Functional analysis of thermo-sensitive TRPV1 in an aquatic vertebrate,masu salmon (Oncorhynchus masou ishikawae)

Transient receptor potential vanilloid 1 (TRPV1) is mainly expressed in nociceptive primary sensory neurons and acts as a sensor for heat and capsaicin. The functional properties of TRPV1 have been reported to vary among species and, in some cases, the species difference in its thermal sensitivity is likely to be associated with thermal habitat conditions. To clarify the functional properties and physiological roles of TRPV1 in aquatic vertebrates, we examined the temperature and chemical sensitivities of TRPV1 in masu salmon (Oncorhynchus masou ishikawae, Om) belonging to a family of salmonids that generally prefer cool environments. First, behavioral experiments were conducted using a video tracking system. Application of capsaicin, a TRPV1 agonist, induced locomotor activities in juvenile Om. Increasing the ambient temperature also elicited locomotor activity potentiated by capsaicin. RT-PCR revealed TRPV1 expression in gills as well as spinal cord. Next, electrophysiological analyses of OmTRPV1 were performed using a two-electrode voltage-clamp technique with a Xenopus oocyte expression system. Heat stimulation evoked an inward current in heterologously expressed OmTRPV1. In addition, capsaicin produced current responses in OmTRPV1-expressing oocytes, but higher concentrations were needed for its activation compared to the mammalian orthologues. These results indicate that Om senses environmental stimuli (heat and capsaicin) through the activation of TRPV1, and this channel may play important roles in avoiding environments disadvantageous for survival in aquatic vertebrates.     

Individual growth variation of red-spotted masu salmon,Oncorhynchus masou rhodurus,in a mountain stream

Individual growth patterns in red-spotted masu salmon,Oncorhynchus masou rhodurus, were examined by mark and recapture in a mountain stream, central Japan. The growth pattern varied substantially among individuals of the same age cohort in the stream. Mean absolute growth rates of the individuals were neither significantly different between years, study sections along the stream course, or sexes, but showed significant differences between seasons. No correlation was found between the individual growth rate of fish and the area of pools they inhabited. However, there was considerable growth variation among inhabitants of the same pool. Within a pool, larger individuals grew more rapidly than smaller ones, despite there being no significant relationship between individual growth rates and initial body weights in the stream overall. Individual growth differences probably resulted from growth depensation caused by intraspecific competition within individual pools.     

Proteomic analysis of inviable salmonid hybrids between female masu salmon Oncorhynchus masou masou and male rainbow trout Oncorhynchus mykiss during early embryogenesis

Early embryos of inviable hybrids between female masu salmon Oncorhynchus masou masou and male rainbow trout Oncorhynchus mykiss at 9, 12, 15 and 20 days after fertilization were examined for protein expression profiles. A total of 44 proteins, mostly down-regulated products of house-keeping genes and those involved in nucleic acid metabolism or chromatin replication, were identified in hybrid embryos by mass spectrometry analysis and protein database searching. The identified down-regulated proteins may be responsible for the inviability in the hybrids.     

Anadromous red-spotted masu salmon (Oncorhynchus masou ishikawae), a southernmost sea-migration form of salmonid,displays low variation in both age at seaward migration and sea age

We examined variations in age at seaward migration and sea age for the anadromous form of red-spotted masu salmon (Oncorhynchus masou ishikawae) in two Japanese rivers. The anadromous form of red-spotted masu salmon expressed only two sea migration patterns in the two rivers: (a) the majority of the salmon (95%, n = 81) were of age-0, and age-1 migrants were rare (n = 4); and (b) all the salmon examined (n = 22) made a return migration within a year, with 23% of the salmon exhibiting potamodromy in the river. Owing to low variation in their sea migratory patterns, the anadromous form of red-spotted masu salmon is likely vulnerable to environmental fluctuations.     

The lateral line system and its innervation in the masu salmon Oncorhynchus masou masou (Salmonidae)

Ichthyological Research - The lateral line system and its innervation were examined in the masu salmon Oncorhynchus masou masou. The species has 8 cephalic canals (supraorbital, infraorbital, otic,...     

