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Directional cell movement during early development of the teleost Blennius pholis: I. Formation of epithelial cell clusters and their pattern and mechanism of movement.

Embryos of the teleost Blennius pholis provide exceptional material for observation of the formation and movement of cell clusters in vivo because the clusters are packed with melanosomes and migrate beneath the transparent enveloping layer. These clusters arise from two pigmented cell masses (PCM) which appear precociously on either side of the embryonic axis at 3/5 epiboly, at the future level of somites 1 and 2. As development proceeds, each PCM enlarges and spreads on its lateral margins to form an epithelial sheet. As spreading continues, the sheet fragments, forming small cell clusters that move in a distad direction in the yolk sac. The highly motile lateral marginal cells of the spreading PCM, as well as those of the marginal cells of each moving cluster, invariably protrude highly flattened lamellipodia, which terminate in long, fine, often branched filopodia. As cell clusters leave the PCM, they form long, taut retraction fibers. The rate of spreading of both the lateral edge of the PCM and the initial phase of cluster movement, is higher (1.0 micron/min or greater) than the later rate of cluster movement, apparently because at this phase, motile activity is confined to the distal borders of each. This directional migration ceases in 24 h at 16 degrees - 18 degrees C, when the farthest clusters have reached the ventral region of the yolk sac. By then, all clusters are spaced more or less evenly, apparently due to cessation of all cluster movement at about the same time. Once movement ceases, the clusters remain immobile for 2-4 days, depending on the temperature.

Animals↗

Teleost vision: seeing while growing.

Teleost fish eyes grow throughout life without compromising visual performance of the animal. This is made possible by a set of novel adaptations in the growth and development of the eye. Increased retinal area is achieved both by stretching the existing retina and by generation of new tissue at the retinal germinal zone at the margin of the eye. Rods are added in a fundamentally different fashion than are all other retinal cell types: they appear last as new retina is produced at the margin and they are inserted throughout the functional retina as it stretches. In this way, the animal maintains a constant rod density to preserve vision in low light level. Because the larger eye produces a larger image, visual acuity improves slightly as the animal grows. Adaptations responsible for regulation of retinal growth are analyzed and discussed.

Accommodation, Ocular↗

Identification of a stretch-activated monovalent cation channel from teleost urinary bladder cells.

The urinary bladder of euryhaline teleost is an important osmoregulatory organ which absorbs Na+, Cl-, and water from urine. Using patch clamp technique, single stretch-activated channels, which were permeable to K+ and Na+ (PNa/PK approximately 0.75) and had conductances of 55 and 116 pS, were studied. In excised, inside-out patches which were voltage-clamped in the physiological range of membrane potential, the single-channel open probability (Po) was low (approximately 0.02), and increased to a maximum of 0.9 with applied pipette suction. Single-channel conductance also increased with suction. The channels showed adaptation to applied suction and relaxed to a steady-state activity about 20 seconds after application of suction. The Po increased up to 0.9 with strong membrane depolarization (Vm = 0 to +80 mV); however, there was little dependence of Po on membrane potential in the physiological range. The kinetic data suggest that there is one conducting state and at least two non-conducting states of the channel. The open-time constant increased with suction but remained unchanged with membrane potential (Vm = -70 to +60 mV). The mean closed-time of the channel decreased with suction and membrane depolarization. These results demonstrate the presence of a non-selective monovalent cation channel which may be involved in cell volume regulation in the goby urinary bladder. Additionally, this channel may function as an enhancer of Na+ influx and K+ efflux across the bladder cell as part of transepithelial ion transport if it is located in apical membrane.

Animals↗

A quantitative autoradiographic study of 125I atrial natriuretic factor in the heart of a teleost fish (Conger conger).

Quantitative receptor autoradiographic study of 125I-atrial natriuretic peptide factor (ANF) in the heart of a teleost fish Conger conger has shown that a heterogenous distribution of 125I-ANF binding exists in the different cardiac regions. Elevated ANF binding densities (3,790 fmol/mg protein) were encountered in the innermost layer (tunica intima) of the bulbus arteriosus while lower binding levels (293-403 fmol/mg protein) were revealed in atrium and ventricle. In order to determine 125I-ANF binding characteristics (KD, Bmax) in the above cardiac sites, saturation binding assays were carried out. The results show that low 125I-ANF KD values (28.8-52.6 pM) were found in the atrium and in the bulbus arteriosus with respect to the higher KD values (373 pM) of the ventricle. The number of binding sites were respectively 632 and 1,279 fmol/mg protein for the atrium and the ventricle, while a substantially elevated Bmax of 7,235 fmol/mg protein was found for the bulbus arteriosus. These results may furnish some insights concerning ANF receptor binding activity and its putative regulatory role of different cardiac functions.

