The effect of copper and zinc on the metabolism of the mussel Mytilus edulis

The effects of copper and zinc on the metabolism of the mussel Mytilus edulis (L.) and its component tissues were studied. 500 ppm copper sodium citrate inhibited oxygen consumption of the whole animal and gill tissue, but no similar effect was observed on digestive gland tissue. 500 ppm zinc sodium citrate exerted no effect upon gill or digestive gland respiration, and neither metal salt affected the respiration of homogenates of gill, digestive gland or gonad. Direct observation of gill tissues during exposure to the metals revealed that 500 ppm copper sodium citrate caused inhibition of ciliary activity; exposure of tissues to 2 ppm Cu for 24 h resulted in only partial inhibition of the cilía. It is suggested that metabolic suppression noted in whole animals and gill tissues is due to the inhibition of an energy-consuming process such as ciliary activity rather than interference with respiratory enzyme systems.     

Effect of sexual maturation on the tissue biochemical composition of<Emphasis Type="Italic"> Octopus vulgaris</Emphasis> and<Emphasis Type="Italic"> O</Emphasis>.<Emphasis Type="Italic"> defilippi</Emphasis> (Mollusca: Cephalopoda)

Changes in the protein, lipid, glycogen, cholesterol and energy contents, total amino acid and fatty acid profiles of Octopus vulgaris and O. defilippi tissues (gonad, digestive gland and muscle) during sexual maturation (spermatogenesis and oogenesis) were investigated. Both species showed an increase of amino acids and protein content in the gonad throughout sexual maturation (namely in oogenesis), but allocation of these nitrogen compounds from the digestive gland and muscle was not evident. The major essential amino acids in the three tissues were leucine, lysine and arginine. The major non-essential amino acids were glutamic acid, aspartic acid and alanine. With respect to carbon compounds, a significant increasing trend (P<0.05) in the lipid and fatty acid contents in the three tissues was observed, and, consequently, there was also little evidence of accumulated lipid storage reserves being used for egg production. It seems that for egg production both Octopus species use energy directly from food, rather than from stored products. This direct acquisition model contrasts with the previous model for Octopus vulgaris proposed by ODor and Wells (1978: J Exp Biol 77:15–31). Most of saturated fatty acid content of the three tissues was presented as 16:0 and 18:0, monounsaturated fatty acid content as 18:1 and 20:1 and polyunsaturated fatty acid content as arachidonic acid (20:4n-6), eicosapentaenoic acid (20:5n-3) and docosahexaenoic acid (22:6n-3). Though cholesterol is an important precursor of steroid hormones, this sterol content exhibited variations that do not seem to be related with the maturation process. Moreover, significant differences (P<0.05) were obtained between genders, suggesting that perhaps there is a greater physiological demand for cholesterol during spermatogenesis than oogenesis. If the component sterols of octopus are of a dietary origin, considerable variation in the cholesterol content between species might be expected on the basis of the sterol composition of their prey. The glycogen reserves increased significantly in the gonad and decreased significantly (P<0.05) in the digestive gland and muscle of O. vulgaris (these trends were not evident in O. defilippi). Glycogen may play an important role in the maturation process and embryogenesis of these organisms, because carbohydrates are precursors of metabolic intermediates in the production of energy. It was evident that sexual maturation had a significant effect upon the gonad energy content, but the non-significant energy variation (P>0.05) in the digestive gland and muscle revealed no evidence that storage reserves are transferred from tissue to tissue. The biochemical composition of digestive gland and muscle may not be influenced by sexual maturation, but rather by other biotic factors, such as feeding activity, food availability, spawning and brooding.Communicated by S.A. Poulet, Roscoff     

Energy maximization by selective feeding on tissues of the venerid clam <Emphasis Type="Italic">Marcia hiantina</Emphasis> in the marine scavenger <Emphasis Type="Italic">Nassarius festivus</Emphasis> (Gastropoda: Nassariidae)

Feeding selectivity of the intertidal scavenging gastropod Nassarius festivus on four types of tissue (soft tissue, adductor muscle, foot, mantle margin) of the venerid clam Marcia hiantina was studied by field observations and laboratory experiments. Both approaches showed similar results with the soft tissue and adductor muscle being much preferred. As the energy content of the four types of tissue was similar and individuals of N. festivus rearing in different types of tissue obtained similar growth rates both in shell length and in body wet weight, differential consumption was most likely determined by the palatability of the tissues. Energy intake in a meal for N. festivus was estimated at 10.92, 9.17, 3.86 and 2.02 cal meal⁻¹ ind⁻¹ for soft tissue, adductor muscle, mantle margin, and foot, respectively. In view of the small size and sporadic distribution of the carrion, and intense competition among the conspecifics, selective feeding on more palatable tissues maximizes energy intake for scavengers like N. festivus.     

