Persistent and reparative neurogenesis in the juvenile masu salmon <Emphasis Type="Italic">Oncorhynchus masou</Emphasis> telencephalon after mechanical injury

A superficially located periventricular proliferative area with PCNA-immunopositive (PCNA+) cells, which corresponds to the pallial periventricular zone (PVZ) of other fish species, including its dorsal, lateral, and medial compartments, is discovered in the telencephalon of the juvenile masu salmon Oncorhynchus masou. The PCNA+ cells are also identified in the parenchyma of the masu salmon intact brain, and their maximum concentration is observed in the medial zone. After a mechanical injury, the zones of induced neurogenesis—neurogenic niches and sites of secondary neurogenesis surrounded by radial glial fibers—appear in the masu salmon telencephalon. The PVZ of the juvenile masu salmon pallium contains clusters of undifferentiated HuCD-immunopositive (HuCD+) neurons. A change in the HuCD+ cell topography is observed in the mechanically injured masu salmon telencephalon, namely, neurogenic niches in the lateral zone and an increase in the cell distribution density and cell migration patterns in the medial zone. A high level of persistent neurogenesis is characteristic of the juvenile masu salmon brain.     

Neurochemical Organization and Connections of the Cerebral Preglomerular Complex of the Masu Salmon

Using immunohistochemical labeling of the cells containing neuronal NO synthase (nNOS), tyrosine hydroxylase (TH), GABA, and parvalbumin (PA), as well as histochemical marking of choline acetyltransferase-containing neurons, we examined the neurochemical organization of the glomerular nuclei and preglomerular complex in the brain of the masu salmon (Oncorhynchus masou). Injections of the carbocyanine dye DiI allowed us to examine projections of neurons of the preglomerular and mammillary nuclei in the salmon brain. We showed that cholinergic, GABA-, PA-, TH-, and nNOS-immunopositive neurons belonging to different morphological types are present in the glomerular and medial preglomerular nuclei. The analysis of correlations between morphometric characteristics of the cells belonging to different neurochemical types and densitometric estimates of amounts of neurochemical agents present in these cells allowed us to hypothesize that there are close morphofunctional interrelations in cell populations possessing different neurochemical and morphometric characteristics. These interrelations of the cells belonging to different chemotypes are, probably, realized as mediatory/modulatory ones. The presence of a great number of small slightly differentiated cells in the preglomerular and glomerular nuclei allows us to suppose that the growth of the greatest sensory center of the salmon brain is provided by neuroblasts that migrate from the proliferative zones in the course of postembryonal neurogenesis. It is also hypothesized that NO, TH, and GABA are involved in paracrine control of the postnatal morphogenesis of the salmon preglomerular complex. The data obtained by hodological analysis indicate that the nuclei of the preglomerular complex obtain afferent projections from the dorsomedial and ventroventral telencephalic regions, preoptic nucleus, periventricular layer of the tectum, and posterior central thalamic nucleus. Our study demonstrated the existence of reciprocal functional connections between the preglomerular complex (most important diencephalic center for transmission of sensory information) and dorsomedial and ventral regions of the telencephalon in the masu salmon.     

Gaseous transmitters in the brain of the masu salmon, <Emphasis Type="Italic">Oncorhynchus masou</Emphasis> (Salmoniformes,Salmonidae)

Distribution of nitroxidergic and H₂S-producing neurons in the brain of the masu salmon Oncorhynchus masou was studied by methods of histochemical labeling of NADPH-diaphorase and by immunohistochemical labeling of the neuronal nitric oxide synthase and cystathionine β-synthase (CBS). The established distribution of CBS and nNOS/NADPH-d of neurons and fibers in the masu salmon telencephalon, optic tectum, and cerebellum allows suggesting that the NO- and H₂S-producing systems represent individual, non-overlapping neuronal complexes performing specialized functions in the activity of local neuronal networks. In the medullar part, the nNOS-ir and NADPH-d-positive neurons were detected in the composition of viscerosensory (V, VII, and IX–X) and visceromotor (III, IV, and VI) nuclei of craniocerebral nerves, octavolateral afferent complex, reticulospinal neurons, and medial reticular formation. CBS in the masu salmon medulla was revealed in neurons of the nerve X nucleus, reticulospinal neurons, and ventrolateral reticular formation. Distribution of NO-ergic and H₂S-producing neurons in the masu salmon medullar nuclei indicates that NO in masu salmon is the predominant neuromodulator of the medullar viscerosensory systems, while H₂S seems to modulate only the descending motor systems. The results of the performed study allow suggesting that NO in the masu salmon medulla periventricular area can act as a regulator of postnatal ontogenesis.     

