Oral absorption of biologically active insulin in common carp (Cyprinus carpio L.).

1. Bovine insulin dissolved in 0.05 M deoxycholic acid was absorbed through oral intubation in fish weighing 200-300 g. 2. The peak of absorption appeared in all fish, 30-45 min after intubation and was followed by a gradual decrease. The extent of absorption was extremely variable, with up to a 20-fold difference between individuals. No detectable insulin was found in fish intubated with vehicle. 3. The absorbed hormone retained 17-88% of its lipogenic bioactivity in vitro. The absorbed insulin lowered the levels of several amino acids in the intubated fish, indirectly indicating that its bioactivity was also retained in vivo.     

Water flow dynamics in the respiratory tract of the carp (Cyprinus carpio L.).

Simultaneous measurements of water velocity in the buccal chamber, and buccal and opercular hydrostatic pressure of carp have revealed surprisingly high water velocities. The high flow velocities mean that, at times, the kinetic energy of flow makes a substantial contribution to the total fluid energy. This suggests that there may be unequal distribution of hydrostatic pressures within the buccal chamber. Anatomical examinations showed that fluid channels in the buccal chamber and gill raker sieve are complex and can be expected to vary spatially and temporally throughout the respiratory cycle. It appears that there is a potential for error in many of the previous analyses of 'gill resistance and energetics of fish breathing based solely on hydrostatic pressure measurements and the simplifying assumption of steady-state conditions.     

Corticosteroids affect the testicular androgen production in male common carp (Cyprinus carpio L.).

Our previous experiments to study the effect of stress adaptation on pubertal development in carp showed that repeated temperature stress and prolonged feeding with cortisol-containing food pellets, which mimics the endocrine stress effects, retarded the first waves of spermatogenesis and decreased 11-ketotestosterone (11KT) plasma levels. The objective of the present study was to investigate whether the decrease in plasma 11KT is caused by a direct effect of cortisol on the steroid-producing capacity of the testis or by an indirect effect, such as a decrease in plasma LH. Pubertal and adolescent isogenic male common carp (Cyprinus carpio L.) were fed with either cortisol-containing food pellets or control food pellets over a prolonged period. Our results indicate that cortisol has a direct inhibitory effect on the testicular androgen secretion independent of the LH secretion. Furthermore, the pubertal period is critical to the influence of cortisol regarding testicular androgen secretion, because the effect is no longer observed at adolescence.     

Microsatellite markers in common carp (Cyprinus carpio L.)

Microsatellite markers of the poly (CA) type in common carp ( Cyprinus carpio L.) are described. Clones containing a (CA) repeat were isolated from a common carp genomic library and sequenced. The number of repeats found was high compared to mammals but comparable with other teleost fishes. Classification of the repeats (perfect, imperfect and compound) are compared with the Atlantic cod ( Gadus morhua L.), rainbow trout ( Oncorhynchus mykiss ), and Atlantic salmon ( Salmo salar L.). A total of 41 primer sets were designed and tested for polymorphism on a test panel of eight animals (derived from outbred lines, inbred lines and gynogenetic clones). Thirty-two markers were found to be polymorphic. The heterozygosity in the outbred animals was 60·4%, 51·1% in the inbred animals and 0% in the gynogenetic clones. The average number of alleles among the eight animals was 4·7 per marker. Six markers (18·8%) gave an additional polymorphic amplification product besides the polymorphic amplification product in the expected size range. The possibility that these loci are tetraploid is discussed. The polymorphic loci described for common carp will be valuable as genetic markers for use in population, breeding, and evolutionary studies.     

Investigation on carp, Cyprinus carpio L. gynogenesis

A reliable method for gynogenetic mass-production has been developed in carp ( Cyprinus carpio L.). The effect of different cold shock treatments applied after fertilization were examined. The yield of viable diploid gynogenetic animals could be significantly increased when the cold shock was applied 5 min or 15 min after fertilization. A negative correlation has been found between the effectiveness of these two treatments, so both were used on separate batches of eggs. By means of gynogenesis, the crossover frequency and map distance of the transferrin locus have been determined. Successful pilot experiments were made on grass carp ( Ctenopharyngodon idella Val.) and sheatfish ( Silurus glanis L.) with respect to gynogenetic mass production.     

Segregation of microsatellite alleles and residual heterozygosity at single loci in homozygous androgenetic common carp (Cyprinus carpio L.).