Effects of melatonin feeding on smoltification in masu salmon (Oncorhynchus masou)

Influences of photoperiod on plasma melatonin profiles and effects of melatonin administration on long-day-induced smoltification in masu salmon (Oncorhynchus masou) were investigated in order to reveal the roles of melatonin in the regulation of smoltification in salmonids. Under light-dark (LD) cycles, plasma melatonin levels exhibited daily variation, with higher values during the dark phase than during the light phase. The duration of nocturnal elevation under short photoperiod (LD 8:16) was longer than that under long photoperiod (LD 16:8). Melatonin feeding (0.01, 0.1 and 1 mg/kg body weight) elevated plasma levels of melatonin in a dose-dependent manner for at least 7 h but not for 24 h. When masu salmon reared under short photoperiod were exposed to long photoperiod (LD 16:8) and fed melatonin (1 mg/kg body weight) 7 hours before the onset of darkness, a significantly smaller proportion of smolts appeared in the melatonin-fed group after 32 days than in the control group. However, after 59 days of the treatment, there was no difference in the proportion of smolts between the control and melatonin-treated groups. Thus, melatonin feeding mimicked the effects of short photoperiod, which delays but does not completely suppress smoltification. These results indicate that the day length is transduced into changes in the duration of nocturnal elevation in plasma melatonin levels, and that artificial modification of the plasma melatonin pattern possibly delays the physiological processes of smoltification induced by long-day photoperiodic treatment.     

Effects of stocking hatchery fish on the phenotype of indigenous populations in the amago salmon Oncorhynchus masou ishikawae in Japan

The expression of colour marks (parr marks, red and black spots) of the amago salmon Oncorhynchus masou ishikawae was compared with microsatellite information, to see the effects of stocking hatchery fish on the phenotype of indigenous populations, which face extinction through extensive stocking. A Bayesian-based assignment test suggested introgression of two exotic clusters into one indigenous cluster in the stocked area and its vicinity. The number of parr marks was significantly higher in one hatchery-origin population, which exclusively comprised one exotic cluster. An increased number of red spots in stocked hatchery fish was probably a consequence of hatchery feeding conditions. The number of black spots was correlated with body size in many populations, except for hatchery and heavily introgressed populations. Coefficients of correlation and regression of black spots with body size, which were largest in indigenous populations, decreased with an increase of introgression by hatchery fish. As indigenous populations have low genetic diversity with high relatedness, it was inferred that the height of correlation and regression coefficients in black spots is caused by high genetic homogeneity and fixation of alleles in loci related to the increase of black spots, both of which might have collapsed with introgression by hatchery fish. These results suggest the possibility that introgression by stocked fish causes a change of phenotype in indigenous populations.     

Restricted movement of the fluvial form of red-spotted masu salmon,Oncorhynchus masou rhodurus,in a mountain stream,Central Japan

Movement of the fluvial form of red-spotted masu salmon (1⁺ and older),Oncorhynchus masou rhodurus, was studied using mark-recapture methods in a Japanese mountain stream. Most (63–91%) adult salmon were recaptured in the pool in which they were marked. The rest of the salmon moved upstream or downstream <20m during the non-breeding period. The proportion of the salmon moving increased slightly during the breeding period, but did not exceed 66%. The distance moved was also more variable during this period. The proportion of the smaller salmon which moved was larger than that of the larger fish during the non-breeding period. Conversely, during the breeding period, larger fish moved more frequently. Sedentary behaviour and local movements of adult salmon seem to be affected by their social relationships.     

Genetic diversity and differentiation of masu salmon (Oncorhynchus masou masou) between and within cultured populations inferred from microsatellite DNA analysis

Masu salmon, Oncorhynchus masou masou, is one of the most valuable fishery species that has been introduced to China, though to date no studies on the genetic diversity and genetic relationship among hatchery populations has been performed with molecular markers. We undertook such a study and sampled 120 individuals from three hatchery stocks and analyzed 20 microsatellite loci. All loci were polymorphic and a total of 91 alleles were detected. A relatively low level of genetic diversity was revealed with effective number of allele of 3.1094, 3.3299 and 3.1894 and expected heterozygosity of 0.6600, 0.6648 and 0.6638 in the three stocks, respectively. Deviations from Hardy-Weinberg equilibrium were found due to heterozygote deficit. Accordingly, evidence of genetic bottlenecks were found in the three stocks. An individual assignment test demonstrated that 85% of individuals were correctly assigned into their original stocks. Pairwise Fst revealed that significant differentiation occurred between these three stocks. The results of the study indicated that disequilibrium of genetic structure and differentiation has occurred in all three stocks. This information collectively provides a basis for measures to avoid of loss of genetic diversity and introgression in Chinese aquaculture.     