Lactate formation in Callianassa californiensis and Upogebia pugettensis (Crustacea: Thalassinidea)

The ability to accumulate lactate as a result of laboratory exposure to anoxia was examined in 2 species of mud-dwelling shrimp, the ghost shrimp Callianassa californiensis and the mud shrimp Upogebia pugettensis. Hemolymph lactate accumulated to a much greater degree in the mud shrimp, even though the overall levels of lactate dehydrogenase activity were similar in tissues of the two species. Carbohydrate reserves, estimated as glycogen, were not significantly depleted in muscle or midgut gland of either species as a result of anoxic stress. The results are discussed in relation to possible metabolic strategies employed to cope with the hypoxic environment of these crustaceans.     

Concentration and distribution of heavy metals in tissues of two cephalopods,Eledone cirrhosa and Sepia officinalis,from the French coast of the English Channel

The concentrations of 11 heavy metals (Ag, Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, V and Zn) were measured in the tissues (digestive gland, branchial hearts, gills, digestive tract, kidney, genital tract, muscle, skin, shell) of the two cephalopods Eledone cirrhosa (d'Orb.) and Sepia officinalis (L.) collected from the French coast of the English Channel in October 1987. The tissues of both species displayed a similar pattern of heavy-metal accumulation: the digestive gland, branchial hearts and kidney were the major sites of concentration for all 11 metals; the digestive gland accumulated silver, cadmium, cobalt, copper, iron, lead and zinc, the branchial hearts high concentrations of copper, nickel and vanadium, and the kidney high concentrations of manganese, nickel and lead. The digestive gland, which constituted 6 to 10% of the whole-animal tissue, contained >80% of the total body burden of Ag, Cd and Co and from 40 to 80% of the total body burden of the other metals. The ratios between heavy metal concentrations in the digestive gland and those in the muscle separated the elements into three groups, those with a ratio 10 (Cr, Mn, Ni, Pb, V, Zn), those with a ratio >10 to <50 (Co, Cu, Fe), and those with a ratio 50 (Ag, Cd). The digestive gland of cephalopods (carnivorous molluscs whose age can be easily calculated with great accuracy) would seem to constitute a good potential indicator of heavy metal concentrations in the marine environment.     

Lipid-class composition of organs and tissues of the tiger prawnPanaeus esculentus during the moulting cycle and during starvation

Neutral lipid and phospholipid fractions and their component lipid classes in the digestive gland, abdominal muscle, epidermis and cuticle ofPenaeus esculentus Haswell were analysed and compared during the moulting cycle and during starvation. The prawns were collected from Moreton Bay, Queensland, Australia, by trawling during 1985–1987, and were fed on a standard, semi-purified diet. The digestive gland appears to be a major site of lipid synthesis, storage and mobilisation in preparation for moulting. Neutral lipid, 59 to 80% of which was triacylglycerol, was the larger fraction. It accumulated during early premoult, mainly due to the increase in triacylglycerol. The digestive gland contained only 18% of the total body lipid, or 8% of body lipid as triacylglycerol. Thus, the reserve lipid available for energy production is very small. Digestive gland triacylglycerol was markedly depleted after 4 d starvation and was almost completely absent after 8 d. In the other tissues, the major fraction was phospholipid, of which over 50% was phosphatidylcholine and up to 20% phoshatidylethanolamine; cholesterol was the major class in the neutral lipid fraction and appeared to be very stable. Most of this lipid was probably a component of cellular membranes. The lipid composition of muscle changed very little during the moulting cycle: total lipid levels in the epidermis were high in late premoult and early postmoult, when new cuticle is being secreted, but the proportions of the component lipids were closely similar. Cuticle lipid, together with other major components, was resorbed from the old cuticle prior to ecdysis, but the cuticle phospholipids appeared to be labile at all moult stages. The total of all lipids in fedP. esculentus was about 3.6% dry weight, of which about 70% was phospholipid. Earlier research had shown that when digestive gland lipid is exhausted after a short period of starvation, muscle is metabolised for energy. The present research showed that in the remaining muscle only about 13% of lipid was lost after 21 d starvation, mostly as phosphatidylcholine. This is in keeping with the need to maintain this tissue in a functional state. In contrast, epidermal lipid levels were markedly reduced after only 4 d starvation and the proportions of phospholipids changed significantly. This sensitivity of the cuticle lipids to starvation may be the cause of delayed moulting, which is characteristic of poor nutrition.     

