Day-Night Variations in Malate Concentration, Osmotic Pressure, and Hydrostatic Pressure in Cereus validus

Malate concentration and stem osmotic pressure concomitantly increase during nighttime CO₂ fixation and then decrease during the daytime in the obligate Crassulacean acid metabolism (CAM) plant, Cereus validus (Cactaceae). Changes in malate osmotic pressure calculated using the Van't Hoff relation match the changes in stem osmotic pressure, indicating that changes in malate level affected the water relations of the succulent stems. In contrast to stem osmotic pressure, stem water potential showed little day-night changes, suggesting that changes in cellular hydrostatic pressure occurred. This was corroborated by direct measurements of hydrostatic pressure using the Jülich pressure probe where a small oil-filled micropipette is inserted directly into chlorenchyma cells, which indicated a 4-fold increase in hydrostatic pressure from dusk to dawn. A transient increase of hydrostatic pressure at the beginning of the dark period was correlated with a short period of stomatal closing between afternoon and nighttime CO₂ fixation, suggesting that the rather complex hydrostatic pressure patterns could be explained by an interplay between the effects of transpiration and malate levels. A second CAM plant, Agave deserti, showed similar day-night changes in hydrostatic pressure in its succulent leaves. It is concluded that, in addition to the inverted stomatal rhythm, the oscillations of malate markedly affect osmotic pressures and hence water relations of CAM plants.     

In vitro effect of hydrostatic pressure exposure on hydroxyl radical production in fish red muscle

The effects of hydrostatic pressure on reactive oxygen species (ROS) production have been studied in vitro on fish red muscle fibres. In the eel, Anguilla anguilla, previous studies have shown that hydrostatic pressure acclimatization improves oxidative phosphorylation efficiency together with a supposed concomitant decrease in electron leak and ROS production. In order to test the hypothesis of an electron leak decrease under pressure, hydroxyl radical (HO*) production and oxygen consumption were measured on fish red muscle fibres directly exposed to hydrostatic pressure. Experiments were performed under two conditions--atmospheric pressure and hydrostatic pressure (16.1 MPa)--on eel and trout (which exhibit low- and high-pressure sensitivity, respectively). This work has permitted, first, the validation of an indirect HO* measurement (in vitro) on fish red muscle and the documentation of reference values for fish. Second, at atmospheric pressure, results show higher oxygen consumption for trout (+40%) than for eel which is accompanied by higher HO* production (+90%); in addition, both species present a positive relationship between HO* production and oxygen consumption. Hydrostatic pressure exposure reverses this relationship for eel but not for trout. These preliminary results only partially verify the proposed hypothesis and further experiments are needed.     

Effects of Ocean Acidification on Juvenile Red King Crab (Paralithodes camtschaticus) and Tanner Crab (Chionoecetes bairdi) Growth,Condition, Calcification,and Survival

Ocean acidification, a decrease in the pH in marine waters associated with rising atmospheric CO₂ levels, is a serious threat to marine ecosystems. In this paper, we determine the effects of long-term exposure to near-future levels of ocean acidification on the growth, condition, calcification, and survival of juvenile red king crabs, Paralithodes camtschaticus, and Tanner crabs, Chionoecetes bairdi. Juveniles were reared in individual containers for nearly 200 days in flowing control (pH 8.0), pH 7.8, and pH 7.5 seawater at ambient temperatures (range 4.4–11.9 °C). In both species, survival decreased with pH, with 100% mortality of red king crabs occurring after 95 days in pH 7.5 water. Though the morphology of neither species was affected by acidification, both species grew slower in acidified water. At the end of the experiment, calcium concentration was measured in each crab and the dry mass and condition index of each crab were determined. Ocean acidification did not affect the calcium content of red king crab but did decrease the condition index, while it had the opposite effect on Tanner crabs, decreasing calcium content but leaving the condition index unchanged. This suggests that red king crab may be able to maintain calcification rates, but at a high energetic cost. The decrease in survival and growth of each species is likely to have a serious negative effect on their populations in the absence of evolutionary adaptation or acclimatization over the coming decades.     