Animals↗

Immunohistochemical detection of P-glycoprotein in teleost tissues using mammalian polyclonal and monoclonal antibodies.

Mammalian P-glycoprotein is a highly conserved 170-kD integral plasma membrane protein functioning as an energy-dependent efflux pump of exogenous and endogenous lipophilic aromatic compounds entering the cell by diffusion. In this study, the tissue specificities of one polyclonal (pAb) and three monoclonal (mAbs) antibodies to mammalian P-glycoprotein were identified in paraffin-embedded, parasagittal whole-body sections of the guppy Poecilia reticulata. Polyclonal antibody mdr(Ab-1) and mAbs C219, C494, and JSB-1 demonstrated differential staining patterns in the following tissues: bile canaliculi in the liver, exocrine pancreas, lumenal surface of the intestinal epithelium, renal tubules, interrenal tissue, branchial blood vessels, gas gland, pseudobranch, and the gill transverse septa. Positive P-glycoprotein expression in P. reticulata correlates well with published results for homologous mammalian tissues of secretory and excretory function. These data indicate that one or more highly conserved members of the P-glycoprotein transporter family exist in a teleost species and can be detected using commercially available mammalian antibodies.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

Cortisol receptor blockade and seawater adaptation in the euryhaline teleost Fundulus heteroclitus.

To examine the role of cortisol in seawater osmoregulation in a euryhaline teleost, adult killifish were acclimated to brackish water (10 per thousand) and RU486 or vehicle was administered orally in peanut oil daily for five days at low (40 mg.kg(-1)) or high dose (200 mg.kg(-1)). Fish were transferred to 1.5 x seawater (45 per thousand) or to brackish water (control) and sampled at 24 h and 48 h after transfer, when Cl- secretion is upregulated. At 24 h, opercular membrane Cl- secretion rate, as Isc, was increased only in the high dose RU486 group. Stimulation of membranes by 3-isobutyl-1-methylxanthine and cAMP increased Isc in vehicle treated controls but those from RU486-treated animals were unchanged and membranes from brackish water animals showed a decrease in Isc. At 48 h, Isc increased and transepithelial resistance decreased in vehicle and RU486 groups, compared to brackish water controls. Plasma cortisol increased in all groups transferred to high salinity, compared to brackish water controls. RU486 treated animals had higher cortisol levels compared to vehicle controls. Vehicle treated controls had lower cortisol levels than untreated or RU486 treated animals, higher stimulation of Isc, and lower hematocrit at 24 h, beneficial effects attributed to increased caloric intake from the peanut oil vehicle. Chloride cell density was significantly increased in the high dose RU486 group at 48 hours, yet Isc was unchanged, suggesting a decrease in Cl- secretion per cell. Thus cortisol enhances NaCl secretion capacity in chloride cells, likely via glucocorticoid type receptors.

Adaptation, Physiological↗

Different somatostatin receptor subtypes are operating in the brain of the teleost fish, Coris julis.

Characterization of somatostatinergic (sst) neuronal activity through the application of nonpeptidyl agonists L-779,976 and L-817,818 which are highly specific for the sst receptors (sstr) sstr(2) and sstr(5), respectively, shows for the first time that sstr2, 5-like subtypes are the two major sstr subtypes operating in the brain of the teleost sea wrasse, Coris julis. A somewhat high but heterogeneous distribution pattern (> 30 < 180 fmol/mg wet tissue weight) of neurons expressing sstr2, 5 was reported in the different diencephalic regions plus in mesencephalon and telencephalon while low values were obtained in the cerebellum. Application of the above nonpeptidyl agonists permitted us to identify sstr2-like as the predominant subtype in telencephalic areas such as the entopeduncular nucleus (E) and postcommissural nucleus of the ventral telencephalon (Vp) as well as in hypothalamic and thalamic areas. At the same time high levels of neurons expressing sstr5-like, that greatly overlap those of sstr2-like in the diencephalic areas such as the anteroventral part of the preoptic nucleus (NPOav), the dorsal habenular nucleus (NHd) and the ventrolateral thalamic nucleus (VL), indicate that sstr2-like is very likely not the only sstr subtype acting in this fish brain. The predominance of sstr5-like in other brain areas is confirmed by the high quantities of this subtype in mesencephalic areas such as the torus longitudinalis (TLo). Overall, the discriminately differing densities of neurons expressing both subtypes seem to point to this system as a key molecular basis accounting for the distinct neurophysiological and behavioral sst-dependent activities in Coris julis.