Pax‐6 is expressed early in the differentiation of a corneal epithelial model system

Pax‐6 is a regulatory gene with a major role during visual system development, but its association with corneal epithelial differentiation is not clearly established. Using the RCE1‐(5T5) cell line, which mimics corneal epithelial differentiation, we analyzed Pax‐6 biological role. Immunostaining of proliferating colonies and confluent sheets showed that Pax‐6‐positive cells were also K3 keratin‐positive, suggesting that Pax‐6 is expressed in differentiating cells. Pax‐6 mRNA was barely expressed in early cell cultures; but after confluence, its levels raised up to fivefold as demonstrated by Northern blot and RT‐qPCR. The raise in Pax‐6 expression preceded for 9 h the increase in LDH‐H and LDH‐M mRNAs, previously shown as early markers of corneal epithelial cell differentiation. The full‐length mRNAs encoding for the two major Pax‐6 isoforms were found at very low levels in proliferating cells, and abundantly expressed in the confluent stratified epithelia; Pax‐6 mRNA was 2‐ to 2.5‐fold more abundant than Pax‐6(5a) mRNA. The ectopic expression of Pax‐6 or Pax‐6(5a) decreased proliferative ability leading to the formation of abortive, non‐proliferative colonies. In contrast, culture conditions that delay or block corneal epithelial cell differentiation reduced or inhibited the expression of Pax‐6. Collectively, results show that Pax‐6 is the earlier differentiation marker expressed by corneal epithelial cells, and open the possibility for a major role of Pax‐6 as the main driver of the differentiation of corneal epithelial cells. J. Cell. Physiol. 220: 348–356, 2009. © 2009 Wiley‐Liss, Inc.     

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.     

Comparative anatomy of the dorsal hump in mature Pacific salmon

Mature male Pacific salmon (Genus Oncorhynchus ) demonstrate prominent morphological changes, such as the development of a dorsal hump. The degree of dorsal hump formation depends on the species in Pacific salmon. It is generally accepted that mature males of sockeye (O. nerka ) and pink (O. gorbuscha ) salmon develop most pronounced dorsal humps. The internal structure of the dorsal hump in pink salmon has been confirmed in detail. In this study, the dorsal hump morphologies were analyzed in four Pacific salmon species inhabiting Japan, masu (O. masou ), sockeye, chum (O. keta ), and pink salmon. The internal structure of the dorsal humps also depended on the species; sockeye and pink salmon showed conspicuous development of connective tissue and growth of bone tissues in the dorsal tissues. Masu and chum salmon exhibited less‐pronounced increases in connective tissues and bone growth. Hyaluronic acid was clearly detected in dorsal hump connective tissue by histochemistry, except for in masu salmon. The lipid content in dorsal hump connective tissue was richer in masu and chum salmon than in sockeye and pink salmon. These results revealed that the patterns of dorsal hump formation differed among species, and especially sockeye and pink salmon develop pronounced dorsal humps through both increases in the amount of connective tissue and the growth of bone tissues. In contrast, masu and chum salmon develop their dorsal humps by the growth of bone tissues, rather than the development of connective tissue.     

