SOME OBSERVATIONS ON THE FINE STRUCTURE OF THE LATERAL LINE ORGAN OF THE JAPANESE SEA EEL LYNCOZYMBA NYSTROMI.
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The fine structure of the lateral line organ of the Japanese sea eel Lyncozymba nystromi has been studied with the electron microscope. The sensory epithelium of the lateral line organ consists of a cluster of two major types of cells, the sensory hair cells and the supporting cells. The sensory cell is a slender element with a flat upper surface provided with sensory hairs, Two different types of synapses are distinguished on the basal surface of the receptor cell. The first type is an ending without vesicles and the second type is an ending with many vesicles. These are presumed to correspond to the afferent and the efferent innervations of the lateral line organ. The fine structure of the supporting cells and the morphological relationship between the supporting cells and the receptor cells were observed. The possible functions of the supporting cells are as follows: (a) mechanical and metabolic support for the receptor cell; (b) isolation of the individual receptor cell; (c) mucous secretion and probably cupula formation; (d) glial function for the intraepithelial nerve fibers. Both myelinated and unmyelinated fibers were found in the lateral line nerve. The mode of penetration of these fibers into the epithelium was observed. (+info)
THE ADENOSINE-TRIPHOSPHATASE SYSTEM RESPONSIBLE FOR CATION TRANSPORT IN ELECTRIC ORGAN: EXCLUSION OF PHOSPHOLIPIDS AS INTERMEDIATES.
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1. Subcellular fractions were prepared from the electric organs of Electrophorus and Torpedo and assayed for adenosine-triphosphatase activity. 2. Treatment of the ;low-speed' fraction from Torpedo with m-urea gave an adenosine-triphosphatase preparation that was almost completely (98%) inhibited by ouabain (0.1mg./ml.) and dependent on the simultaneous presence of Na(+) and K(+). 3. The adenosine-triphosphatase preparations were exposed to [gamma-(32)P]ATP for 30sec. in the presence of (i) Na(+), (ii) K(+), (iii) Na(+)+K(+) and (iv) Na(+)+K(+)+ouabain. No significant labelling of phosphatidic acid, triphosphoinositide or any other phospholipid was observed. 4. The results suggest that phospholipids do not act as phosphorylated intermediates in the ;transport adenosine-triphosphatase' system of electric organ. (+info)
Interrenal steroid 21-hydroxylase in eels: primary structure, progesterone-specific activity and enhanced expression by ACTH.
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Cytochrome P450 21-hydroxylase (P450c21) is a key enzyme for corticosteroidogenesis. To understand the regulatory mechanisms of cortisol production in fish, we have cloned a cDNA encoding P450c21, for the first time in non-mammalian vertebrates, from the head kidney of the eel (Anguilla japonica). The overall similarity of the deduced P450c21 sequence was modest (41-44% amino acid identity) between the eel and mammals. However, the functional domains for steroid-binding, heme-binding and proton-transfer sites were well conserved (74-100% identity). The eel P450c21 mRNA was expressed abundantly in the anterior quarter of the head kidney, but was undetectable in the remaining three-quarters or in other tissues including the gill, heart, liver, intestine, kidney, immature gonad and skeletal muscle. Functional expression of the cDNA clone in non-steroidogenic COS-1 cells produced a protein with high 21-hydroxylase activity to convert progesterone to 11-deoxycortisterone but not 17alpha-hydroxyprogesterone to 11-deoxycortisol, although the latter is a preferred substrate for mammalian P450c21. To examine whether 21-hydroxylated progesterone is actually 17alpha-hydroxylated in the eel interrenal, 11-deoxycorticosterone and (3)H-corticosterone were respectively incubated with the interrenal-containing anterior quarter of the head kidney. The separation of the steroids produced by two HPLC systems revealed that cortisol was produced from both substrates, showing the 17alpha-hydroxylation of 11-deoxycorticosterone and corticosterone in the eel interrenal. ACTH infused at 3 pmol/kg per min for 5 h consistently increased plasma cortisol levels and interrenal P450c21 mRNA levels in seawater eels. These results showed that the interrenal-specific eel P450c21 cloned in this study is involved in cortisol production through conversion of progesterone to 11-deoxycorticosterone in the interrenal-containing anterior quarter of eel head kidney. Furthermore, ACTH stimulates cortisol production in part through enhanced P450c21 expression in the eel interrenal. (+info)
Integrated responses of Na+/HCO3- cotransporters and V-type H+-ATPases in the fish gill and kidney during respiratory acidosis.