Amides↗

Scaling effects on metabolism of a teleost.

Scaling effects on citrate synthase (CS), glucose-6-phosphate dehydrogenase (G6-PDH), RNA. RNA/DNA ratio and protein contents of brain, liver and skeletal muscle were studied in a teleost, Clarias batrachus. The activity of white skeletal muscle CS decreased significantly as a function of increasing body mass of the fish. It shows that the fulfilment of energy demand in white skeletal muscle is not dependent on aerobic metabolism. The activity of liver G6-PDH decreased with the increasing body mass showing reduction in NADPH generation for lipogenic activity. However, increase in G6-PDH activity showed enhancement in reductive synthesis in skeletal muscle of the larger-sized individuals. A positive scaling of RNA, RNA/DNA ratio and protein contents reflects changes in macromolecular turnover for ATP-supplying enzymes and proteins.

Analysis of Variance↗

Synaptic organization of the nucleus rotundus in some teleosts.

Synaptic organization of the nucleus rotundus was studied with the electron microscope in three teleost species belonging to the same order. In spite of the different histological organization (non-laminated, incompletely laminated, and laminated), the same kinds of axon terminals (S and F) are observed in all species. A fibrous layer which is clearly formed only in the laminated nucleus is composed of F1 terminals and dendrites from a layer of small cells. The same kind of synapses formed between F1 terminals and dendrites of small cells are also found among glomeruli in the non-laminated and incompletely laminated nuclei. The main constituents of glomeruli are S and F2 terminals and dendrites of large cells in the non-laminated and incompletely laminated nuclei, and are S terminals and star-like structures which correspond to the tips of the dendrites of large cells in the laminated nucleus. The star-like structure contains numerous mitochondria and clusters of small polymorphic vesicles. Some of the vesicles aggregate at thickened cell membranes of the structure as in presynaptic dendrites.

Animals↗

Structure and carbohydrate histochemistry of the stomach in eight species of teleosts.

The histology and carbohydrate histochemistry of eight teleostean stomachs are compared. Three gross anatomical types of stomachs are described and their shapes appear to correlate somewhat with feeding habits. Each type can be divided histologically into a corpus and pylorus. Gastric glands, containing only one cell type, occur in the copora of all species, but are present in the pylori of esocids only. As a single cell can produce both enzymes and hydrochloric acid such cells may be comparable to those of amphibians but not mammals. Lamina propria and submucosa are indistinctly separated in corpora but better defined in pylori by an intervening muscularis mucosa. The arrangement of the muscularis into inner circular and outer longitudinal layers is the opposite of that seen in the esophagus. Gastric mucous cells show species variations in localization of epithelial mucosubstances, which in broad terms are recognized as sulfomucins, sialomucins and neutral mucosubstances. A piscivorous diet does not appear to demand any particular type of carbohydrate. Within the Centrarchidae, gastric pit cells vary in carbohydrate content from only neutral mucosubstance to only weakly acidic sulfomucin; two species contain both types. A positive PAS reaction on the surface of gastric epithelial cells is suggestive of a striated border and thus possibly absorptive function. The absence of stomachs in some teleosts and the evolutionary and dietary significances are discussed.

Animals↗

Spatial organization in the saccule and lagena of a teleost: hair cell pattern and innervation.

The relationship between the hair cell orientation pattern and innervation in the saccule and lagena of the teleost Helostoma temmincki (the kissing gourami) was investigated with scanning electron microscopy and the Winkelmann-Schmitt silver impregnation technique. The hair cell pattern in the saccule consists of four orthogonally oriented groups. The anterior two groups are oriented along the animal's rostrocaudal axis, and the posterior two are oriented along its dorsoventral axis. The pattern of hair cell orientations in the lagena is a typical bidirectional one. Two divisions of the eighth nerve innervate the saccule. The anterior division innervates the horizontally oriented hair cell groups, and the posterior division innervates the dorsoventrally oriented groups. A single nerve innervates the lagena, with the majority of fibers innervating one or the other of the two lagenar hair cell groups. The segregated pattern of innervation according to hair cell orientation groups in the saccule was confirmed in other species. Individual types of axonal terminations appear to innervate hair cells of specific ciliary bundle types.

