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[Studies on the morphology of blood of the European eel (Anguilla agnuilla). III. Studies on monocytic and lymphoid cells].

Investigations concerning the monocytic and lymphocytic cell series were carried out on the peripheral blood and dabbings of the kidney, the spleen and intestines of healthy eels (anguilla anguilla), and eels taken ill with vibrio anguillarum. By means of histological, cytochemical, phaseoptical, and electronmicroscopical examinations independent development lines of the monocytes and lymphocytes were established. The place of origination for the monocytes was found to be the kidney, for the lymphocytes the spleen and the intestines additionally. The evidence of the cytochemical assay of enzymes for the differentiation of these blood cells and their cellular systems is primarily dependent on the distribution of the activity of the unspecific esterase, the naphthol-AS-D-chloroacetate-esterase, the peroxidase and the alkalic phosphatase. Macrophages were - on the basis of enzymocytochemical results - seen as derivatives of the monocytes. Plasma cells and lymphoid cells are transformation forms of the lymphocytes. Our results emphasize the significance especially of cytochemistry for the hematological diagnostics of fish diseases.

Animals

[Structural modifications of the esophagus linked to osmoregulation in eels].

In comparison with the fresh-water Eel (Anguilla anguilla L.), the oesophagus of the sea-water Eel displays an almost complete regression of the multilayered epithelium and consequently the disappearance of mucous cells. In contrast, a large development of a columnaŕ epithelium is associated with a tremendous proliferation of the subepithelial vascularization. These modifications enable one to interpret the changes in ionic permeability of the oesophagus during transfer from freshwater to seawater.

Animals

Pancreatectomy in a teleost fish, Anguilla rostrata (American eel).

A technique for pancreatectomy was described for the American eel. In this fish, the operation was not followed by the typical signs of diabetes mellitus. Histodensitometric determination of liver glycogen showed no major differences between operated controls and pancreatectomized eels. The absence of a specific hyperglycemia, previously reported with the hexokinase method, was confirmed with the glucose oxidase technique; however, the more specific hexokinase method gave consistently lower values. Breakdown of adipose tissue and hyperlipemia were absent.

Animals

Relative estimation of ventilatory fow rate in the eel (Anguilla anguilla L.)

Freshwater eels (Anguilla anguilla L.) were experimented on in order to measure ventilatory flow rate (by means of a direct method of collecting expired water), and simultaneously, to record sub-opercular pressure changes and opercular displacements (operculogram). Significant correlations (p less than 0.01) were found between the ventilatory flow rate (V) and various characteristics of operculiogram records. The relationships which are significant in each fish experimented on are; V = a(f.lo) + b; V = a(f.Ao) % b and V = a(f.Ao2) + b(f: ventilatory frequency: Ao and lo: mean amplitude and result of integration of opercular tracings). Considering the technical difficulties in measuring the ventilation flow rate of the eel, the easier operculographic recording methods lead to a correct estimation of relative changes in ventilatory flow rate, by using the refined relationship: V = a(Ao1.63).

Animals

Long term effect of hypophysectomy and prolactin treatment on kidney structure in the eel in fresh water.

Long term effects of hypophysectomy and ovine prolactin treatment on the histological picture of the kidney were investigated in European eels. The epithelial cell height is very slowly affected after hypophysectomy, the decrease being highly significant after 50 days only in the distal and the initial collecting tubules. Nuclear surfaces were more rapidly reduced, the regressive changes being first detected in the initial collecting tubule (5 days), then in the distal portion and the two proximal segments. No degenerative changes were observed. These structural responses may be correlated with the slow reduction of the urine flow and of the plasma electrolytes in the same species. Ovine prolactin treatment (10mug/g/day) overcompensates the effects of hypophysectomy on the epithelial cell height and the nuclear area, and even after 50 days still induces the proliferation of new tubules although its action remains less intense than in intact eels. Other histological criteria (brush border thickness, mitotic activity, basal structure of proximal 2 cells) and sodium retention do suggest that these structural modifications reflect an increased functional activity of the kidney in addition to the effect of prolactin on sodium fluxes in the gills.

Anguilla

Serotonin and MSH secretion: effect of parachlorophenylalanine on the pituitary cytology of the eel.