Mechanisms of copper toxicity in the shore crab,Carcinus maenas

The influence of copper(II)chloride on selected key enzymes and metabolites in intermediary metabolism and the energy charge potential of the hyperregulating shore crab, Carcinus maenas (L.) were investigated. Crabs, collected in Odense Fjord, Denmark between September 1989 and May 1990, were exposed to 10 ppm copper(II)chloride, in 10 ppt salinity seawater at 15°C for up to 1 wk. Hexokinase activity was 77% lower in the posterior gills and 60% lower in the midgut gland than in control crabs. Phosphofructokinase activity was reduced in the midgut gland by 82% but was unaffected in the gills. Pyruvate kinase activity was undetectable in the gills of exposed crabs and reduced by 47% in the midgut gland. Citrate synthase activity was depressed by 30% in the posterior gills and unaffected in other tissues. Copper exposure had no apparent effect on tissue cytochrome c oxidase activity. Chela muscle was the only tissue in which copper exposure did not bring about reductions in the enzyme activities tested. As a result of 1 wk of copper exposure the average lactate level increased by 3.5-fold in haemolymph, 9.3-fold in posterior gills and 6.5-fold in midgut gland. The average glucose level was raised by a factor of 6 in the haemolymph, 3.4 in the posterior gills and 1.2 in the midgut gland. Energy charge potential was at a very high level (ca. 0.9) and was unaffected by copper exposure.     

Patterns of trace-metal distribution in tissues of Pleoticus muelleri (Crustacea: Decapoda: Solenoceridae)

Trace-metal distribution in tissues of the shrimp Pleoticus muelleri Bate from the Patagonian region, Argentina, was related to sex, size and physiological condition. Concentrations of cadmium, copper, manganese and zinc were determined in the digestive gland, male reproductive system and muscle of adult specimens. Significant (p<0.01) sex-dependent differences in mean metal concentrations (g g^-1 wet wt) were found in the following tissues of males and females, respectively: digestive gland, Cu, 82.9 and 30.8; Zn, 32.5 and 44.9; reproductive system, Zn, 12.8 and 38.6; Mn, 1.7 and 3.1; Cd, 0.29 and 0.58. Metal levels of muscle showed no significant differences between sexes. Cadmium was not detected in muscle, suggesting that its incorporation into this tissue was strictly regulated. Metal concentrations in the male reproductive system were in general weakly correlated (0.001<p<0.05) with either body size or the concentration of metals in the digestive gland. The developing ovary incorporated substantial amounts of zinc, while cadmium and copper levels decreased at proportional rates (p<0.001). The patterns of the variations in the concentrations of these metals in the ovary strongly suggested that a regulatory mechanism operated throughout oogenesis. Manganese was not involved in this presumptive mechanism. The results of this study are discussed in terms of concentrations and in absolute amounts for standardized individuals.     

Seasonal variations in biochemical constituents during the reproductive cycle of the female dog cockle Glycymeris glycymeris

In an attempt to describe the biochemical events associated with the main stages of the annual and reproductive cycles of the female dog cockle Glycymeris glycymeris L., we studied seasonal variations in the various stages of oocyte development of the ovaries, and the glycogen, total protein and total lipid content of five body tissues – adductor muscle, foot, tunic coat, visceral mass and mantle. From November 1991 to November 1994, microscopic examination of the ovaries and measurement of the tissue concentrations of glycogen, total proteins and total lipids in these five body tissues were made monthly on ten female dog cockles originating from the sea area around Douarnenez (south Brittany, France). Morphological studies revealed that in the population investigated the annual cycle is characterised by three major periods: a first period of vitellogenesis extending from February/March to April/May and preceding a spawning in spring; a second period of vitellogenesis extending from May/June to September/October and leading to either no spawning, a single autumnal spawning event, or to two spawning events in summer and autumn; and a third period extending from October/November to February/March and characterised by a high level of oocyte lysis. In the muscular body tissues of the dog cockle, i.e. the adductor muscles, the foot and the tunic coat (the muscular envelope containing the visceral mass), the concentrations of glycogen, total proteins and total lipids underwent very similar variations during the annual cycle. During each stage of vitellogenesis, a typical glycogen–protein–lipid sequence was observed in the muscular tissues that was characterised firstly by a peak of glycogen concentration 2 to 3 mo before spawning, followed by a peak in total proteins 1 mo before spawning, and finally by a peak in lipid content just before spawning. A similar glycogen–protein–lipid sequence was also recorded in the first half of the winter period. However, these events were followed by general atresia affecting all oocytes in the gonad. Maximum energetic value of biochemical constituents in females coincided with peaks in lipid content in the visceral mass and mantle. These biochemical events occurred principally immediately before and at the end of oocyte lysis (December/January). A drop in the total energetic value, affecting mainly the visceral mass and the mantle, was recorded each year during the period January to March, coinciding with the period of shell growth in this species. Our data clearly indicate that in female G. glycymeris all muscular tissues contribute to the storage of glycogen and proteins, and suggest that glycogen may be the source of energy triggering vitellogenesis. Biochemical and microscopic observations revealed that oocyte development takes place during the first half of winter, but that these oocytes undergo atresia in December/January. The metabolites produced from oocyte lysis could contribute to somatic growth, which occurs in late winter. Received: 3 March 1997 / Accepted: 23 July 1997     

