Retention of visible implant tags by juvenile brown trout

The retention and readability of visible implant (VI) tags were evaluated in juvenile brown trout. The tags were implanted in the adipose eyelid tissue posterior to the left eye, in a total of 6697 fish in 18 groups. During the first year, tag retention increased with tagging experience, from 58 to 64% in 3–summer–old fish and from 86 to 96% in 4–summer–old fish. The effect of fish size at tagging on tag loss was clear in the 3–summer–old groups in 1991 (ANOVA, P = 0.00001). Tag retention in 3–year–old and 4–summer–old fish in 1992 and 1993 varied from 96 to 99% and the handling of the fish after tagging affected retention significantly ( t –test, P = 0.0002), the average being 98.7% for those transposed by submerging and 96.5% in those transposed by dropping. After inspection the fish were released into a natural environment. On recapture after 6 months or more the red tags (faded almost to white) and yellow tags (originally close to white) were difficult to distinguish. No infection or tissue damage was found after either inspection or stocking.     

RNA-DNA ratios in white muscle tissue biopsies reflect recent growth rates of adult brown trout

Recently developed fluorescence techniques were used to quantify RNA-DNA ratios in white epaxial muscle tissue biopsies taken from live adult brown trout. RNA-DNA ratios from small biopsies (4–24 mg wet weight) were measured concurrently with growth over 5 weeks in two groups of brown trout fed different rations, 5% BW day^{– 1} (reference) and 1% BW week^{– 1}(treatment). Reduced rations had significant effects on RNA-DNA ratios in muscle tissue biopsies. The treatment group had significantly lower mean RNA–DNA ratios than the reference group after the first week. After 5 weeks of treatment the mean RNA–DNA ratios of the treatment and reference groups were 1–98 and 4–31, respectively. RNA-DNA ratios from muscle tissue biopsies reflected recent growth rates in the two groups of fish. This technique allows a biochemical measurement of growth rates in adult fish without mortality.     

Sex differentiation in the European eel: histological analysis of the effects of sex steroids on the gonad

The effects of sex steroids on sex differentiation in the European eel were studied. The steroids, 17α-methyltestosterone (MT) and 17α-ethynylestradiol (EE), were given in the diet to 6–8 cm elvers and to 15–18 cm and 22–25 cm yellow eels. In our rearing conditions a very large percentage of the untreated eels developed as males. No masculinizing effect of MT could be demonstrated. The EE, administered at a dose of 10 mg kg^-1 of diet to 6–8 cm elvers and 15–18 cm eels, induced ovarian differentiation in about 90 and 65% of eels respectively, while in the control <5% of females was recorded. In 22–25 cm yellow eels a moderated feminizing effect was observed.
Histological analysis of the gonads of treated eels showed that sex steroids affect the gonadal structure. The androgen stimulates hypertrophy of compact connective tissue, early differentiation of Leydig cells, Sertoli cells and early formation of the spermatic duct. Oestrogen inhibits the differentiation of these structural components and stimulates the differentiation of follicular cells and an ovarian structure.
The involvement of gonadal structural components is discussed in relation to the effect of steroid treatment and to the peculiarities of sex differentiation in the European eel.     

Fluorometric determination of DNA in fixed tissue using ethidium bromide

The measurement of DNA in tissue samples fixed in ethanol/acetic acid is described. Small, fixed tissue samples are digested by warm alkaline treatment followed by neutralization with HCl, and DNA is determined by complex formation with the dye ethidium bromide (EB). When standard DNA from calf thymus was treated similarly, a hyperchromicity of 8–12% and a reduction in fluorescence intensity of the EB-DNA complex to 55% was observed. The NaOH concentration (0.5–2.0 mol/liter) or the temperature (50–60°C) used for the digestion of tissue, as well as subsequent ribonuclease or protease treatment had no effect on the observed tissue DNA concentrations.     

Tolerance of Pomacanthus imperator to hypoosmotic salinities: changes in body composition and hepatic enzyme activities

Emperor angel fish Pomacanthus imperator were acclimated to various salinities (33, 22, 15, 10 and 7%) for 1 month and changes in serum chemistry, body composition and liver enzyme activities were monitored. No mortality was observed in fish adapted throughout the salinity range of 7–33% and the limit of survival was around 5%. Serum Na^+; and Cl ^– exhibited a stepwise decrease as the salinity was reduced from 33–7% but no concomitant tissue hydration could be observed. Branchial Na-K-ATPase activity was generally higher in hypoosmotic (7–15%) than in hyperosmotic salinities (22–33%o). Serum concentrations of glucose, lipid, α-amino acids and protein as well as the haematocrit value were all elevated in the salinity range of 15–22%. Serum cortisol was markedly lower but serum thyroxine was significantly elevated at 15–22%. Fish adapted to 15–22% had higher hepatosomatic indices, liver glycogen and muscle lipid contents and hepatic glucose-6-phosphate dehydrogenase activity. These data indicate that Pomacanthus imperator is a' physiologically euryhaline' species and metabolism in isoosmotic salinities has been fundamentally re-organized to favour carbohydrate and lipid retention.     