Parachlorophenylalanine (pCPA), an inhibitor of tryptophan hydroxylase which depletes brain serotonin in higher vertebrates, was injected into freshwater eels. After 4 or 6 injections (200 mg/kg/day) or 10 injections (100 and 140 mg/kg/day), the animals are paler, with a low melanophore index. In the pituitary gland, granules tend to accumulate in the basal part of the MSH cells and in the perinuclear area. Cells appear smaller with a decreased nuclear area (P less than 0.001). In the neurohypophysis, the amount of neurosecretory material is often reduced. Conversely, injections of 5-hydroxytryptophan induce a strong darkening, a result similar to that previously reported in some amphibian species and in one lacertilian species. These data substantiate the hypothesis of a stimulatory influence of 5-hydroxytryptamine on MSH release and possibly its synthesis in the eel and other lower vertebrates.

5-Hydroxytryptophan

Morphochemical analysis of mucosubstances in some epithelial tissues of the eel (Anguilla japonica).

In attempts to gain insight into the nature of mucosubstances in the eel epithelial tissues, the epidermis and epithelia lining the gill and intestine of the fish have been reacted for a series of light microscopic stainings of mucosaccharide and protein histochemistry under normal and experimental conditions. The present morphochemical analyses indicate that (1) the mucosubstances of the epidermis are found within two well differentiated cell types, goblet and clavate cells which elaborate a neuraminic acid containing mucosaccharide with vicinal hydroxyl, sulfate and carboxyl groupings and a glycoprotein respectively and (2) the mucosubstances of the gill and intestinal epithelia are found within goblet cells which elaborate a mucosaccharide with histochemical properties comparable to those of mucosaccharide within the epidermal goblet cells. The histochemical characteristics of all these mucosubstances have been briefly discussed in relation to the activities of the epithelial tissues in the eel.

Anguilla

HCO-3-activated adenosine triphosphatase in intestinal mucosa of the eel.

An HCO-3-activated and SCN--inhibited ATPase (ATP phosphohydrolase, EC 3.6.1.3) found in homogenates of intestinal mucosa of the eel was solubilized by Triton X-100. Optimal HCO-3-concentration and pH for the enzyme were 25 mM and 8.7, respectively. HCO-3-ATPase activity in both homogenate and solubilized preparations increased after seawater adaptation. This adaptive increase in enzyme activity was also observed in the gills and the kidney. The HCO-3-ATPase seems to be related to transport mechanisms, especially for Cl-, in osmoregulatory surfaces of the eel.

Adenosine Triphosphatases

Detergent inactivation of sodium- and potassium-activated adenosinetriphosphatase of the electric eel.

The stability of the sodium- and potassium-activated adenosinetriphosphatase (Na,K-ATPase) of the electric eel, Electrophorus electricus, was studied in five detergents in an effort to establish conditions for reconstitution of this membrane protein into defined phospholipids. The Na,K-ATPase activity of purified electric organ membranes as well as the ATPase is stable for at least 1 month of storage at 0 degrees C in the absence of detergents. At low concentrations of detergents, the enzyme is also stable for several days, but irreversible inactivation occurs rapidly as the detergent concentration is further increased. This inactivation begins at well-defined threshold concentrations for each detergent, and these concentrations generally occur in the order of the detergent critical micelle concentrations. Increasing the concentration of the electric organ membranes causes a linear increase in the inactivation threshold concentrations of Lubrol WX, deoxycholate, and cholate. The onset of inactivation evidently occurs when the mole fraction of detergent associated with the membrane lipids reaches a critical value in the narrow range of 0.2-0.4, in contrast to the large differences in the bulk concentrations of these detergents. The eel Na,K-ATPase is more sensitive to detergents than the sheep kidney enzyme.

Animals

Active-site determinations on forms of mammalian brain and eel acetylcholinesterase.