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Using degenerate primers, followed by 3' and 5' RACE and "long" PCR, a continuous 4050-bp cDNA was obtained and sequenced from rainbow trout (Oncorhynchus mykiss) gill. The cDNA included an open reading frame encoding a deduced protein of 1088 amino acids. A BLAST search of the GenBank protein database demonstrated that the trout gene shared high sequence similarity with several vertebrate Na(+)/HCO(3)(-) cotransporters (NBCs) and in particular, NBC1. Protein alignment revealed that the trout NBC is >80% identical to vertebrate NBC1s and phylogenetic analysis provided additional evidence that the trout NBC is indeed a homolog of NBC1. Using the same degenerate primers, a partial cDNA (404 bp) for NBC was obtained from eel (Anguilla rostrata) kidney. Analysis of the tissue distribution of trout NBC, as determined by Northern blot analysis and real-time PCR, indicated high transcript levels in several absorptive/secretory epithelia including gill, kidney and intestine and significant levels in liver. NBC mRNA was undetectable in eel gill by real-time PCR. In trout, the levels of gill NBC1 mRNA were increased markedly during respiratory acidosis induced by exposure to hypercarbia; this response was accompanied by a transient increase in branchial V-type H(+)-ATPase mRNA levels. Assuming that the branchial NBC1 is localised to basolateral membranes of gill cells and operates in the influx mode (HCO(3)(-) and Na(+) entry into the cell), it would appear that in trout, the expression of branchial NBC1 is transcriptionally regulated to match the requirements of gill pHi regulation rather than to match trans-epithelial HCO(3)(-) efflux requirements for systemic acid-base balance. By analogy with mammalian systems, NBC1 in the kidney probably plays a role in the tubular reabsorption of both Na(+) and HCO(3)(-). During periods of respiratory acidosis, levels of renal NBC1 mRNA increased (after a transient reduction) in both trout and eel, presumably to increase HCO(3)(-) reabsorption. This strategy, when coupled with increased urinary acidification associated with increased vacuolar H(+)-ATPase activity, ensures that HCO(3)(-) levels accumulate in the body fluids to restore pH. (+info)
Multiplex PCR assay for detection of bacterial pathogens associated with warm-water Streptococcosis in fish.
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A multiplex PCR-based method was designed for the simultaneous detection of the main pathogens involved in warm-water streptococcosis in fish (Streptococcus iniae, Streptococcus difficilis, Streptococcus parauberis, and Lactococcus garvieae). Each of the four pairs of oligonucleotide primers exclusively amplified the targeted gene of the specific microorganism. The sensitivity of the multiplex PCR using purified DNA was 25 pg for S. iniae, 12.5 pg for S. difficilis, 50 pg for S. parauberis, and 30 pg for L. garvieae. The multiplex PCR assay was useful for the specific detection of the four species of bacteria not only in pure culture but also in inoculated fish tissue homogenates and naturally infected fish. Therefore, this method could be a useful alternative to the culture-based method for the routine diagnosis of warm-water streptococcal infections in fish. (+info)
Dopamine inhibits luteinizing hormone synthesis and release in the juvenile European eel: a neuroendocrine lock for the onset of puberty.