Effects of dietary polyunsaturated fatty acid deficiencies on mortality, growth and gill structure in the turbot, Scophthalmus maximus

The preparation of fish oil concentrates containing only ( n -3) polyunsaturated fatty acids (PUFA) with different ratios of 20:5 ( n -3)/22:6 ( n -3) is described. Three groups of turbot were maintained on different diets containing: 1, 10% of the dry weight of the diet as natural fish oil, equivalent to 2.5% ( n -3) PUFA and 0–23% ( n -6) PUFA; 2, 10% of the dry weight of the diet as palmitic acid, i.e. no PUFA; 3, 8–7% palmitic acid and 1–3% of the dry weight as ( n -3 PUFA and negligible ( n -6) PUFA. Only the fish on the diet containing natural fish oil showed significant growth over a 15-week period. In addition there were high mortalities on the two experimental diets (2 and 3). Changing the ratio of 20:5 ( n -3)/22:6 ( n -3) from 13–8 to 2–2 in the diet containing 1 3% (n-3) PUFA and negligible ( n -6) PUFA markedly decreased the mortalities. Fish fed the two experimental diets (2 and 3) developed gross changes in gill structure involving the disappearance of chloride cells, a 'sloughing off' of the epithelium along the primary and secondary filaments and an accumulation of cellular material in the inter-lamellar spaces. The tissue ultimately disintegrated to leave a skeleton of connective tissue and a mass of cellular material in the inter-lamellar spaces. It is concluded that 22:6 (n-3) is an essential fatty acid for turbot and that the gill epithelium is a sensitive indicator of this deficiency in this species.     

Water relations of Picea abies. I. Comparison of water relations parameters of spruce shoots examined at the end of the vegetation period and in winter

Seasonal changes of some water relations parameters of Norway spruce shoots ( Picea abies [L.] Karst.) were studied during two experiments using the pressure-volume analysis. For each experiment only shoots of a single tree were used.
During the first study, the course of the turgor loss point (as bulk osmotic pressure when turgor first reaches zero, πp) of shoots developed in late 1986 vegetation period, were measured in 1987. The turgor loss point decreased temporarily from –2.5 MPa at the beginning of the year to –3.3 MPa at the end of March, but then increased to the original level for the rest of the year.
During the second study, water relations parameters were measured in late summer 1987 and in late winter 1988. Winter shoots at full water saturation contained up to 20% less water than in late summer. Accordingly, the bulk osmotic pressure at full water saturation (π_p) decreased from –1.7 MPa in late summer to –1.9 MPa in winter, π_p decreased also from –2.2 MPa to –2.8 MPa. However, the amount of osmotically active substances (mOsmol, N) remained unchanged. The relative amount of apoplastic water in the total shoot water content appeared to drop insignificantly from 17% to 15%.
The results show that the decrease in π_o and π_p in late winter is not due to an accumulation of osmotically active substances in the vacuoles but is due to a decrease in tissue water content. The temporary reduction of the symplastic volume by deposition of osmotically inert substances seems to be the most probable cause of this phenomenon.     

COMPARISON OF THE EFFECTS OF DEPOLARIZING AGENTS AND NEUROTRANSMITTERS ON REGIONAL CNS CYCLIC GMP LEVELS IN VARIOUS''ANIMALS

Abstract— The effects of 121 m m -K⁺, 10 m m -glutamate, 5 m m -GABA, 1 m m -glycine, 0.1 m m -NE, and 1–10 μ m ACh on cyclic GMP levels in tissue slices prepared from cerebral cortex and cerebellum of mouse, rabbit, guinea-pig, cat, and rat were studied. Basal levels of cyclic GMP in the cerebella of mice, guinea-pigs and cats were 4–15 and 70 pmol/mg prot in rat, whereas in the cerebral cortex of the same animals, levels were only 0.6–2 pmol/mg prot. In contrast, basal levels of the cyclic nucleotide were 1–2 pmol/mg prot in both of these regions in rabbit brain. Only 121 m m -K⁺ was capable of increasing cyclic GMP levels in all the tissues studied. Elevations ranged from 30% in rat cerebral cortex to 2800% in mouse cerebellum. Glutamate produced a 30–1000% rise of cyclic GMP levels in all tissues except rabbit cerebellum. NE elevated levels of cyclic nucleotide 2- to 3-fold in slices of cerebellum from all species studied but had no effect in cerebral cortex. GABA and glycine had no effect in any tissue except mouse cerebellum. ACh had no consistent effect on levels of cyclic GMP in any brain region investigated. These results suggest that mechanisms regulating cyclic GMP levels in mammalian CNS vary among brain regions and among animal species.     