Three forms of brain acetylcholinesterase were purified from bovine caudate-nucleus tissue and determined by calibrated gel filtration to have mol.wts. of approx. 120 000 (C), 230 000 (B) and 330 000 (A). [3H]Di-isopropyl phosphorofluoridate (isopropyl moiety labelled) was purified from commercial preparations and its concentration estimated by an enzyme-titration procedure. Brain acetylcholinesterase preparations and enzyme from eel electric tissue were allowed to react with [3H]di-isopropyl phosphorofluridate in phosphate buffer until enzyme activity was inhibited by 98%. Excess of [3H]di-isopropyl phosphorofluoridate that had not reacted was separated from the labelled enzyme protein by gel filtration, or by vacuum filtration or by extensive dialysis. The specificity of active-site labelling was confirmed by use of the enzyme reactivator, pyridine 2-aldoxime. The forms of brain acetylcholinesterase were calculted to contain approximately two (C) four (B) and six (A) active sites per molecule respectively. Acetylcholinesterase (mol.wt. 250 000) from electric-eel tissue was estimated to contain two active sites per molecule. Gradient-gel electrophoresis was used to confirm the estimation of molecular weights of brain acetylcholinesterase forms made by gel filtration. Under the conditions of electrophoresis acetylcholinesterase form A was stable, but form B was converted into a species of approx. 120 000 mol. wt. Similarly, form C of the brain enzyme was converted into a 60 000-mol.wt. form during electrophoresis. These results are in general accord with the suggestion that the multiple forms of brain acetylcholinesterase may be related to the aggregation of a single low-molecular-weight species.

Acetylcholinesterase

Gustatory responses of eel palatine receptors to amino acids and carboxylic acids.

The gustatory receptors of the eel palate were found to be extremely sensitive to amino acids and carboxylic acids. The results obtained are as follows: (a) 11 amino acids which are among naturally occurring amino acids elicited responses in the palatine nerve, but 9 amino acids did not elicit a response even at a high concentration. The effect of D-amino acids was always much less than that of their corresponding L-isomers. There was no appreciable difference in the effectiveness of an alpha-amino acid (alpha-alanine) and beta-amino acid (beta-alanine). (b) The threshold concentrations of the most potent amino acids (arginine, glycine) were between 10(-8) and 10(-9) M. A linear relation between the magnitude of the response and log stimulus concentration held for a wide concentration range for all the amino acids examined. (c) The palatine receptors responded sensitively to various carboxylic acid solutions whose pH was adjusted to neutral. The threshold concentrations varied between 10(-4) and 10(-7) M. The magnitude of the response at 10(-2) M increased with an increase of carbon chain length. (d) The extent of cross-adaptation was examined with various combinations of amino acids. A variety of the response patterns showing complete cross-adaptation, no cross-adaptation, or synergetic interaction was observed. The synergetic interaction was also observed when one amino acid below its threshold concentration was added to the other amino acid below its threshold concentration was added to the other amino acid. No cross-adaptation was observed between amino acids and fatty acids. (e) The treatment of the palate with papain led to loss of the responses to arginine, glycine, and histidine without affecting those to proline and acetic acid. The treatment with pronase E eliminated selectively the response to proline. The possibility that the eel gustatory receptors are responsible for sensing food at a distance was discussed.

Adaptation, Physiological

[Ultrastructure of gonadotropic cells of the normal eel and after estradiol injection].

Male and female silver eels, in fresh water or sea water, have few gonadotrophs (GTH) poorly differentiated. An oestradiol (E2) treatment induces their development: a well developed Golgi area, abundant secretory granules (200-500 nm), dilated cisterna and large globules 1.2-2.2 micron) are observed. A positive feedback may control gonadotropin synthesis in immature eels. No macroscopic effect is discernible on the gonad.

Anguilla

A study of the fine structure of the pit organ of the common Japanese sea eel Conger myriaster.

The fine structure of the pit organ of the sea eel has been studied by means of electron microscopy. The sensory epithelium of the pit organ consists of sensory cells and supporting cells. The apical surface of the sensory cell is studded with sensory hairs consisting of a kinocilium and stereocilia. The sensory cells are divided into two groups. In one, the kinocilium points dorsally and in the other the kinocilium points ventrally. The total number of sensory cells in one pit organ is about 100, and the ratio of cells with opposite polarity is about 1:1. On the basis of these structural features, the pit organ is considered to be a mechano-receptor sensitive to the movement of liquid in a dorso-ventral direction. It may also serve as an ion receptor, sensitive to environmental ion concentration. Efferent nerve terminals make rare synaptic contacts on the afferent nerve fiber.

