Profiles of mRNA expression for prolactin,growth hormone,and somatolactin in Japanese eels,Anguilla japonica: The effect of salinity,silvering and seasonal change

For understanding the functions of the growth hormone (GH)/prolactin (PRL)/somatolactin (SL) family of hormones, we examined pituitary mRNA expression of these hormones in anguillid eels in relation to salinity difference, silvering, and seasonal change. Female Japanese eels (Anguilla japonica) were collected in the brackish Hamana Lake and its freshwater rivers from July to December. To clarify the effect of salinity, the habitat use history of the eels were determined using otolith microchemistry. Expression levels of mRNA of each hormone were determined using real time PCR. Although GH and PRL have been known to be osmoregulatory hormones, there were no consistent differences in expression levels of these hormones between different salinity habitats. In contrast, SL mRNA expression was higher in eels from freshwater rivers than from the brackish lake. GH mRNA expression clearly decreased during silvering, whereas PRL and SL mRNA expression did not change. We also showed that PRL mRNA and SL mRNA decreased in the brackish lake and PRL mRNA increased in freshwater rivers from autumn to early winter. These findings provide basic knowledge for a further understanding of the role of these hormones.     

Influence of thyroidal status on pituitary content of thyrotropin beta- and alpha-subunit, growth hormone, and prolactin messenger ribonucleic acids

Thyroidectomized rats were used to study the effects of a single injection of T3 on pituitary mRNA synthesis and hormone secretion. T3 was injected ip at doses of 0, 0.2, 1, or 5 micrograms/100 g body weight, and and animals were killed 24 h later. T3 caused a significant decrease in serum TSH, but caused no significant change in either serum GH or PRL. Pituitary mRNA was quantified by slot blot hybridization with cDNA probes specific for alpha-TSH, beta-TSH, PRL, and GH. We found that both the alpha and beta mRNA subunits decreased, that PRL mRNA remained relatively unchanged, and that GH mRNA increased with increasing T3 dose. The data show that a single dose of T3 can profoundly influence mRNA levels in the anterior pituitary; the lowest dose of T3 caused maximum inhibition of alpha-TSH mRNA while beta-TSH mRNA declined further in a dose-dependent manner.     

The sulfide tolerance of milkfish and tilapia in relation to fish kills in farms and natural waters in the Philippines

Fish kills of milkfish Chanos chanos and tilapia Oreochromis spp. now occur frequently in brackish, marine, and freshwater farms (ponds, pens, and cages) in the Philippines. Aquafarms with high organic load, limited water exchange and circulation, no aeration, and high stocking and feeding rates can become oxygen-depleted and allow sulfide from the sediments to appear in the water column and poison free-swimming fish. The sulfide tolerance of 2–5 g milkfish and 5–8 g O. mossambicus was determined in 25-liter aquaria with flow-through sea water (100 ml min-1) at 26–30 °C and sulfide stock solutions pumped in at 1ml min-1. Total sulfide concentrations in the aquaria were measured by the methylene blue method and used in the regression against the probits of % survival. Four experiments showed that the two species have similar sulfide tolerance. In sea water of pH 8–8.5, about 163 ± 68 μM or 5.2 ± 2.2 mg l-1 total sulfide (mean ± 2 se) or 10 μM or 313 μg l-1 H2S was lethal to 50% of the fish in 4–8 h, and 61 ± 3 μM total sulfide or 4 μM H2S in 24–96 h (to convert all sulfide concentrations: 1 μM = 32 μg l-1). Earthen pond bottoms had 0–382 μM total dissolved sulfide (mean ± sd = 54 ± 79 μM, n = 76); a tenth of the samples had >200 μM. The water column may have such sulfide levels under hypoxic or anoxic conditions. To simulate some of the conditions during fish kills, 5–12 g milkfish were exposed to an abrupt increase in sulfide, alone or in combination with progressive respiratory hypoxia and decreasing pH. The tests were done in the same flow-through set-up but with sulfide pumped in at 25 ml min-1. The lethal concentration for 50% of the fish was 197 μM total sulfide or 12 μM H2S at 2 h, but 28–53 μM sulfide allowed fish to survive 6–10 h. Milkfish in aquaria with no aeration nor flow-through sea water died of respiratory hypoxia in 5–8 h when oxygen dropped from 6 to 1 mg l-1. Under respiratory hypoxia with 30–115 μM sulfide, the fish died in 2.5–4 h. Tests with low pH were done by pumping a weak sulfuric acid solution at 25 ml min-1 into aquaria with flow-through sea water such that the pH dropped from 8 to 4 in 5 h. Under these conditions, milkfish died in 7–9 h when the pH was 3.5. When 30–93 μM sulfide was pumped in with the acid, the fish died in 2–6 h when the pH was still 4.5–6.3. Thus, sulfide, hypoxia, and low pH are each toxic to milkfish at particular levels and aggravate each other's toxicity. Aquafarms must be well oxygenated to prevent sulfide toxicity and fish kills. This revised version was published online in July 2006 with corrections to the Cover Date.     