Thermal acclimation in selected tissues of the fathead minnow, Pimephales promelas rafinesque

Environmental variables besides temperature such as length, weight, sex, and photoperiod were shown to affect the metabolic rate in gill and white muscle tissue. This required statistical adjustment to account for these factors. Oxygen consumption rates for gill tissue were greater following cold acclimation at all determination temperatures. This suggests thermal compensation had occurred. Metabolic rates for white muscle tissue showed inverse compensation between 7 °C and 15 °C, but little compensation was evident between 15 °C and 23 °C. Both gill and white muscle tissue had Q₁₀ values ranging between 1–00 and 2–00. These are lower than values previously recorded for fathead minnows.     

Protochlorophyllide forms in non-greening epicotyls of dark-grown pea (Pisum sativum)

Low-temperature fluorescence emission spectra of 6.5-day-old dark-grown epicotyls of pea ( Pisum sativum ) revealed the presence of protochlorophyll(ide). The upper part of the epicotyl contained 30% of the protochlorophyll(ide) content per fresh weight found in pea leaves, whereas the lower part contained 3%. Three discrete spectral forms of protochlorophyll(ide) were clearly distinguished after Gaussian deconvolution of fluorescence excitation and emission spectra. Adding the satellite bands of the Qy_(0-0) transitions (the emission vibrational (Emv) bands with correlated amplitudes, gave the following delineation: Ex439–Em629–Emv684, Ex447–Em636–Emv700 and Ex456–Em650–Emv728. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) followed by immunodetection of whole tissue extracts of the epicotyl indicated the presence of NADPH-protochlorophyllide oxidoreductase (EC 1.3.1.33). Electron micrographs showed prolamellar bodies in at most 11 % of the plastid profiles of the epicotyl cells. These prolamellar bodies were smaller, and many of them showed less regular structure than those of the leaves. Taken together, the results indicate that the protochlorophyll(ide) in epicotyls is arranged in a different way than in leaves.     

Seasonal changes in energy and the energy cost of spawning in Gulf of Alaska Pacific cod

In the Gulf of Alaska, adult Pacific cod exhibited an annual cycle of condition, gonad index and liver index in which maximum values occurred in ripe fish in March and minima in July. About 30–31 % of prespawning stored energy was expended during the spawning effort. The energy associated with spawning derived from liver (24% and 18%), somatic tissue (22% and 33%) and gonad (53% and 48%) for females and males, respectively. Liver index and gonad index at the time of sampling were directly related in females, but in males gonad index was best related to liver index 1–3 months earlier.
The Pacific cod is very similar to the Atlantic cod in terms of energy cycling, maximum gonad sizes, energy expended during spawning and gonadal contribution to energy expenditure. However, in Pacific cod, somatic tissue contributes markedly to energy expended during reproduction. The Pacific cod cod differs from the walleye pollock with respect to gonad index (13% and 20%ν. 20% and 8% for females and males, respectively), spawning weight loss (25%ν. 38%), liver energy loss during spawning (71%ν. 55%) and energy cost of spawning.     

Fiber development in preanthesis cotton ovules

A tissue culture method was developed to investigate the production of cotton (Gossypium hirsutum L. cv. Texas Marker-1) fibers in vitro. Ovules were excised from 3, 5, 7 and 9 days preanthesis ovaries and placed on an agar-solidified, modified Murashige and Skoog medium containing 2.3 μ M kinetin and 0.45 μ M –2,4–dichlo-rophenoxyacetic acid or 2.3 μ M kinetin and 10.7 μ M naphthaleneacetic acid. Ovules formed fibers and callus tissue. Fibers formed in vitro were up to 10 mm long, 10–22 μ wide and the cell wall was 1–3 μ M thick. Callus tissue cells were subcultured for over 25 weeks and their degree of elongation was monitored. The ability of ovule-derived cells to direct expansion in a longitudinal direction diminished, while lateral expansion increased with time in culture.     