Animals

Interactions of lectins with (Na+ + K+)-ATPase of eel electric organ.

Interaction of lectins with a detergent-solubilized ATPase from eel electric organ was studied. Concanavalin A, which binds to alpha-mannosides, altered the rate of enzyme migration in agar and inhibited the formation of an antigen-antibody precipitate: other lectins had no such effects. Concanavalin A similar amounts partially inhibited (Na+ + K+)-ATPase; this inhibition was reversible by alpha-methylglucoside. There was no corresponding effect of concanavalin A on the potassium p-nitrophenylphosphatase. Concanavalin A also did not interfere with ouabain binding. Thus, concanavalin A binds to an antigenic region also involved in Na+ and/or ATP binding, but does not interact with a K+ site.

4-Nitrophenylphosphatase

High affinity calcium binding peptides from the gills, gut and kidneys of the freshwater eel (Anguilla anguilla).

We report the isolation of high affinity calcium-binding peptides from homogenates of gills, gut and kidneys of the freshwater eel Anguilla anguilla. These peptides show identical gel-filtration profiles on Biogel P-6, from which a molecular weight of 3500 was estimated. The average binding constant (Kf) for calcium is 7 . 10(7)M-1, and MgCl2 up to 5 mM does not displace calcium.

Animals

Effect of acetylcholine on postjunctional membrane permeability in eel electroplaque.

Acetylcholine (ACh) was applied iontophoretically to the innervated face of isolated eel electroplaques while the membrane potential was being recorded intracellularly. At the resting potential (about -85 mV) application of the drug produced depolarizations (ACh potentials) of 20 mV or more which became smaller when the membrane was depolarized and reversed in polarity at about zero membrane potential. The reversal potential shifted in the negative direction when external Na+ was partially replaced by glucosamine. Increasing external K+ caused a shift of reversal potential in the positive direction. It was concluded that ACh increased the permeability of the postjunctional membrane to both ions. Replacement of Cl- by propionate had no effect on the reversal potential. In Na+-free solution containing glucosamine the reversal potential was positive to the resting potential, suggesting that ACh increased the permeability to glucosamine. Addition of Ca++ resulted in a still more positive reversal potential, indicating an increased permeability to Ca++ as well. Analysis of the results indicated that the increases in permeability of the postjunctional membrane to K+, Na+, Ca++, and glucosamine were in the ratios of approximately 1.0:0.9:0.7:0.2, respectively. With these permeability ratios, all of the observed shifts in reversal potential with changes in external ionic composition were predicted accurately by the constant field equation.

Acetylcholine

The eel retina. Ganglion cell classes and spatial mechanisms.

We have been able to separate optic fibers in the eye of the eel Anguilla rostrata into two distinct classes on the basis of spatial summation properties. X fibers, the first class, are like X ganglion cells in the cat: they have null positions for contrast reversal sine gratings; they respond at the modulation frequency; and many have a strong surround mechanism. X fibers, the second class, respond with an "on-off" response to local stimulation, to diffuse light modulation, to coarse drifting gratings, and to contrast reversal gratings. We have put forward a model for the receptive field of X fibers which involves two subunits, with rectification before the subunits add their signals. This model accounts for many of the quirks of X fibers.

Animals

The effect of serotonin and transcapillary pressure on the capillary permeability in the rete mirabile of the eel (Anguilla vulgaris l.).

Measurements of transcapillary exchange of high-molecular dextrans, K+ and THO in the rete mirabile of eel (Anguilla vulgaris L.) have been made under steady-state conditions while the transcapillary pressure was varied and pharmaca added to the perfusates. Raising the transcapillary input pressure increased the exchange of dextrans and K+ but lowered that of THO. Histamine and ouabain had no effect on the transcapillary exchange of the test molecules. The highest dose (100 mug/ml) of serotonin reversibly increased the capillary permeability of D-75 000, K+ and THO while the medium dose (50 mug/ml) only raised the transcapillary exchange of K+ and THO. The lowest dose (10 mug/ml) had no permeability effect. Our main conclusion is that serotonin reversibly increases the area of the intercellular pores by affecting the capillary wall directly.

Air Sacs