Occurrence of a hybrid between endemic Miyabe charr Salvelinus malma miyabei and introduced masu salmon Oncorhynchus masou masou in the Lake Shikaribetsu system, Hokkaido, Japan

A hybrid specimen between endemic Miyabe charr Salvelinus malma miyabei and introduced masu salmon Oncorhynchus masou masou was collected in an inlet stream (Yamada Creek) of the Lake Shikaribetsu. This specimen showed intermediate external characteristics between two species and was also confirmed as a hybrid by DNA markers. The mtDNA of the specimen was identical with that of S. malma miyabei, suggesting that the mating occurred between a female Miyabe charr and a male masu salmon. Hybridization with introduced masu salmon may cause deleterious effects on S. malma miyabei.     

Genetic differentiation between collections of hatchery and wild masu salmon (<Emphasis Type="Italic">Oncorhynchus masou</Emphasis>) inferred from mitochondrial and microsatellite DNA analyses

There has been very little effort to understand genetic divergence between wild and hatchery populations of masu salmon (Oncorhynchus masou). In this study, we used mitochondrial (mt) NADH dehydrogenase subunit 5 gene (ND5) and six polymorphic nuclear microsatellite DNA loci to compare the genetic variability in three hatchery broodstocks of masu salmon with the variability in eight putative wild masu populations sampled in five rivers including one known source river for the hatchery broodstocks. Both ND5 and microsatellites showed no significant genetic divergence (based on F_ST estimates) between four annual collections from the source river population, suggesting no change in genetic diversity over this time period. The F_ST estimates, an analysis of molecular variance (AMOVA), and a neighbor-joining tree using both DNA markers suggested significant differentiation between the three hatchery and all eight putative wild populations. We conclude that genetic diversity of hatchery populations are low relative to putative wild populations of masu salmon, and we discuss the implications for conservation and fisheries management in Hokkaido.     

GFAP and PCNA Marking in the cerebellum of masu salmon’s (<Emphasis Type="Italic">Oncorhynchus masou</Emphasis>) juvenile after mechanical injury

The objective of this work was to study proliferation processes and the role of glia and neural stem cells in the event of injurious action on cerebellum of masu salmon’s (Oncorhynchus masou) juvenile. Using the immunoperoxidase staining of the glial fibrillary acidic protein (GFAP) and proliferating cells nuclear antigen (PCNA), processes of proliferation and gliogenesis after mechanical trauma of cerebellum of cherry salmon’s (Oncorhynchus masou) juvenile were studied. After the trauma, the intensity of proliferation and migration processes varies in different zones. Proliferation processes decrease after the trauma in lateral and basal zones, and migration increases. In the dorsal zone, on the contrary, migration processes significantly decrease and proliferation increases. In the dorsal matrix zone of a cerebellum, intense cell proliferation was detected. In the dorsal, lateral, and basal zone of the molecular layer of cerebellum after traumatic damage, neurogenic niches containing PCNA and cells, as well as a heterogeneous population of PCNA-cells, were identified. At the location of neurogenic niches, fibers of radial glia and small single intensely or moderately labeled GFAP cells were discovered. As a result of damaging action, GFAP+ fibers of radial glia, which form differently directed radially oriented bundles, appeared in the dorsal matrix zone. Such structural formations have not been discovered in intact animals. We suppose that, after the trauma, structural reconstruction connected with partial spatial reorientation of the radial glia fibers and formation of specific directions for cells formed in this zone occurs in the dorsal matrix zone. As a result of the trauma, in masu salmon’s cerebellum, elements of the radial glia, including both cells possessing typical morphology and cell fragments presented as long radially oriented processes or cell body containing initial fragments of radial fibers, appeared.     

Genetic differentiation and the problems of conservation of masu salmon (<Emphasis Type="Italic">Oncorhynchus masou</Emphasis> Brevoort, 1856 (Pisces: Salmonidae)) populations