Effects of spironolactone and RU486 on gene expression and cell proliferation after freshwater transfer in the euryhaline killifish

We have explored the possible mechanisms by which mineralocorticoid (MR) and glucocorticoid (GR) receptors regulate the response to freshwater transfer in the gills of the euryhaline killifish Fundulus heteroclitus. Killifish were implanted with RU486 (GR antagonist) or spironolactone (MR antagonist) at doses of 0.1–1.0 mg g⁻¹, and subsequently transferred from 10‰ brackish water to freshwater. Compared to brackish water sham fish, mRNA expression of CFTR and NKCC1 decreased in the gills of sham fish transferred to freshwater, whereas Na⁺,K⁺–ATPase α_1a mRNA expression and α protein abundance, as well as cell proliferation (detected using BrdU) increased. Spironolactone inhibited the normal increase in cell proliferation and Na⁺,K⁺-ATPase expression after freshwater transfer. RU486 increased plasma cortisol levels and may have slightly inhibited Na⁺,K⁺–ATPase activity, but did not change α _1a expression. RU486 had no effect on cell proliferation in the non-lamellar region of the gills, but increased proliferation in the lamellar region. Neither antagonist inhibited the suppression of CFTR or NKCC1 expression after freshwater transfer. Glucocorticoid receptor expression was reduced in all sham and antagonist treatments compared to untreated controls, but no other consistent differences were observed. The effects of spironolactone suggest that MR is important for regulating ion transport in killifish gills after freshwater transfer.     

The physical factor inducing spawning of the Biwa catfish, Silurus biwaensis

The physical factor inducing spawning of the Biwa catfish, Silurus biwaensis (Siluriformes: Siluridae), an endemic species of Lake Biwa, was investigated on the basis of field observations on 150 nights at the spawning site, the rocky shore of the Seta River, the lake's outlet, and environmental data from April to July in 1989–1994. The catfish gathered at the spawning site when the water level of the river rapidly rose and the rocky shoal became submerged, within a wide range of water temperature (15.0°–30.0°C); their spawning was induced in relation to increases in the submerged portion of the rocky shoal. This observation suggests that water level control operations in Lake Biwa might critically affect the spawning of this and other fish species. Received: December 17, 1999 / Revised: September 13, 2000 / Accepted: January 16, 2001     

Growth hormone and Prolactin-1 gene transcription in natural populations of the black-chinned tilapia Sarotherodon melanotheron acclimatised to different salinities

The effects of salinity on the expression of genes coding for growth hormone (GH) and prolactin-1 (PRL1) were studied in various natural populations of the black-chinned tilapia Sarotherodon melanotheron from West Africa. Individuals were sampled in June 2005 in six locations in Senegal and the Gambia, at various salinities between 0 and 101. The poorest condition factors were recorded in the most saline sampling site and the best growth in the fish from a marine environment. The pituitary GH mRNA levels were significantly higher in fish adapted to seawater, whereas the PRL1 mRNA levels were highest in fish adapted to fresh- and brackish water. These results show that the PRL1 mRNA levels seem to reflect relatively well the differences in environmental salinity, in contrast to those of GH, which would tend instead to reflect the individual growth in each environment. However, no relation could be found between growth in the hypersaline areas and the expression profile of GH. Although the fish analysed were morphologically identical, the expression of genes coding for GH and PRL1 showed large differences between individuals. This inter-individual variation in gene expression remains poorly understood.     