Why can the eel, unlike the trout, migrate under pressure

In order to elucidate the difference between pressure resistance in trout (Onchorhyncus mykiss) and eel (Anguilla anguilla), oxygen consumption of red muscle permeabilised cells and mitochondria were measured at 101 ATA hydrostatic pressure per se. Such an experiment involved the setting up of a special system allowing measurements under high pressure. The results show that hydrostatic pressure strongly alters the oxidative phosphorylation in trout but not in eel, which exhibits mitochondrial pressure resistance. It is hypothesised that the eel has a supranormal mitochondria functioning at atmospheric pressure in order to cope with the high pressure environment encountered during its migration.     

Chronic effect of atrazine on hydromineral balance in the crab

Effect in vivo of atrazine on total body weight, hydration level, haemolymph volume, inorganic electrolytes was studied in the gill, hepatopancreas and haemolymph of the crab. Significant changes were seen in the tissues indicating disturbances in the hydromineral balance of the crab as a consequence of atrazine.     

Effects of the piezo-tolerance of cultured deep-sea eel cells on survival rates, cell proliferation, and cytoskeletal structures

We investigated the pressure tolerance of deep-sea eel (Simenchelys parasiticus; habitat depth, 366–2,630 m) cells, conger eel (Conger myriaster) cells, and mouse 3T3-L1 cells. Although there were no living mouse 3T3-L1 and conger eel cells after 130 MPa (0.1 MPa = 1 bar) hydrostatic pressurization for 20 min, all deep-sea eel cells remained alive after being subjected to pressures up to 150 MPa for 20 min. Pressurization at 40 MPa for 20 min induced disruption of actin and tubulin filaments with profound cell-shape changes in the mouse and conger eel cells. In the deep-sea eel cells, microtubules and some actin filaments were disrupted after being subjected to hydrostatic pressure of 100 MPa and greater for 20 min. Conger eel cells were sensitive to pressure and did not grow at 10 MPa. Mouse 3T3-L1 cells grew faster under pressure of 5 MPa than at atmospheric pressure and stopped growing at 18 MPa. Deep-sea eel cells were capable of growth in pressures up to 25 MPa and stopped growing at 30 MPa. Deep-sea eel cells required 4 h at 20 MPa to finish the M phase, which was approximately fourfold the time required under atmospheric conditions.     

Physiological responses to emersion in the intertidal green crab,Carcinus maenas (L.)

The green shore crab, Carcinus maenas, undergoes on average 6?h periods of emersion during each low-tide cycle during the summer months. Under those conditions, the crab is cut off from its normal water environment and is exposed to potential stress from a suite of environmental and physiological changes: dehydration, compromised gas exchange and resultant internal hypoxia and hypercapnia, thermal stress, and ammonia toxicity. This study examined the comprehensive responses of the green crab in water and to a 6?h emersion period laboratory simulation of a tidal cycle followed by a 1?h re-immersion period, measuring indicators of dehydration, hemolymph osmolality, oxygen uptake, hemolymph acid–base status, heart and ventilatory rate, and hemolymph ammonia and ammonia excretion. Green crabs showed physiological responses of varying magnitude to 6?h of emersion. Individuals were found in the field exclusively under rocks and large clumps of seaweed where temperatures were approximately half those of exposed surfaces and relative humidity was about twice as high as ambient air. During emersion, crabs lost less than 5% of their wet weight, and hemolymph osmolality did not increase significantly. Oxygen uptake continued in air at about 50% of the control, aquatic values; and the gills continued to be ventilated by the scaphognathite, albeit at lower rates. Hemolymph lactate concentrations increased, indicating a shift to a greater reliance on anaerobic metabolism to support energetic needs. A slight acidosis developed in the hemolymph after 1?h of emersion, but it did not increase thereafter. Ammonia concentrations in the hemolymph were unchanged, as ammonia was volatilized by the gills and excreted into the air as NH₃ gas. These results show that the green crab copes with emersion by seeking refuge in microhabitats that mitigate the changes in the physical parameters of intertidal emersion. Physiologically, desiccation is avoided, cardio-respiratory processes are maintained at reduced levels, and hemolymph acid–base balance is minimally affected. Ammonia toxicity appears to be avoided by a shift to excreting NH₃ gas directly or indirectly to air.     