This study focuses on the strategy for the conservation of masu salmon, Oncorhynchus masou, in the northern part of the species range (via the masu populations in Sakhalin Oblast), based on data of its population structure. It is shown that masu populations that inhabit different rivers genetically differ from each other in allele frequencies at microsatellite markers. In the Naiba River basin, at least two genetically distinct masu populations exist: in the upper reaches and in a tributary, the Bolshoy Takoy River. The masu populations on Iturup Island significantly differ from those on Sakhalin Island; within Sakhalin, the masu salmon from the Chernaya River in the southwestern part of the island is genetically distinct from the southeastern Sakhalin and Aniva Bay populations. The genetic diversity of Iturup populations is substantially lower than that on Sakhalin, probably due to their small sizes. The measures for the conservation and recovery of masu salmon populations should be based primarily on their own genetic resources, or, in the case of a lack of spawners, on the base populations of their ecological/geographical region. In the latter case, masu populations of large rivers can be considered as base ones: for southeastern Sakhalin, this is masu salmon of the Naiba River; for Aniva Bay, this is masu salmon of the Lyutoga River. Transplantation of fish, fertilized eggs, or any other genetic material from a population that is different genetically and inhabits the waters with different ecological gradients should be strongly restricted. The formosan masu salmon from Taiwan Island is studied as an example of a strict genetic isolate.     

Reproduction ecology of masu salmon <Emphasis Type="Italic">Oncorhynchus masou</Emphasis> in the Kol basin (Western Kamchatka)

In the Kol basin, one of the typical salmon rivers of Western Kamchatka, special traits of reproduction ecology of masu salmon Oncorhynchus masou are studied (characteristics and behavior of spawners during spawning, localization of spawning grounds, topography and hydrology of redds, and interaction with other species of salmonids). The masu salmon spawns in the Kol basin in tributaries of the upper reaches of the river and only in downwelling. Spawning grounds of masu samlmon are confined to stretches with overhanging banks, log jams creating shelters for spawners and favorable hydrological conditions for spawning and egg development. All over its range, masu salmon requires similar conditions for reproduction. In the north of its range, in Kamchatka, masu salmon retains the properties of the most warm-water species in the genus Oncorhynchus and selects for spawning the grounds characterized by the highest temperature in the period of spawning and development.     

Strategies for the conservation and management of isolated salmonid populations: lessons from Japanese streams

1. Endangered native populations of stream salmonids in Japan face three major threats: (i) negative interactions with introduced hatchery‐reared fish, (ii) fragmentation of habitat by impassable dams and (iii) recreational angling. 2. To prevent imminent extinction of many local populations, we evaluated these threats and possible conservation actions for red‐spotted masu salmon (Oncorhynchus masou ishikawae) and white‐spotted charr (Salvelinus leucomaenis japonicus) in the Fuji River system in central Japan. 3. Red‐spotted masu salmon and white‐spotted charr occupied only 0.73 and 2.4% of suitable thermal habitats, respectively, with masu salmon typically occupying habitats closer to human population centres. 4. Population viability analysis resulted in a 100‐year probability of extinction of 78.1% for masu salmon and 48.1% for charr. However, extinction risk of both species was predicted to be <5% if the carrying capacity increased from 141 to 303 for masu salmon and from 94 to 125 for charr, by allowing fish passage at the lower end of the habitat, and if annual adult survival rate increased by 0.04. Adult survival rate was the principal factor associated with population persistence. 5. To conserve isolated populations of stream‐dwelling salmonids, we recommend (i) assessing the distribution of remnant native and non‐native fish populations, (ii) that fishing regulations are modified to improve adult survival and population persistence and (iii) that fragmented reaches be reconnected to adjacent habitat, for example by removing or modifying artificial barriers to increase the carrying capacity of the isolated populations. Reconnection of fragmented reaches should, however, be avoided if it results in non‐native fish invading isolated populations.     

Structure and developmental expression of hatching enzyme genes of the Japanese eel<Emphasis Type="Italic"> Anguilla japonica</Emphasis>: an aspect of the evolution of fish hatching enzyme gene