Animals↗

Horizontal cell processes in teleost retina.

Contacts between horizontal and bipolar cells are described in the retina of the teleost Eugerres plumieri. A single, long expansion observed in the external cone horizontal cells makes contact by means of a terminal button with the cell body of a bipolar. It represents the only contact between this class of horizontal cell and the bipolar soma. On the other hand, the medial and internal cone horizontal cells and the rod horizontal cells, which lack such a single, long expansion, display instead numerous short and fine expansions that terminate by means of a terminal knob on a bipolar cell body. The bipolar-destined, short expansions of the rod horizontal cell make contact with large bipolar cell bodies, whereas corresponding short expansions of cone horizontal cells contact small bipolar cell bodies. It is suggested that the ascending horizontal cell process forms presynaptic terminals in the photoreceptor triad complex, and that the single, long and the multiple, short bipolar-destined expansions are postsynaptic to the bipolar cell body.

Animals↗

Microtubules and actin filaments in teleost visual cone elongation and contraction.

Teleost retinal cones contract in light and elongate in darkness. This paper describes the disposition of microtubules and cytoplasmic filaments in cone cells of 2 species of fish (Haemulon sciurus and Lutjanus griseus). In Haemulon, the neck-like "myoid" region of the cone changes in length from 5 mu to 75 mu. Maximal observed rates of elongation and contraction are comparable to that of chromosome movement in mitosis (2-3 mu/min). Microtubules presumably participate in cone elongation, since numerous longitudinal microtubules are present in the myoid region, and colchicine blocks dark-induced elongation. Myoid shortening, on the other hand, appears to be an active contractile process. Disruption of microtubules in dark-adapted cones does not produce myoid shortening in the absence of light, and light-induced myoid shortening is blocked by cytochalasin-B. Cone cells possess longitudinally-oriented thin filaments which bind myosin subfragment-1 to form arrowhead complexes typical of muscle actin. Myoid thin filaments are clearly observed in negatively stained preparations of isolated cones which have been disrupted with detergent after attachment to grids. These myoid filaments are not, however, generally preserved by conventional fixation, though bundles of thin filaments are preserved in other regions of the cell. Thus, actin filaments are poorly retained by fixation in precisely the region of the cone cell where contraction occurs. Cone cells also possess longitudinally-oriented thick filaments 130-160 A in diameter. That these thick filaments may be myosin is suggested by the presence of side-arms with approximately 150 A periodicity. The linear organization of the contractile apparatus of the retinal cone cell makes this cell a promising model for morphological characterization of the disposition of actin and myosin filaments during contraction in a nonmuscle cell.

Actins↗

A novel seminal plasma glycoprotein of a teleost, the Nile tilapia (Oreochromis niloticus), contains a partial von Willebrand factor type D domain and a zona pellucida-like domain.

Our previous study shows that seminal plasma of a teleost, the Nile tilapia, contains a glycoprotein Mr = 120,000 named as SPP (Seminal plasma glycoprotein)120 which forms a homopolymer that has sperm immobilizing activity. In order to elucidate the mechanisms of the formation of the homopolymer and the immobilization of sperm, molecular cloning of SPP120 was conducted. The cDNA for SPP120 contains a complete open reading frame encoding 797 amino acid residues with 14 potential N-glycosylation sites. The predicted amino acid sequence of SPP120 contains a partial von Willebrand factor type D domain and a zona pellucida domain, that are involved in protein-protein adhesion that form filamentous structures in various kinds of cells. This result suggests that SPP120 forms a homopolymer via these domains in seminal plasma and probably interacts with spermatozoa. Northern blotting reveals that the gene is also expressed in ovary, even in ovulated eggs. The results of in situ hybridization indicate that in testis the gene is expressed in Sertoli cells and epithelial cells of sperm ducts, and the localization corresponds to that of the protein analyzed by immunohistochemistry. In the ovary, the gene is expressed at the perinucleolus stage of oocytes; however, the protein is not detected in any cells other than oocytes.