Ocean Acidification Affects Hemocyte Physiology in the Tanner Crab (Chionoecetes bairdi)

We used flow cytometry to determine if there would be a difference in hematology, selected immune functions, and hemocyte pH (pH_i), under two different, future ocean acidification scenarios (pH = 7.50, 7.80) compared to current conditions (pH = 8.09) for Chionoecetes bairdi, Tanner crab. Hemocytes were analyzed after adult Tanner crabs were held for two years under continuous exposure to acidified ocean water. Total counts of hemocytes did not vary among control and experimental treatments; however, there were significantly greater number of dead, circulating hemocytes in crabs held at the lowest pH treatment. Phagocytosis of fluorescent microbeads by hemocytes was greatest at the lowest pH treatment. These results suggest that hemocytes were dying, likely by apoptosis, at a rate faster than upregulated phagocytosis was able to remove moribund cells from circulation at the lowest pH. Crab hemolymph pH (pH_e) averaged 8.09 and did not vary among pH treatments. There was no significant difference in internal pH (pH_i) within hyalinocytes among pH treatments and the mean pH_i (7.26) was lower than the mean pH_e. In contrast, there were significant differences among treatments in pH_i of the semi-granular+granular cells. Control crabs had the highest mean semi-granular+granular pH_i compared to the lowest pH treatment. As physiological hemocyte functions changed from ambient conditions, interactions with the number of eggs in the second clutch, percentage of viable eggs, and calcium concentration in the adult crab shell was observed. This suggested that the energetic costs of responding to ocean acidification and maintaining defense mechanisms in Tanner crab may divert energy from other physiological processes, such as reproduction.     

Water transport in maize roots : measurement of hydraulic conductivity, solute permeability, and of reflection coefficients of excised roots using the root pressure probe

A root pressure probe has been used to measure the root pressure (P_r) exerted by excised main roots of young maize plants (Zea Mays L.). Defined gradients of hydrostatic and osmotic pressure could be set up between root xylem and medium to induce radial water flows across the root cylinder in both directions. The hydraulic conductivity of the root (Lp_r) was evaluated from root pressure relaxations. When permeating solutes were added to the medium, biphasic root pressure relaxations were observed with water and solute phases and root pressure minima (maxima) which allowed the estimation of permeability (P_Sr) and reflection coefficients (σ_sr) of roots. Reflection coefficients were: ethanol, 0.27; mannitol, 0.74; sucrose, 0.54; PEG 1000, 0.82; NaCl, 0.64; KNO₃, 0.67, and permeability coefficients (in 10⁻⁸ meters per second): ethanol, 4.7; sucrose, 1.6; and NaCl, 5.7. Lp_r was very different for osmotic and hydrostatic gradients. For hydrostatic gradients Lp_r was 1·10⁻⁷ meters per second per megapascal, whereas in osmotic experiments the hydraulic conductivity was found to be an order of magnitude lower. For hydrostatic gradients, the exosmotic Lp_r was about 15% larger than the endosmotic, whereas in osmotic experiments the polarity in the water movement was reversed. These results either suggest effects of unstirred layers at the osmotic barrier in the root, an asymmetrical barrier, and/or mechanical effects. Measurements of the hydraulic conductivity of individual root cortex cells revealed an Lp similar to Lp_r (hydrostatic). It is concluded that, in the presence of external hydrostatic gradients, water moves primarily in the apoplast, whereas in the presence of osmotic gradients this component is much smaller in relation to the cell-to-cell component (symplasmic plus transcellular transport).     