Thirty-three androgenetic progeny groups of common carp were analysed using 11 microsatellite markers to (i) verify the homozygous status of the 566 androgenetic individuals, (ii) analyse the microsatellite allele segregation, and (iii) study the possible association of microsatellite alleles with phenotypic traits. In total, 92% of the androgenetic individuals proved to be homozygous at all 11 loci. Forty-three of the 47 heterozygous individuals were heterozygous at a single locus only. This heterozygosity was probably due to DNA fragments caused by UV irradiation of the eggs. although the maternal origin of the fragments could not be proved beyond doubt. Screening with 11 microsatellites also revealed two linkage groups, a segregation distortion at two microsatellite loci, and the possible association of some microsatellites with mass, length, stress-related plasma cortisol levels, and basal plasma glucose levels. The success of the linkage and association study could be explained by a low recombination frequency due to high chiasma interference. This would imply a relatively short genetic map for common carp.     

Zinc-induced damage to carp (Cyprinus carpio L.) erythrocytes in vitro

Fish erythrocytes were used to elucidate the effect of zinc ions on the cell antioxidant defence system. It was detected that an increase of the Zn2+ concentration (0.01-1 mM) leads to a marked decrease (p < 0.05) in the catalase and the glutathione peroxidase activities. We observed a loss of 14-39% activity of glutathione peroxidase, and 16-20% diminution for catalase. No significant changes were found in case of the superoxide dismutase. Incubation of red blood cells with zinc brought about a decrease of the erythrocyte thiol group content. Treatment of carp erythrocytes with zinc ions also resulted in enhanced hemolysis and in the induction of significant (p < 0.001) changes in the intracellular glucose level. The increase of glucose concentration in the erythrocytes was correlated with increased concentration of metal in the incubation medium. It was proposed that Zn could affect transport systems across the red blood cells and therefore increased the permeability of the membranes to small molecules (e.g. hexose), and led to hemolysis. Zinc ions could act as a potential cell toxicant, leading to disturbances in functions of the antioxidant defence system and to alterations in the erythrocyte membrane properties.     

Studies of the antibody heterogeneity in carp (Cyprinus carpio L.). I. Electrophoretic and isoelectric spectra of anti-DNP-antibodies]

Carp IgM as well as carp anti-DNP-antibodies migrate electrophoretically very well as a diffuse band into polyacrylamide gel of large pore size. The isoelectric spectra of the carp anti-DNP-antibodies are heterogeneous and show bands in the pI-range of 4.0 to 6.4. The activity of focused anti-DNP-antibodies could be demonstrated in the pI range between 5.4 to 6.4 even in high antibody dilutions. The investigated structural heterogeneity of the anti-DNP-antibodies of carp is a further proof for the phylogenetically early onset of a large antibody heterogeneity of lower vertebrates.     

Response of common carp (Cyprinus carpio L.) leucocytes to hormone-induced stress

The influence of cortisone on leucocytes composition in the blood of common carp (Cyprinus carpio L.) is studied. Following the hormone injection, the relative number of leucocytes decreased and the number of neutrophils and blast-form cells increased in the leucocyte spectrum of experimental fish.     

The effect of oral immuno-stimulation in juvenile carp (Cyprinus carpio L.)

The effect of a 2-week period of oral immuno-stimulation from the age of 2 or 6 weeks post-fertilisation (wpf; before and after reaching the ability to produce antibodies) onwards was investigated on various immune functions of the common carp, Cyprinus carpio. The immuno-stimulants Aeromonas salmonicida lipopolysaccharide, Yeast DNA (containing unmethylated CpG motifs) or high-M alginate (an extract of algae containing poly-mannuronic acid) were used. The effect of this treatment was studied on the kinetics of B cells in head kidney and peripheral blood leucocytes using flow cytometry, on the total plasma IgM level using ELISA, on cytokine and inducible nitric oxide synthase (iNOS) expression in the intestine, and acute phase protein expression in the liver, using real time quantitative PCR, and on exposure to Vibrio anguillarum. Oral administration of immuno-stimulants from 6 wpf resulted in decreased WCI12(+) (B) cell percentages in PBL (only after administration of LPS) and head kidney (all test groups), and a decreased total IgM level in plasma, suggesting that suppressive effects are strongly indicative of oral or juvenile tolerance. After administration from 2 wpf, the effects on WCI12(+) (B) cell percentages were less pronounced: the group fed with Yeast DNA showed higher percentages compared to the control group at 6 wpf, but lower percentages at 8 wpf. No changes were observed in the cytokine or iNOS expression levels in the intestine or acute phase protein expression in the liver. A challenge with V. anguillarum resulted in an initially higher cumulative mortality in the group fed with LPS, but lower mortality in the groups fed with Yeast DNA or high-M alginate compared to the control group, providing a provisional warning especially for the use of pathogen-derived immuno-stimulants, such as A. salmonicida LPS, in larval and juvenile fish.     