Effects of salinity stress on neurotransmission, energy metabolism, and anti-oxidant biomarkers of Carcinus maenas from two estuaries of the NW Iberian Peninsula

This study investigated the effects of salinity on biomarkers of oxidative stress, energy metabolism, and neurotransmission of Carcinus maenas from an estuary low impacted by pollution and from an estuary under chemical stress in the NW Iberian Peninsula. Crabs were collected in the field and, following an acclimation period, they were exposed for 7 days to five salinity levels ranging from 4 to 45 psu. At the end of the exposure period, stress biomarkers were determined in samples of muscle and digestive gland. The biomarkers assessed in the muscle were the activities of the enzymes cholinesterases (ChE), of which acetylcholinesterase is involved in neurotransmission, and lactate dehydrogenase (LDH) and isocitrate dehydrogenase (IDH) that are involved in energy metabolism. The biomarkers assessed in the digestive gland were (1) the activities of the enzymes glutathione S-transferases (GST), glutathione reductase (GR), and glutathione peroxidase (GPx), involved in phase II biotransformation and the anti-oxidant defence system; (2) the levels of total glutathiones (TG), also belonging to the anti-oxidant system; and (3) the levels of lipid peroxidation as a measure of oxidative damage. The results showed a significant influence of salinity on neurotransmission, energy metabolism, anti-oxidant status, and oxidative damage of C. maenas. For some biomarkers, this influence was dependent on whether the crabs were collected at the low-polluted estuary or at the contaminated estuary. In particular, crabs collected at the low-polluted estuary showed altered neurotransmission and anti-oxidant defences (GR). Crabs collected at the impacted estuary showed alterations in neurotransmission, energy metabolism (IDH and LDH), biotransformation, and anti-oxidant defences (GST, GR, GPx, and TG), as well as in oxidative damage, indicating that salinity change superimposes higher stress on these organisms. For ChE, IDH, and TG, altered responses were induced by both hypo- and hypersalinity.     

Effect of dissolved oxygen level on respiratory metabolism, nutritional physiology, and immune condition of southern king crab Lithodes santolla (Molina, 1782) (Decapoda, Lithodidae)

Episodes of hypoxia are common in the marine environment, and their ecological effects depend, in part, on their severity and duration. Many species of decapod crustaceans reside in areas with fluctuating oxygen regimens. Physiological mechanisms enhance the ability of these crustaceans to cope with acute episodes of hypoxia. Southern king crab, Lithodes santolla, fishery is important in the south of South America, and some data describe fishing zones with low dissolved oxygen (DO) levels (3.5 mgO₂ l⁻¹, i.e., 8.3 kPa). Our main objective was to evaluate the effect of dissolved oxygen level on respiratory metabolism, nutritional physiology, and immunological condition of L. santolla juveniles. Individual animals were exposed for 10 days to different oxygen tensions (2.1, 4.2, 8.5, 12.7, and 21.1 kPa) to quantify the oxygen consumption rate; thereafter, blood oxyhemocyanin (Hc), protein concentration, as well as hemocytes, were sampled. Freeze-dried animals were dissected, and digestive gland metabolites (glycogen, protein, glucose, cholesterol, acylglycerol, and lactate) and digestive enzyme activity (general protease, trypsin, and chymotrypsin), as well as gill lactate dehydrogenase (LDH) activity, were quantified. In the present study, Lithodes santolla showed a critical oxygen tension between 4 and 9 kPa, indicating that this crab species is more sensitive to DO than other crustacean species. Protein and Hc concentrations followed a similar pattern to that of oxygen consumption. Digestive gland glycogen and protein concentration did not change after 10 days at different oxygen exposures, but glucose, cholesterol, and acylglycerol concentrations decreased linearly and proportionally to the available oxygen in the water. As in other decapods, chymotrypsin showed over 90% of the total quantified proteases activity. Chymotrypsin activity together with total proteases and trypsin was not affected by the environmental oxygen tension. Gill LDH and digestive gland lactate followed a similar increase at lower environmental oxygen tension but dropped sharply at the lowest tension (2.1 kPa). Dissolved oxygen affected also the immune system through reduction of hemocytes. This could provide a critical window for opportunistic pathogens to become established when crabs are exposed to hypoxic conditions. L. santolla juveniles show a moderate tolerance to low oxygen availability by modifying the concentration of hemolymph proteins, mainly OxyHc, some digestive gland metabolites, and by activating the anaerobic metabolism. This allows L. santolla juveniles to inhabit temporarily low oxygen zones in the deep ocean and suggests an advantage for culture conditions.     