Amino Acid Sequence↗

Sox9 in a teleost fish, medaka (Oryzias latipes): evidence for diversified function of Sox9 in gonad differentiation.

Sox9 is a transcription factor containing the Sry-related high-mobility-group (HMG) box. Mutations in human SOX9 gene cause skeletal defects and male-to-female sex reversal, indicating its essential roles in chondrogenesis and testis development. Comparative studies have shown that Sox9 is expressed in chondrogenic tissues and testis in other vertebrates. Therefore, it was suggested that roles of Sox9 in cartilage and male gonad development are conserved among vertebrates. To investigate the evolutional significance of Sox9 in the gonad and cartilage development of teleost fish, we isolated medaka sox9 and analyzed its expression. Two kinds of transcripts (sox9 and sox9lf) were isolated by cDNA library screening. The sox9 encoded 487 amino acids and showed approximately 70% amino acid identity with known vertebrate SOX9 proteins. The sox9lf was a longer form of the sox9, which was transcribed from an additional exon in the 5' upstream region. Interestingly, the expression of medaka sox9 was predominantly observed in the adult ovary by northern blot and in situ hybridization analyses, whereas in the testis, its expression was detectable only by RT-PCR. During medaka embryogenesis, its expression was observed in the cranial cartilage and pectoral fin endoskeleton. These observations suggest that the function of Sox9 in the cartilage is conserved among vertebrates, while that in the gonad is quite different in medaka.

Amino Acid Sequence↗

Role of substrate on larval development of the freshwater teleost Pelvicachromis pulcher.

It is known that, in an aquatic environment, the mineralogical composition of the substrate can affect the structure of settled communities. In marine environments, the presence of quartz negatively influences the formation of biofilm, as well as the selection and the colonization of the substrate by benthic organisms. Direct laboratory observation revealed that the freshwater teleost Pelivicachromis pulcher selects, when available, nonquartzitic brooding substrate. To monitor the effects of substrate on larvae development, ten lots of embryos were distributed in grid nurseries; carbonatic gravel was laid in five of the nurseries, while freshly fractured quartz gravel was used in the remaining ones. All the embryos laid in the two nurseries hatched, and 90% of the carbonate developing larvae reached adulthood, while 100% of those reared on quartz grain died 120 hr post hatching. Examination was made, both in larvae developed on carbonatic substrates and in those developed on quartz substrates, of the expression of the fetal growth factor, the insulin growth factor-II (IGF-II), of the molecular chaperone, the heat shock protein 70 (HSP70), which is involved in the folding of the nascent polypeptide chain, of the key enzyme of the glycolytic pathway, the glyceraldehyde-3-phosphate dehydrogenase (GADPH), and of the housekeeping gene, the beta-actin. All the data were normalized against 18S RNA expression. In larvae reared on quartz substrate, the genes IGF-II and the beta-actin showed a lower expression, while the GADPH was totally suppressed and the expression of HSP70 increased. In conclusion, the data presented in this article demonstrated, for the first time, that the presence of quarzitic substrates is sufficient to stop larvae development through the inhibition of gene transcription in this African cichlid, leading to its death.

Actins↗

The effects of ciguatoxin on the nerves of the teleost fish, Sillago ciliata.

The absolute refractory period, relative refractory period, and the duration and magnitude of the supernormal period were measured after incubation of fish nerves with ciguatoxin and other channel modifying compounds, tetrodotoxin, veratridine, verapamil, and lignocaine. In vitro electrophysiological studies were carried out on the lateral line nerve of the whiting, Sillago ciliata Cuvier. Electrophysiological changes in fish nerves after exposure to ciguatoxin (0.3 MU.ml-1) and veratridine (1 x 10(-5) M) are similar to changes that occur in mammalian nerves and include an increase in the absolute refractory period, the relative refractory period, and the magnitude and duration of supernormality. The effects of ciguatoxin (0.3 MU.ml-1) in fish nerves were antagonised by tetrodotoxin (5 x 10(-10) M), verapamil (5 x 10(-7) M), and lignocaine (1 x 10(-5) g/ml-1). The nerves of Sillago ciliata used in this study responded to ciguatoxin and its antagonists in a similar manner to mammalian nerves, suggesting that these teleost nerves have no specific electrophysiological mechanism to cope with this toxin.

Animals↗