Monoaminergic innervation of the kidney of the carp (Cyprinus carpio L.)

By means of fluorescent-histochemical method distribution of monoaminergic structures has been studied in the sympatho-adrenal system of the carp (Cyprinus carpio L.) mesonephros. Catecholaminergic nervous fibers are revealed in the walls of adrenal and venous vessels and glomerular arterioles. In the glomerular capillaries they are not found. In the walls of the venous vessels, besides catecholamine-containing nervous fibers, chromaffin cells with long processes running along the vessel are observed. Some contacts of catecholamine-containing structures of the mesonephric vascular wall with the proximal and, very seldom, with distal parts of the nephron canaliculi are demonstrated. Structures containing indolamines are not revealed.     

Microspectrophotometric and scanning microphotometric studies of carp (Cyprinus carpio L.) erythrocytes

Carp (Cyprinus carpio) hemoglobin readily autoxidizes in blood smears. Quantification of Soret-band absorbance in individual erythrocytes by means of scanning cytophotometry therefore requires more elaborate methods of preparation of blood samples. Of the fixatives that have been tested, suspension of whole blood in isotonic salt solutions containing glutaraldehyde was most suitable. Glutaraldehyde-fixed red blood cells are totally resistant to hemolysis. In the course of fixation, hemoglobin is transformed to methemoglobin. Spectrophotometry indicated extensive similarities between glutaraldehyde-fixed carp methemoglobin and human methemoglobin. In aqueous solutions, the intensity of the Soret-peak was pH-dependent. The allosteric modifier organic polyphosphate caused an R----T transition, resulting in increased molar extinctions. Dried preparations showed Soret-spectra that were not influenced from either pH or organic polyphosphate concentration of the aqueous suspensions in which the erythrocytes had been stored. The same was true for slide preparations of cyanomethemoglobin, easily derived from methemoglobin on addition of potassium cyanide. In the absence of oxygen fresh blood cells from carp slowly transform their hemoglobin into deoxyhemoglobin. Spectra of the intermediate stages of deoxygenation, Hb4(O2)3, Hb4(O2)2 and Hb4(O2), as well as mixtures of these intermediates, could be monitored.     

Observations on the ultrastructure of the carp liver (Cyprinus carpio L.)

The paper presents information about the fine structure of the sinusoids, the space of Dissé, the development of the bile canaliculi and some compartments of the hepatocytes in the liver of the carp. The sinusoids are covered completely by flat endothel cells. The plasma of these cells contains numerous vesicles. The endothelial cells possess plasmatic processes, which extend into the space of Dissé. The fine structure of the space of Dissé corresponds to that of mammals, the existence of fibrebundles included. The bile canaliculi don't develop intracellularly as described by other authors. They run intercellularly as in mammals, but they form diverticles which reach into the plasma sideways. The rough ER was found in two types. Outwardly the liver is limited by a layer of connective tissue existing in two different layers.     

Morphometry of muscle fibre types in the carp (Cyprinus carpio L.)

Summary Ultrastructural parameters of muscle fibre types of the carp (Cyprinus carpio L.) were measured and compared with their contractile properties. In red fibres, which are slower than pink fibres, the relative length of the junction between the T system and the sarcoplasmic reticulum (T-SR junction) is smaller and the Z lines are thicker than in pink fibres. Pink fibres have a smaller relative length of T-SR junction than white fibres from the axial muscles. The two types of red fibres present in carp muscle also differ in their relative lengths of T-SR junction. Significant differences in the relative areas of the SR were not found.The relative volume of myofibrils in red fibres is two-thirds that in pink fibres, a difference that is not reflected in the maximal isometric tetanic tensions of these types. Red fibres, which are less easily fatigued than pink fibres, have larger relative volumes of subsarcolemmal and intermyofibrillar mitochondria. Small pink fibres have a larger relative volume of subsarcolemmal mitochondria than large pink fibres, but have a similar relative volume of intermyofibrillar mitochondria. Small and large pink fibres differ in the relative volumes of their membrane systems, but have similar relative lengths of T-SR junction.