Potassium Flux and Leaf Movement in Samanea saman : II. Phytochrome Controlled Movement

Phytochrome, a membrane-localized biliprotein whose conformation is shifted reversibly by brief red or far-red light treatments, interacts with the rhythmic oscillator to regulate leaflet movement and potassium flux in pulvinal motor cells of Samanea. Darkened pinnae exposed briefly to red light (high P_fr level) have less potassium in motor cells in the extensor region, more potassium in motor cells in the flexor region, and smaller angles than those exposed to far-red light (low P_fr level). Increase in temperature from 24° to 37° increases the differential effect of the light treatments during opening (the energetic phase) but not during closure, implying that phytochrome controls an energetic process. It seems likely that phytochrome interacts with rhythmically controlled potassium pumps in flexor and extensor cells. During nyctinastic closure of white-illuminated pinnae, exposure to far-red light before darkening results in larger angles than does exposure to red. As in rhythmic opening, the angles of all pinnae and the differential effect of the light treatments increases with increasing temperature.     

Metabolic effects of CO2 anaesthesia in Drosophila melanogaster

Immobilization of insects is necessary for various experimental purposes, and CO₂ exposure remains the most popular anaesthetic method in entomological research. A number of negative side effects of CO₂ anaesthesia have been reported, but CO₂ probably brings about metabolic modifications that are poorly known. In this work, we used GC/MS-based metabolic fingerprinting to assess the effect of CO₂ anaesthesia in Drosophila melanogaster adults. We analysed metabolic variation of flies submitted to acute CO₂ exposure and assessed the temporal metabolic changes during short- and long-term recovery. We found that D. melanogaster metabotypes were significantly affected by the anaesthetic treatment. Metabolic changes caused by acute CO₂ exposure were still manifested after 14 h of recovery. However, we found no evidence of metabolic alterations when a long recovery period was allowed (more than 24 h). This study points to some metabolic pathways altered during CO₂ anaesthesia (e.g. energetic metabolism). Evidence of short-term metabolic changes indicates that CO₂ anaesthesia should be used with utmost caution in physiological studies when a short recovery is allowed. In spite of this, CO₂ treatment seems to be an acceptable anaesthetic method provided that a long recovery period is allowed (more than 24 h).     

The synergism between hydrocarbon pollutants and UV radiation: a potential link between coastal pollution and larval mortality

Coastal, benthic invertebrates with complex life history strategies are exposed to stage- and habitat-specific selective forces. In the coastal environment, benthic adults are exposed to polycyclic aromatic hydrocarbon pollutants (PAHs) due to their proximity to human activities (shipping, urbanization, and industrialization). Benthic invertebrates produce lipid-rich eggs or larvae that absorb PAHs from polluted estuaries and coastal waters. The larvae of many coastal invertebrates move offshore following release from benthic adults. During development in offshore waters, larvae of some species are exposed to relatively high levels of ultraviolet (UV) radiation. Marine organisms vary in their tolerance to PAHs and UV radiation. The purpose of this study was to examine the effects of the sequential exposure of the larvae of marine crabs to PAHs and UV radiation.Using laboratory experiments, the larvae of four crab species were exposed to PAHs and UV radiation. There was a significant synergistic effect of exposure to PAH (fluoranthene or pyrene) and UV radiation on larvae of the spider crab (Libinia dubia), the stone crab (Menippe adina) and the mud crab (Panopeus herbstii). Larvae of blue crabs (Callinectes sapidus) were exposed to PAHs and UV radiation in both laboratory and solar UV experiments. Significantly higher mortality occurred for C. sapidus larvae using either type of UV-artificial or solar.Larvae of coastal invertebrates with complex life history strategies are susceptible to the combined effects of PAHs and UV radiation. In this study, the exposure of crab larvae to PAHs and UV radiation resulted in mortality to crab larvae using laboratory and solar UV experiments. There were no effects on larval crab mortality due to PAH or UV radiation independently but mortality was as high as 100% when both factors were present.     