Targeted ablation of gonadotrophs in transgenic mice depresses prolactin but not growth hormone gene expression at birth as measured by quantitative mRNA detection

We previously reported that transgenic ablation of gonadotrophs results in impaired development of cells immunostainable for prolactin (PRL) but not of cells immunostainable for growth hormone (GH) or proopiomelanocortin (POMC) in pituitary of newborn mice. The question remained whether this reduction in PRL protein is a reflection of reduced PRL mRNA expression, or whether this regulation is only situated at the translational level. We therefore generated a new series of transgenic mice in which gonadotrophs were ablated by diphtheria toxin A targeting, and analyzed hormone mRNA levels instead of hormone protein around the day of birth. Pituitary mRNA expression levels of luteinizing hormone- (LH), PRL and GH were quantified using real-time TaqMan RT-PCR. Of the 13 transgenic mice obtained, 8 showed a clear-cut reduction (ranging from 62 to 98%) in LH mRNA levels. PRL mRNA values were significantly reduced in the transgenic mice (p=0.0034), while GH mRNA expression was unaffected (p=0.93). An additional observation was that female newborn mice produce 5 times more LH mRNA than male mice whereas no sex difference was observed for expression levels of PRL and GH mRNA. Moreover, in the wild-type mice, LH mRNA expression was 20-fold higher than GH mRNA expression which in turn was 500- to 1,000-fold higher than PRL mRNA expression, suggesting a low expression level of the PRL gene at birth. In conclusion, the present data support the hypothesis that embryonic development of PRL gene expression is stimulated by gonadotrophs.     

Levels of cadmium and mercury in the hair of Atlantic walruses (Odobenus rosmarus rosmarus) from Svalbard,Norway

Hair samples of 15 adult male Atlantic walruses (Odobenus rosmarus rosmarus) collected from anaesthetized individuals at Svalbard, Norway, were analysed for cadmium and total mercury. The mean level of cadmium was 0.860 ± 0.321 μg/g dry weight (median = 0.811, range = 0.349–1.51 μg/g dry weight) and the mean level of mercury was 0.235 ± 0.100 μg/g dry weight (median = 0.251, range = 0.121–0.424 μg/g dry weight). Levels of cadmium and mercury in hair of walruses from other areas are not known. Both cadmium and mercury levels in hair of walruses from Svalbard are relatively low compared to the levels found in the hair of other marine mammal species. It has been documented from a number of marine species, including marine mammals such as ringed seals and polar bears, that both cadmium and mercury levels at Svalbard are lower than in other areas. It is uncertain as to what degree levels in hair reflect levels in internal organs in walruses. In rare and highly endangered species or populations tissue samples can be difficult to collect. In walruses, it is possible to collect hair from anaesthetized individuals or at the haul-out sites during moult, to monitor heavy metal levels of the population. Accepted: 6 December 1998     

Maternal and neonatal scalp hair concentrations of zinc, copper, cadmium, and lead

Postpartum scalp hair samples from 82 term-pregnancy mother/neonate pairs were analyzed for their concentration of zinc (Zn), copper (Cu), cadmium (Cd), and lead (Pb), using inductively coupled plasma-mass spectrometry. Maternal and neonatal Zn concentrations had geometric means (and 99% confidence intervals) of 122.5 μg/g (117.9–131.5 μg/g) and 146.9 μg (141.5–156.7 μg/g) respectively. Corresponding Cu values were 18.4 μg/g (17.6–23.8 μg/g) and 6.7 μg/g (6.3–7.6 μg/g). Those of Cd were 0.49 μg/g (0.47–0.69 μg/g) in the mothers and 0.57 μg/g (0.55–0.86 μg/g) in the neonates. For Pb, they were 7.95 μg/g (7.60–9.32 μg/g) and 4.56 μg/g (4.39–5.56 μg/g). Cigaret smoking, despite its relatively low prevalence (19.5%), was associated with lower Zn and higher Cd and Pb concentrations and in lower Zn/Cd and Zn/Pb molar concentration ratios. Smoking also altered interelemental relationships, particularly those of Zn with Cd and Pb and those between Cd and Pb. Smoking frequency appeared to show negative dose-response effects on maternal and neonatal Zn concentrations, Zn/Pb molar concentration ratios, and birth weight. Mothers with a history of oral contraceptive (OC) usage had significantly higher Cu concentrations and lower Zn/Cu molar concentration ratios than nonusers, with the highest Cu concentrations and lowest Zn/Cu values being associated with third-generation OCs. No similar effects were elicited in the respective neonatal Cu concentrations. Neither alcohol consumption nor prenatal supplementation with iron and/or folic acid had discernible effects on the maternal or neonatal elemental concentrations. The data from this study suggest that in a given population of term-pregnancy mothers and neonates, significant interindividual variations in hair trace element concentrations can occur, irrespective of commonality of general environment, and that lifestyle factors, including cigaret smoking and OC usage history, can be significant contributory factors to such variations. The data are discussed in relation to the effects of smoking-associated exposure to Cd and Pb exposure on Zn availability for placental transfer, as well as on the quantitative maternal Zn supply levels to the fetus resulting from the known tendency of smokers to have lower dietary intakes of Zn. The higher Cu concentrations in OC users are discussed in relation to altered Cu metabolism, characterized by increased synthesis of the Cu-binding protein, ceruloplasmin, as an acute-phase antioxidant response to altered lipid profile and increased lipid oxidation.     