Genetic diversity and differentiation of masu salmon (Oncorhynchus masou masou) between and within cultured populations inferred from microsatellite DNA analysis

Masu salmon,Oncorhynchus masou masou,is one of the most valuable fishery species that has been introduced to China,though to date no studies on the genetic diversity and genetic relationship among hatchery populations has been performed with molecular markers.We undertook such a study and sampled 120 individuals from three hatchery stocks and analyzed 20 microsatellite loci.All loci were polymorphic and a total of 91 alleles were detected.A relatively low level of genetic diversity was revealed with effective number of allele of 3.1094,3.3299 and 3.1894 and expected heterozygosity of 0.6600,0.6648 and 0.6638 in the three stocks,respectively.Deviations from Hardy-Weinberg equilibrium were found due to heterozygote deficit.Accordingly,evidence of genetic bottlenecks were found in the three stocks.An individual assignment test demonstrated that 85% of individuals were correctly assigned into their original stocks.Pairwise Fst revealed that significant differentiation occurred between these three stocks.The results of the study indicated that disequilibrium of genetic structure and differentiation has occurred in all three stocks.This information collectively provides a basis for measures to avoid of loss of genetic diversity and introgression in Chinese aquaculture.     

Interpopulation variation in reproductive traits of female masu salmon, Oncorhynchus masou

Several life history models predict that larger eggs and lower fecundity should be favored in a low-growth environment. We applied the model of Sibly et al. to seven Japanese populations of masu salmon ( Oncorhynchus masou ) in order to test the predictions that populations in which individual growth rate is low are characterized by larger eggs and lower fecundity. Two populations, Shumarinai and Shikaribetsu, had the lowest growth rates of individuals, largest egg sizes and lowest fecundities of all populations examined. In contrast, the Shiribetsu, Chitose and Uono populations had highest growth rates of individuals, smallest egg sizes and highest fecundities. The Shibetsu and Toya populations were intermediate between these two groups. The gonadsomatic index (GSI) of the Shumarinai and Shikaribetsu populations was smaller than that of all other populations. Correlation analysis indicated that populations with lower individual growth rates had larger eggs and lower fecundities. The results were consistent with the predictions of the Sibly et al. model: increased egg size which results in decreased fecundity is probably an adaptation to low-growth environments. Therefore, in masu salmon, growth differences among populations may explain interpopulation variation in egg size and fecundity.     

Serum immunoglobulin M (IgM) during early development of masu salmon (Oncorhynchus masou).

1. The development of serum immunoglobulin M (IgM) was studied by measuring IgM concentration in early developmental stages of masu salmon (Oncorhynchus masou) using single radial immunodiffusion. 2. IgM concentration was measurable from 88 days after hatching (46.5 micrograms/ml) and was maintained at a relatively constant level (less than 100 micrograms/ml) until 235 days after hatching. 3. IgM concentration increased significantly to a level of 691 +/- 37 micrograms/ml (mean +/- SE, N = 59) during the period from 251 to 429 days after hatching. The highest level of serum IgM was maintained from 429 days to 489 days after hatching. 4. The increase of IgM concentration was not accompanied by parallel increases in total serum protein concentration.     

Purification of serum precursor proteins to vitelline envelope (choriogenins) in masu salmon,Oncorhynchus masou

Three vitelline envelope-related proteins (VERPs), very-high-molecular-weight VERP (vhVERP), high-molecular-weight VERP (hVERP) and low-molecular-weight VERP (lVERP) were purified from female masu salmon serum. The apparent molecular weights of vhVERP, hVERP and lVERP, in their native state, were 520, 88 and 54 kDa, respectively, by gel-filtration chromatography. Very-high-molecular-weight VERP comprises two subunits, corresponding to 175 and 126 kDa. On SDS-PAGE, hVERP and lVERP migrate at 53 and 47 kDa, respectively. Amino acid analysis of vhVERP and hVERP showed that they share a high content of glutamic acid and proline. By contrast, lVERP is rich in glutamic acid and asparatic acid. These features are in good agreement with the amino acid composition of the vitelline envelope. Immuno-biochemical analysis suggested that vhVERP is derived from hVERP by polymerization and/or aggregation. Antibodies against hVERP and lVERP specifically immunostained the vitelline envelope and liver of female masu salmon. In addition, both hVERP and lVERP were induced in the serum of estrogen-treated male fish. Taken together, it is suggested that hVERP and lVERP are homologous molecules with choriogenin H and choriogenin L in medaka, respectively. These results indicate that hVERP and lVERP are precursor proteins to the vitelline envelope (choriogenins) in masu salmon.     