A review of glass eel migratory behaviour,sampling techniques and abundance estimates in estuaries: implications for assessing recruitment,local production and exploitation

Recruitment of the three northern hemisphere eel species (European eel Anguilla anguilla, American eel Anguilla rostrata and Japanese eel Anguilla japonica) has reduced significantly over the past thirty-five years. The stock of the European eel is described as being outside safe biological limits, with urgent action required by European Union Member States to assist recovery of the panmictic stock. Stock recruitment models and estimates of silver eel output from a river catchment are strongly influenced by the degree of certainty in estimating key population parameters of each life history stage. Therefore, management decisions aimed at enhancing eel populations rely on sound scientific evidence, based upon a fundamental understanding of the complex anguillid eel life cycle. This review paper focuses on the estuarine entry phase of the eel life cycle and synthesises the current scientific knowledge with regard to glass eel migratory behaviour, sampling methods and abundance estimates within estuaries. Although the behavioural and environmental processes modulating glass eel migration patterns are reasonably well understood, site specific factors play a significant role in determining fine scale distribution patterns at an individual estuary level. Given the large resource commitment required to adequately sample this key life history stage, behavioural studies of migration patterns on a local scale are crucially important to aid the design of robust sampling programmes aimed at quantifying seasonal abundance and annual recruitment.     

Hypothermal effects on survival,energy homeostasis and expression of energy-related genes of swimming crabs Portunus trituberculatus during air exposure

Previously, dry or semi-dry approach under the hypothermal condition is proved to be an alternative method in transport of live swimming crabs Portunus trituberculatus. However, we wondered whether this method can improve crab survival when temperature is kept as cool as possible. In this study, we hypothesized that there is a thermal threshold below which dry or semi-dry approach (air exposure) could cause crab physiological disruption and therefore aggravate their mortality. To test the above hypothesis, crabs (23 °C) were exposed to air at temperatures ranging from 4 to 16 °C. Results showed that crabs had a worse survival and vigor at temperatures below 12 °C. Then we tested crab energy metabolism to explore the possible reason. It was shown that total adenine nucleotide and adenylate energy charge in gills were remarkably reduced by air exposure of below 12 °C. This increased the need for crabs to re-balance energy metabolism, which was indicated by the upregulation of AMPKα and HIF-1α. Meanwhile, there was a significant increase of the expression of Na⁺/K⁺-ATPase, V-type ATPase and HSP90 at temperatures below 12 °C, while all treatments shared a similar level of hemocyanin, urate and lactate in hemolymph and expression of cytochrome c oxidase and NADH-ubiquinone reductase in gills. These results implied that dry or semi-dry approach below 12 °C could exert detrimental effects on P. trituberculatus, and perturbation of energy homeostasis, which is more related with changes of energy-demanding physiological pathways, is a possible reason of crab death and poor vigor.     

Cell-Specific Thioautotrophic Productivity of Epsilon-Proteobacterial Epibionts Associated with Shinkaia crosnieri

In this study, we report experimental evidence of the thioautotrophic activity of the epibiotic microbial community associated with the setae of Shinkaia crosnieri, a galatheid crab that is endemic to deep-sea hydrothermal systems in the Okinawa Trough in Japan. Microbial consumption of reduced sulfur compounds under in situ hydrostatic and atmospheric pressure provided evidence of sulfur-oxidizing activity by the epibiotic microbial community; the rate of sulfur oxidation was similar under in situ and decompressed conditions. Results of the microbial consumption of reduced sulfur compounds and tracer experiments using ¹³C-labeled bicarbonate in the presence and absence of thiosulfate (used as a thioautotrophic substrate) convincingly demonstrated that the epibiotic microbial community on S. crosnieri drove primary production via an energy metabolism that was coupled with the oxidation of reductive sulfur compounds. A combination of tracer experiments, fluorescence in situ hybridization (FISH) and nano-scale secondary ion mass spectrometry (Nano-SIMS) indicated that the filamentous cells of the genus Sulfurovum belonging to the class Epsilonproteobacteria were thioautotrophs in the epibiotic community of S. crosnieri. In conclusion, our results strongly suggest that thioautotrophic production by Sulfurovum members present as the epibiotic microbial community play a predominant role in a probable nutritional ectosymbiosis with S. crosnieri.     