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In various adult teleost fishes, LH ovulatory peak is under a dual neurohormonal control that is stimulatory by GnRH and inhibitory by dopamine (DA). We investigated whether DA could also be involved in the inhibitory control of LH at earlier steps of gametogenesis by studying the model of the European eel, Anguilla anguilla, which remains at a prepubertal stage until the oceanic reproductive migration. According to a protocol previously developed in the striped bass, eels received sustained treatments with GnRH agonist (GnRHa), DA-receptor antagonist (pimozide), and testosterone (T) either alone or in combination. Only the triple treatment with T, GnRHa, and pimozide could trigger dramatic increases in LH synthesis and release as well as in plasma vitellogenin levels and a stimulation of ovarian vitellogenesis. Thus, in the prepubertal eel, removal of DA inhibition is required for triggering GnRH-stimulated LH synthesis and release as well as ovarian development. To locate the anatomical support for DA inhibition, the distribution of tyrosine hydroxylase (TH) in the brain and pituitary was studied by immunocytochemistry. Numerous TH-immunoreactive cell bodies were observed in the preoptic anteroventral nucleus, with a dense tract of immunoreactive fibers reaching the pituitary proximal pars distalis, where the gonadotrophs are located. This pathway corresponds to that mediating the inhibition of LH and ovulation in adult teleosts. To our knowledge, this is the first demonstration of a pivotal role for DA in the control of LH and puberty in a juvenile teleost. These data support the view that DA inhibition on LH secretion is an ancient evolutionary component in the neuroendocrine regulation of reproduction that may have been partially maintained throughout vertebrate evolution. (+info)
Solution structure of an RNA stem-loop derived from the 3' conserved region of eel LINE UnaL2.
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The eel long interspersed element (LINE) UnaL2 and its partner short interspersed element (SINE) share a conserved 3' tail containing a stem-loop that is critical for their retrotransposition. Presumably, the first step of retrotransposition is the recognition of their 3' tails by UnaL2-encoded reverse transcriptase. The solution structure of a 17-nucleotide RNA derived from the 3' tail of UnaL2 was determined by NMR. The GGAUA loop forms a specific structure in which the uridine is exposed to solvent with the third and fifth adenosines stacked. A sharp turn in the phosphodiester backbone occurs between the second guanosine and third adenosine. When the uridine is mutated (but not deleted), all mutants form the loop structure, indicating that the loop structure requires an exposed fourth residue. The retrotransposition assay in HeLa cells revealed that retrotransposition requires the second guanosine, although any nucleoside functions at the fourth position, suggesting that UnaL2 reverse transcriptase specifically recognizes the 5' side of the GGANA loop. (+info)
Pathogenic Aeromonas hydrophila serogroup O:14 and O:81 strains with an S layer.
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Five autoagglutinating Aeromonas hydrophila isolates recovered from eels and humans were assigned to serogroups O:14 and O:81 of the Sakazaki and Shimada (National Institutes of Health) scheme. They had the following properties in common: positive precipitation after boiling, moderate surface hydrophobicity (salt-aggregation-test value around 1.2), pathogenicity for fish and mice (50% lethal dose, 10(4.61) to 10(7.11)), lipopolysaccharides that contained O-polysaccharide chains of homogeneous chain length, and an external S layer peripheral to the cell wall observed by electron microscopy. A strong cross-reactivity was detected by immunoblotting between the homogeneous O-polysaccharide fraction of O:14 and O:81 strains but not between them and the lipopolysaccharide of A. hydrophila TF7 (O:11 reference strain). Outer membrane fractions of these strains contained a predominant 53- to 54-kDa protein which was glycine extractable under low-pH (pH 2.8) conditions and was identified as the surface array protein. The S-layer proteins of the O:14 and O:81 A. hydrophila strains seemed to be primarily different from those previously purified from strains A. hydrophila TF7 and Aeromonas salmonicida A450 on the basis of colony hybridizations with both the structural genes vapA and ahsA. This is the first report of the presence of an S layer in mesophilic Aeromonas strains not belonging to serogroup O:11. (+info)