Clipperton, a possible future for atoll lagoons

Closure of the Clipperton Island atoll (10°17′ N 109°13′ W), now a meromictic lake, is estimated to have occurred between 1839 and 1849. It was still closed in 2005. Brackish waters in the upper layer (0–10 m) were oxygenated, while saline waters in the deep layer (>20 m) were anoxic. Allowing for the methodological difficulties of earlier measurements, the physical characteristics of the lagoon did not seem to have changed significantly since the last expedition (1980). The intermediate layer between brackish and saline waters was characterized by a strong density gradient and a temperature inversion of up to 1.6°C. Microbial activity, water exchange between the deep layer and surrounding oceanic waters and the geothermal flux hypothesis are discussed. The low DIN and SRP concentrations observed in the upper layer, despite high nutrient input by seabird droppings, reflect the high nutrient uptake by primary producers as attested by the elevated overall gross primary production (6.6 g C m⁻² day⁻¹), and high suspended photosynthetic biomass (2.23 ± 0.23 μg Chl a l⁻¹) and production (263 ± 27 μg C l⁻¹ day⁻¹). Phytoplankton composition changed in 67 years with the advent of new taxa and the disappearance of previously recorded species. The freshwater phytoplanktonic community comprised 43 taxa: 37 newly identified during the expedition and 6 previously noted; 16 species previously found were not seen in 2005. The closure of the lagoon, combined with the positive precipitation–evaporation budget characteristic of the region, has induced drastic changes in lagoon functioning compared with other closed atolls.     

Interactions of culture vessels, media volume, culture density, and carbon dioxide levels on lettuce and spearmint shoot growth in vitro

 The influence of culture chamber capacity, medium volume and culture density on the growth yields of lettuce (Lactuca sativa L.) and spearmint (Mentha spicata L.) shoots were determined in an environment containing either 350 or 10,000 μmol mol^–1 CO₂ after 8 weeks of incubation. High positive correlations occurred between the culture vessel capacity and spearmint fresh weight, leaf number, root number, and shoot number. Similarly, high positive correlations occurred between culture vessel capacity and lettuce fresh weight, leaf number, and root number. Higher fresh weights, leaf numbers, and root numbers were obtained from lettuce and spearmint shoots when cultured in 1-quart Mason jars containing 100- or 150-ml aliquots of medium compared to jars containing 25- or 50-ml aliquots of medium within an environment containing either 350 or 10,000 μmol mol^–1 CO₂. High culture density decreased growth yields, and this phenomenon could only be slightly off-set by the employment of an elevated CO₂ environment or larger culture vessels. Received: 22 December 1998 / Revision received: 2 July 1999 / Accepted: 12 July 1999     

Deleterious effects of fungizone on growth hormone and prolactin secretion by cultured GH3 cells

Summary The rates at which growth hormone (GH) and prolactin (PRL) are spontaneously secreted from a rat pituitary tumor cell line (GH₃) were significantly reduced when these cells were maintained in medium containing 2.5 μg/ml Fungizone (Fz). The reduction in hormone secretion was not immediately reversed by removal of Fz during perifusion, but after 3 wk in control medium, secretory rates approached the pre-Fz treatment levels. In plated cells, secretion of GH was reduced by Fz in a dose-dependent manner, whereas PRL secretion was significantly reduced only by the highest concentration (2.5 μg/ml) of Fz. We concluded that Fz is not an acceptable medium constituent for the long-term culture of GH₃ cells. However, because its effects are reversible, its short-term use as a decontaminating agent might eliminate the necessity for reinitiating the culture of cells whose secretory behavior must be followed in long-term protocols. Technical assistance provided by Y. S. Lee. Supported by grant AM33388 to M. E. S. from the National Institutes of Health, Bethesda, MD, and in part by the Medical Research Service of the Veterans Administration.     