Dispersal of wild and domestic masu salmon fry (Oncorhynchus masou) in an artificial channel

The day and night pattern of upstream and downstream dispersal of masu salmon fry of wild and domestic origin was compared in artificial channels (45 m long), for two ages of planting: unfed alevins and eyed eggs. Early dispersal was important for the wild stock (48–50%) compared with the domestic one (16–36%). More wild fry moved downstream than upstream, and more domestic fry dispersed upstream. Upstream movement in wild and domestic fry was more active by day than by night, except for wild fry planted as eyed eggs, where upstream migration was higher at night. In contrast, downstream movement in wild and domestic fry was more common by night than by day, but daylight catches were not negligible for the wild stock.     

Molecular cloning and characterization of three distinct choriogenins in masu salmon, Oncorhynchus masou

Three cDNAs, each encoding a different choriogenin (Chg), were isolated from a female masu salmon (Oncorhynchus masou) liver cDNA library. Two of the cDNA clones, Chg Halpha and Chg Hbeta, showed a close relationship and contained the typical domains of zona pellucida (ZP) B genes in fish, namely proline and glutamine rich repeats, a trefoil factor family domain, and a ZP domain. Specific antibodies against recombinant Chg H products (rmHalpha and rmHbeta) were generated to elucidate the relationship between the Chg H cDNAs and two types of serum Chg H protein, which were previously purified and characterized, and designated as very-high-molecular-weight vitelline envelope-related protein (vhVERP) and Chg H of masu salmon. The immunobiochemical analyses revealed that the Chg Halpha and Chg Hbeta clones encoded vhVERP and Chg H proteins, respectively. The third cDNA clone (Chg L) appeared to be a ZPC gene and, by mapping the N-terminal sequence of purified Chg L, was shown to encode serum Chg L protein. Various types of heteromultimer of the three Chgs were identified immunologically as high molecular weight chorion components, indicating the involvement of complex heterodimerization of multiple Chgs in the construction of chorion architecture in masu salmon.     

Ascending speed and nocturnal activity of adult masu salmon (Oncorhynchus masou Brevoort) during upstream migration

We released five adult masu salmon (Oncorhynchus masou) tagged with external transmitters to track their ascending behaviour. The signals of all specimens were recorded in the upper area of the river system. Two patterns of ascending behaviour were recognized: ascending upward immediately after release and ascending during increased river discharge. The fastest ascending speed was about 1000 m h^–1. Active movements were detected at night. The signal recording duration at each receiver for each fish was generally brief. Most fish did not stay at the pools where the receivers were installed.     

Relationship between eyed-egg percentage and levels of cortisol and thyroid hormone in masu salmon Oncorhynchus masou

A possible cause of the low eyed-egg percentage that afflicts pond-reared masu salmon Oncorhynchus masou at a fish hatchery (Kumaishi Fish Hatchery, Hokkaido, Japan) was investigated. Serum cortisol levels of masu salmon during the spawning period were compared between individuals from Kumaishi and those from another station (Mori). Cortisol, thyroxine (T₄) and triiodothyronine (T₃) hormones were also measured in eyed eggs and their levels were compared to the eyed-egg percentage. Serum cortisol levels were significantly higher at Kumaishi than at Mori in May and July of the breeding season. At the Kumaishi station, there was a linear positive relationship between serum cortisol and fertilized-egg cortisol levels of female masu salmon. As cortisol levels in both ovulating females and eyed eggs increased, the eyed-egg percentage decreased, indicating a highly significant negative relationship between cortisol levels and eyed-egg percentage. In contrast, as T₃ and T₄ levels in eyed eggs increased, the eyed-egg percentage increased. The strong positive correlation between high cortisol levels in serum and in eyed eggs at Kumaishi indicates that the frequently higher cortisol levels may have caused the lowering of the eyed-egg percentage.     

Effect of restricted feeding on early maturation in female and male amago salmon, Oncorhynchus mason ishikawae

Incidence of early maturation of female amago salmon was reduced by >32% by feed restriction beginning in May, 5 months prior to spawning. Male early maturation was unaffected by this feed restriction regime.     

Trace of outbreeding between Biwa salmon (Oncorhynchus masou subsp.) and amago (O. m. ishikawae) detected from the upper reaches of inlet streams within Lake Biwa water system,Japan

Ichthyological Research - The establishment of fluvial fish populations from anadromous populations by natural or artificial barriers obstructing migration is a good research subject to study life...