Potentially pathogenic vibrios associated with mussels from a tropical region on the Atlantic coast of Brazil

Mussels ( Perna perna ) harvested on the coast of Ubatuba, in three different stations in the State of São Paulo, Brazil, were examined for Vibrio spp. over a 1 year period. The ranges of most probable number (MPN 100 g^-1) were: Vibrio alginolyticus (<3–24000), V. parahaemolyticus (<3–24000), V. fluvialis (<3–1100), V. cholerae non-O1 (<3–23), V. furnissi (< 3–30), V. mimicus (< 3–9) and V. vulnificus (< 3–3). The highest incidence was observed for V. alginolyticus (92–100%), followed by V. parahaemolyticus (67–92%), V. fluvialis (34–67%), V. vulnificus (8–17%), V. furnissii (0–17%), V. mimicus (0–17%) and V. cholerae non-O1 (0–8%). Tests for virulence factors were positive in 34.1% of the vibrios in the rabbit ileal loop and 31.7% in the Dean test. Positive results in the Kanagawa test were obtained with 0.51% of V. parahaemolyticus strains. The mean values (MPN 100 g^-1) of faecal coliforms in mussels from the three regions varied from 1100 to 44000, and seawater collected at the same stations gave average values for faecal coliforms in the range 18–3300 MPN 100 ml^-1. These results highlight the potential risks of food poisoning associated with raw or undercooked seafood.     

Light-induced changes in the lipid composition and ultrastructure of plastids from potato tubers

Sandelius, A. S. and Liljenberg, C. 1982. Light-induced changes in the lipid composition and ultrastructure of plastids from potato tubers. – Physiol. Plant. 56: 266–272.
Amyloplasts and starch containing plastids from green tissue – amylochloroplasts – from potato tubers ( Solanum tuberosum L., var. King Edward) were separated from other cell organelles by sedimentation in a discontinuous sucrose gradient. Their lipid composition was analysed with emphasis on galactolipids and phospholipids and the fatty acid compositions of these lipids. Irradiation of the tubers caused increased ratios of monogalactosyl diacylglycerol to digalactosyl diacylglycerol and of total galactolipids to total phospholipids in the plastid membranes. Furthermore, the degree of unsaturation of the fatty acids increased in all lipid classes analysed, this effect being most prominent in the galactolipids. The ultrastructural studies made on tuber tissue revealed that irradiation caused a change in starch grain size distribution concomitant with formation of membrane structures resembling grana within the envelope. In many cases prolamellar bodies and plastoglobuli were present.     

Species of Anguilla as indicators of mercury in the coastal rivers and lakes of Victoria, Australia

Mercury concentrations in the axial muscle tissue of most (243) of the 254 Anguilla australis and most (20) of the 27 A. reinhardtii collected from 30 sites in coastal rivers and lakes in Victoria, Australia, during 1975–78 were well below the Australian statutory health limit (0.5 μg g⁻¹ wet weight). For A. australis the mean mercury concentration was 0.17 μg g⁻¹ (±0.16 s.d. , range 0.01–1.60 μg g⁻¹); for A. reinhardtii the values were 0.37 ± 0.23 μg g⁻¹ (range 0.12–1.10 μg g⁻¹). Statistical analyses showed that variation in mercury concentration due to total length accounted for only 13% of the total variation in A. australis and 2% in A. reinhardtii whereas locality accounted for 54 and 68%, respectively. Both species are thus considered suitable as indicators of mercury pollution.     

The Capacity of Testicular Cells of the Postnatal Rat to Reorganize into Histotypic Structures

Cell reorganization experiments in vitro were performed with dissociated rat testes at different ages of postnatal development namely, newborn, 8–10, 18–25, 35–40, and 90 days. Only newborn and juvenile rat testicular cells reassociated into testicular-like organization in rotation culture. Puberal and adult rat testicular cells show morphogenetic organization when they were deprived of germ cells by busulphan pretreatment. A factor present in testicular tissue of puberal and adult rats inhibits reorganization. The inhibitor is confined to the spermatic cell fraction in the testis.     

Low-temperature preservation of mammalian cells in tissue culture with polyvinylpyrrolidone (PVP), dextrans, and hydroxyethyl starch (HES)

The effects of freezing and thawing on Chinese hamster cells in tissue culture in the presence of PVP, HES, and various dextrans have been investigated. Cooling and thawing rates within a limited range (20–260 °C per min for cooling and 6–115 °C per min for thawing) were studied and best results were achieved with a cooling rate of 20 °C per min and a thawing rate of 115 °C per min in both 10% PVP, and 10% HES. Experiments demonstrated HES to be as good as, and possibly better than PVP. A number of dextrans of average molecular weight (10,000–500,000 daltons) were shown to be poor as cryoprotective agents in contrast to results obtained with this polymer with red cells and bacteria. The presence of 10% serum during all freezing and thawing procedures decreased day-to-day variability and with dextrans increased their limited effectiveness.     