We isolated seven cDNA clones from embryos of the Japanese eel Anguilla japonica. Each deduced amino acid sequence consisted of a signal peptide, a propeptide and a mature enzyme portion belonging to the astacin protease family. A phylogenetic analysis showed that the eel enzymes resembled the high choriolytic enzyme (HCE) of medaka Oryzias latipes, and the hatching enzymes of the zebra fish Danio rerio and masu salmon Oncorhynchus masou. Hatching enzymes of these teleosts belonged to the group of the medaka HCE, and not the medaka low choriolytic enzyme (LCE), another hatching enzyme of medaka. Southern blot analysis showed that the genes of the eel hatching enzymes were multicopy genes like the medaka HCE genes. However, one of the eel hatching enzyme genes comprised eight exons and seven introns, and the exon-intron organization was similar to the medaka LCE gene, which is a single-copy gene. The molecular evolution of the fish hatching enzyme genes is discussed. In addition, whole-mount in situ hybridization and immunocytochemistry showed that the eel hatching enzyme was first expressed in the pillow anterior to the forebrain of early neurula, and finally in the cell mass on the yolk sac of later stage embryos. The early differentiation profile of eel hatching gland cells was similar to that of medaka, masu salmon and zebrafish, whereas the final location of the gland cells was different among fishes.Edited by N. Satoh     

Regularities of the life history strategy adoption in masu salmon Oncorhynchus masou from the Kol River (Western Kamchatka) in regard to the processes of growth and sexual maturation

Growth, age structure, sexual maturation, and peculiarities of the life history strategy adoption were studied for the population of masu salmon Oncorhynchus masou inhabiting the Kol River, Western Kamchatka. The growth rate and gametogenesis depended on the peculiarities of masu salmon generations in certain years and were the limiting factors for the juvenile differentiation and the direction of its ontogenetic development. The adoption of the resident or anadromous life history strategy in masu salmon is the epigenetic process, and the bifurcation during the life span of a single specimen is observed only once in a life, but twice in the generation, at the age of 1+ or 2+. The diversity of the life history strategy patterns was less at the northern boundary of the geographical range of masu salmon compared to the area of ecological optimum (the basin of the Sea of Japan). In the Kol River basin, nearby the northern boundary of its geographical range, the reproductive success and formation of the life history strategy diversity in the population depended strictly on the temperature regime of the water bodies, where the spawning and the freshwater period of life cycle of masu salmon take place. In Kamchatka, all the breeders were monocyclic, i.e. anadromous and resident precocious males spawned only once in a life.     

Responses of immature male masu salmon parr to the urine of mature males

Responses of male masu salmon Oncorhynchus masou parr to mature male urine were investigated in a Y-maze trough. Immature masu salmon made fewer entries into the channel conditioned by mature male urine than that with control water. This phenomenon may suggest that immature males avoid sexually active males in the spawning season.     

Size-structured Interactions between Native and Introduced Species: Can Intraguild Predation Facilitate Invasion by Stream Salmonids?

Dynamics of biological invasions may be complicated in size-structured animal populations. Differences in timing of life history events such as juvenile emergence create complex interaction webs where different life stages of native and non-native species act as predators, competitors, and prey. Stream salmonids are an ideal group for studying these phenomena because they display competition and predation in size-structured populations and have been introduced worldwide. For example, introduced rainbow trout (Oncorhynchus mykiss) are invading streams of Hokkaido Island, Japan and have caused declines in native masu salmon (O. masou) populations. However, age-0 rainbow trout emerge later than age-0 masu salmon and are smaller, which raises the question of why they are able to recruit and therefore invade in the face of a larger competitor. We conducted experiments in laboratory stream channels to test effects of increasing density of age-0 and age-1 rainbow trout on age-0 masu salmon. Age-1 rainbow trout dominated age-0 masu salmon by aggressive interference, relegating them to less favorable foraging positions downstream and reducing their foraging frequency and growth. The age-1 trout also reduced masu salmon survival by predation of about 40% of the individuals overall. In contrast, age-0 rainbow trout had little effect on age-0 masu salmon. Instead, the salmon dominated the age-0 trout by interference competition and reduced their survival by predation of 60% of the individuals. In each case, biotic interactions by the larger species on the smaller were strongly negative due to a combination of interspecific competition and intraguild predation. We predict that together these produce a positive indirect effect in the interaction chain that will allow the recruitment of rainbow trout in the face of competition and predation from age-0 masu salmon, and thereby facilitate their invasion in northern Japan.