An energetic and conceptual model of the physiological role of dietary carbohydrates and salinity on Litopenaeus vannamei juveniles

We are reporting results directed to explain the relation between carbohydrates (CHO), protein metabolism, and the energetic balance of Litopenaeus vannamei juveniles. The interaction of dietary CHO and salinity was measured to try to understand the relation between osmotic control and metabolism, both from a biochemical and energetic point of view. Two experiments were done. In the first experiment, shrimp were fed with 0%, 5%, 33%, and 61% CHO and maintained at 15‰ and 40‰ salinity. Glucose, lactate protein, hemocyanin, ammonia concentration, and osmotic pressure were measured in blood. Digestive gland glycogen (DGG) was measured also. In the second experiment, shrimp were fed with 0% and 38% dietary CHO and maintained at 15‰ and 40‰ salinity. From that shrimp, absorbed energy (Abs) was calculated as: Abs=respiration (R)+ammonia excretion (U) and production (P); assimilated energy (As) was calculated as the product of R×P. Osmotic pressure, hemocyanin, protein, lactate, and blood ammonia increased with the reduction in dietary CHO. In contrast, an increase in blood glucose was observed with an increase in dietary CHO. Digestive gland glycogen (DGG) increased following a saturation curve with a DGG maximum at 33% dietary CHO. Blood metabolites of fasting and feeding shrimp showed the same behavior. Energy balance results showed that shrimp maintained in low salinity and fed without CHO waste more energy in U production than for shrimp maintained in high salinity and fed with high CHO levels. Notwithstanding, the production efficiency was higher in shrimp fed without CHO than that observed in shrimp fed with high CHO independent of salinity. A scheme trying to integrate the relation between CHO and protein metabolism and the way in which both are modulated by salinity is presented. From published and present results, there are two factors that apparently control the use of high dietary CHO levels; α-amylase enzyme-dietary CHO level capacity and glycogen saturation in DG. Production of glucose is limited in shrimp because of saturation of α-amylase when shrimp are fed with diets above 33% CHO. This is the first control point of starch metabolism. The digestive gland is saturated with glycogen in shrimp fed with dietary CHO levels >33%. This is apparently the second control point of CHO metabolism that limits growth rate in such conditions. The high metabolic cost related to high CHO diets could explain why shrimp are well adapted to use protein as a source of energy.     

Effects of High Hydrostatic Pressure on Coastal Bacterial Community Abundance and Diversity

Hydrostatic pressure is an important parameter influencing the distribution of microbial life in the ocean. In this study, the response of marine bacterial populations from surface waters to pressures representative of those under deep-sea conditions was examined. Southern California coastal seawater collected 5 m below the sea surface was incubated in microcosms, using a range of temperatures (16 to 3°C) and hydrostatic pressure conditions (0.1 to 80 MPa). Cell abundance decreased in response to pressure, while diversity increased. The morphology of the community also changed with pressurization to a predominant morphotype of small cocci. The pressure-induced community changes included an increase in the relative abundance of Alphaproteobacteria, Gammaproteobacteria, Actinobacteria, and Flavobacteria largely at the expense of Epsilonproteobacteria. Culturable high-pressure-surviving bacteria were obtained and found to be phylogenetically similar to isolates from cold and/or deep-sea environments. These results provide novel insights into the response of surface water bacteria to changes in hydrostatic pressure.