Biogenic amines and their metabolites in mouse brain tissue: development, optimization and validation of an analytical HPLC method

A simple and fast HPLC method based on an isocratic, reversed-phased ion-pair with amperometric end-point detection for simultaneous measurement of noradrenergic (MHPG/NA and A), dopaminergic (DOPAC, HVA/DA) and serotonergic (5-HIAA/5-HT) compounds in mouse brain tissue was developed. In order to improve the chromatographic resolution (Rs) with an acceptable total analysis time, experimental designs for multivariate optimization of the experimental conditions were applied. The optimal conditions for the separation of the eight neurotransmitters and metabolites, as well as two internal standards, i.e., DHBA and 5-HMT, were obtained using a mixture of methanol–phosphate–citric buffer (pH 3.2, 50 mM) (9:91, v/v) containing 2 mM OSA as mobile phase at 32 °C on a microbore ALF-115 column (150 mm × 1.0 mm, 3 μm particle size) filled with porous C₁₈ silica stationary phase. In this study, a two-level fractional factorial experimental design (½ 2^K) was employed to optimize the separation and capacity factor (k′) of each molecule, leading to a good separation of all biogenic amines and their metabolites in brain tissue. A simple method for the preparation of different bio-analytical samples in phosphate–citric buffer was also developed. Results show that all molecules of interest were stabilized for at least 24 h in the matrix conditions without any antioxidants. The method was fully validated according to the requirements of SFSTP (Société Française des Sciences et Techniques Pharmaceutiques). The acceptance limits were set at ±15% of the nominal concentration. The method was found accurate over a concentration range of 4–2000 ng/ml for MHPG, 1–450 ng/ml for NA, 1–700 ng/ml for A, 1–300 ng/ml for DOPAC, 1–300 ng/ml for 5-HIAA, 1–700 ng/ml for DA, 4–2800 ng/ml for HVA and 1–350 ng/ml for 5-HT. The assay limits of detection for MHPG, NA, A, DOPAC, 5-HIAA, DA, HVA and 5-HT were 2.6, 2.8, 4.1, 0.7, 0.6, 0.8, 4.2 and 1.4 pg, respectively. It was found that the mean inter- and intra-assay relative standard deviations (RSDs) over the range of standard curve were less than 3%, the absolute and the relative recoveries were around 100%, demonstrating the high precision and accuracy, and reliability of the analytical method described to apply in routine analysis of biogenic amines and their metabolites in brain tissue.     

Shoot initiation in light- and dark-grown tobacco callus: the role of ethylene

Shoot-forming tobacco ( Nicotiana tabacum L. cv. Wisconsin 38) callus produces less endogenous ethylene than non-shoot-forming tissue cultured in the light (16 h photoperiod) or the dark. In shoot-forming tissue more ethylene is produced early in culture (days 0–5) than later. Also dark-grown tissue produces much more ethylene than light-grown. On the basis of experiments in which (1) gaseous ethylene was added to or (2) CO₂ removed from the flasks, (3) Ethrel (an ethylene releasing agent) and (4) 1-aminocyclopropane 1-carboxylic acid (an ethylene precursor) were added to the medium, it was determined that this gaseous phytohormone had two contrary effects on shoot initiation (shoot primordium formation). Early in culture (days 0–5) endogenous or exogenous ethylene inhibited organogenesis, but later (days 5–10) exogenous ethylene or increased endogenous ethylene production speeded up primordium formation.     

Free Fatty Acids in the Rat Brain in Moderate and Severe Hypoxia

Abstract: The effects of mild, moderate, and severe hypoxia on cerebral cortical concentrations of free fatty acids (FFAs) were investigated in artificially ventilated rats under nitrous oxide anaesthesia. No change occurred during either mild (arterial Po₂ 35–40 mm Hg) or moderate (Po₂ 25–30 mm Hg) hypoxia. The effects of severe hypoxia (Po₂ about 20 mm Hg) combined with hypotension (mean arterial blood pressure 80–85 mm Hg) varied with the EEG pattern and the tissue energy state. Thus, a major increase in total as well as in individual FFAs occurred first when EEG was severely depressed (almost isoelectric) and energy homeostasis disrupted. On a relative basis the greatest change occurred in free arachidonic acid. It is concluded that hypoxia is associated with an increase in the concentrations of FFAs in brain tissue, provided that tissue oxygen deficiency is severe enough to cause tissue energy failure. However, an increase in FFAs does not invariably accompany minor reductions in the adenylate energy charge (EC) of the tissue.