Effect of dietary nickel and iron on the trace element content of rat liver

The level and/or form of dietary iron, dietary nickel, and the interaction between them affected the trace element content of rat liver. Livers were from the offspring of dams fed diets containing 10–16 ng, or 20 μg, of nickel/g. Dietary iron was supplied as ferric chloride (30 μg/g) or ferric sulfate (30 μg, or 60 μg). In nickel-deprived rats fed 60 μg of iron/g of diet as ferric sulfate, at age 35 days, levels of iron and zinc were depressed in liver and the level of copper was elevated. At age 55 days, iron was still depressed, copper was still elevated, but zinc also was elevated. In rats fed 30 μg of iron/g of diet as ferric chloride, liver iron content was higher in nickel-deprived than in nickel-supplemented rats at 30, but not at 50, days of age. Also manganese and zinc were lower in nickel-deprived than in nickel-supplemented rats at age 35 days if their dams had been on experiment for an extended period of time (i.e., since age 21 days). Thus, the levels of copper, iron, manganese, and zinc in liver were affected by nickel deprivation, but the direction and extent of the affects depended upon the iron status of the rat.     

Abundance of picophytoplankton in the halocline of a meromictic lake, Lake Suigetsu, Japan

Numerous (0.5 to 4.8 × 10⁵ cells/ml), small phytoplankton (smaller than 0.5–1 × 1–2 μm in cell size, picophytoplankton) were distributed in the halocline (depth 2–12 m, 4–14 practical salinity units) of the saline meromictic lake, Lake Suigetsu (35°35′ N, 135°52′ E), located in the central part of the coast of Wakasa Bay along the Japan Sea in Fukui Prefecture, Japan. Vertical distribution of phytoplankton revealed that the maximum number of picophytoplankton was always observed near or a little deeper than the oxic-anoxic boundary layer (depth 5–6 m); they were dominant phytoplankton in the water layer deeper than the oxic-anoxic boundary from July to late September 2005. Spectral analysis of autofluorescence emitted from the particle fractions smaller than 5 μm measured with a spectrofluorometer and from individual cells measured with a microscope photodiode array detector revealed that the major component of picophytoplankton was phycoerythrin-rich, unicellular cyanobacteria (picocyanobacteria). Eukaryotic phytoplankton about 2.5 μm in diameter were also found, but the numbers were low. Fluorescence intensity of chlorophyll a at 685 nm (room temperature) emitted from the particle fractions smaller than 5 μm was increased by the addition of 3-(3,4-dichlorophenyl)-1,1-dimethylurea. These observations indicated that at least some picophytoplankton had a functional photosystem II in the halocline where sulfide, the potential inhibitor of oxygenic photosynthesis, was always present. The large abundance together with their physiological potency suggest that picophytoplankton are one of the important primary producers in the halocline of Lake Suigetsu. Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users.     

Selenium and Mercury in Native and Introduced Fish Species of Patagonian Lakes,Argentina

A survey of mercury (Hg) and selenium (Se) contents was performed in fish collected from lakes located in two National Parks of the northern patagonian Andean range. Two native species, catfish (Diplomystes viedmensis) and creole perch (Percichthys trucha), and three introduced species, brown trout (Salmo trutta), rainbow trout (Oncorhynchus mykiss), and brook trout (Salvelinus fontinalis), were caught from lakes Nahuel Huapi, Moreno, Traful, Espejo Chico, and Guillelmo belonging to Nahuel Huapi National Park and from lakes Futalaufquen and Rivadavia, Los Alerces National Park. In lake Moreno, fish diet items were analyzed and rainbow trout grown in a farm. Hg and Se were measured in muscle and liver tissues by instrumental neutron activation analysis. The average concentrations in muscle of Hg for all species, ages, and lakes are between 0.4 to 1.0 μg g⁻¹ dry weight (DW) with a few fish, mainly native, exceeding the United States Environmental Protection Agency health advisory for freshwater fish limited consumption, and from 0.8 to 1.5 μg g⁻¹ DW for Se. Average concentrations in liver of Hg in all species range from 0.4 to 0.9 μg g⁻¹ DW. Brown trout, the top predator in these lakes, showed the lowest average Hg burden in both tissues. Se concentrations in the liver of brown and rainbow trout, up to 279 μg g⁻¹ DW, are higher than those expected for nearly pristine lakes, exceeding 20 μg g⁻¹ DW, the threshold concentration associated with Se toxicity. These species show lower Hg contents in muscle, suggesting a possible detoxification of Hg by a Se-rich diet. Creole perch and velvet catfish livers have lower Se concentrations, with a narrower span of values (2.3 to 8.5 μg g⁻¹ and 3.3 to 5.5 μg g⁻¹ DW respectively).