviour.     

Color vision, spectral sensitivity, accommodation, and visual acuity in juvenile masu salmon Oncorhynchus masou masou

ABSTRACT:   Color vision, spectral sensitivity, accommodation, and visual acuity were examined in juvenile masu salmon Oncorhynchus masou masou to obtain fundamental information about the visual system. Two types of S-potentials were recorded from 415 horizontal cells in isolated retinas from 34 cultured freshwater masu salmon (114–219 mm standard length, SL). Although horizontal cells recording S-potentials were not identified, the horizontal cells were confirmed because their responses were maintained while the stimulus remained. The variety of chromaticity (C)-type S-potentials indicated well-developed color vision. The analysis of the luminosity (L)-type S-potentials indicated that the peak spectral sensitivity was at 522 nm. S-potentials were also recorded in response to ultraviolet light. The direction and extent of lens movement induced by electrical stimulation was measured in 12 cultured masu salmon (99.0–142.5 mm SL). The results indicated that the visual axis was upward and forward, and that the range of accommodation was from 0.79 × SL in front of the eye to infinity. In histological analysis of the retinas of five wild smolts (100–118 mm SL), the maximum cone densities (276–345 cones/0.01 mm²) were detected in the ventral to temporal regions. The visual acuities assessed by histological methods were 0.069–0.075.