Acid and alkaline phosphatase activities in the clam Ruditapes philippinarum

Acid and alkaline phosphatase activities have been partially characterized in Ruditapes philippinarum (Adams and Reeve, 1850). Two activity peaks at pH=4.5 and pH 10.5 were detected in the gill, digestive gland, mantle, siphon and foot. Acid phosphatase activity was higher than that of alkaline phosphatase. The highest activity for both enzymes was observed in the digestive gland and, in decreasing order, the gill, foot, siphon and mantle. Alkaline phosphatase activity was similar in the mantle, siphon and foot. K _m values were determined for both enzymes in the gill and digestive gland. Hill coefficients were near 1, indicating no allosteric behaviour for either enzyme in the two organs. The optimum temperature was the same for acid phosphatase in both gill and digestive gland (50 °C), while for alkaline phosphatase it differed for these two organs (gill, 40 °C; digestive gland, 35 °C). The apparent activation energy was obtained from Arrhenius plots, and ranged from 8.61 kcal/mol for alkaline phosphatase in the gill, to 10.84 kcal/mol for acid phosphatase in the digestive gland. The effects of metals (1 mM conc) on both enzyme activities were assayed in vitro. Hg strongly inhibited the enzyme activities in the gill and digestive gland, probably because of its affinity for the sulphydryl group. Histochemically, acid phosphatase in the gill was located in a granular form throughout the gill cells, but was undetectable in the ciliate epithelium of the gill filaments. Alkaline phosphatase was located in the gill skeleton. Clam size and phosphatase activities were inversely related, probably reflecting a decrease in shell deposition with inereasing size. As a function of season, both enzymes were present in lowest amounts in winter, when undifferentiated sex cells were predominant in the germinative epithelium, and highest in summer, when ripe individuals of both sexes were more frequent.     

Histoenzymological study on the toxicity of copper sulphate in the digestive glands of Lymnaea luteola

During 24 and 48 hr of exposure, the digestive glands of Lymnaea treated with a lethal concentration of 0.038 mgl(-1) CuSO4 revealed intense activity of acid phosphatase in perilobular margin. On the other hand, same area of the gland showed moderate activity of ATPase during 24 and 48 hr of exposure. However, alkaline phosphatase showed average activity in perialveolar region and perilobular margin during 24 and 48, and 72 hr of exposure respectively The changes in the activity of these enzymes were nonsignificant in alveolar margin and perialveolar region of the gland. It is interesting to note moderate activity of acid phosphatase in perialveolar region during 24 hr of exposure.     

Phenotypic flexibility in response to environmental salinity in the euryhaline crab Neohelice granulata from the mudflat and the saltmarsh of a SW coastal lagoon

This study constitutes a first attempt to investigate intraspecific differences in osmoregulatory capacity and digestive and metabolic responses at the biochemical level in relation to hyper- and hypo-regulation in a single species of estuarine crab inhabiting contrasting habitats within a same intertidal area. We compared hemolymph osmolality, key digestive enzymes, glycemia and energy reserves in Neohelice granulata (Dana in Proc Acad Nat Sci Philadelphia 5:247–254, 1851) from the mudflat and saltmarsh of Mar Chiquita coastal lagoon (37°32′/37°45′S-57°19′/57°26′W) under a wide range of salinities (6–60 psu). Individuals from both sites exhibited high and similar osmoregulatory capacity, but while in individuals from mudflat low and high salinities affected lipase activity in hepatopancreas and triglycerides in muscle, in crabs from saltmarsh, high salinities affected glycogen in anterior gills. Low salinity differentially affected free glucose in anterior gills. The results suggest the occurrence of intraspecific distinct digestive and metabolic adjustments in relation to osmoregulatory responses and habitat.     

Petroleum hydrocarbons in the marine bivalve Venus verrucosa: accumulation and cellular responses

Cellular and subcellular responses of the marine burrowing bivalve Venus verrucosa collected from the north-eastern coastline of Malta from January to June 1985, after exposure to petroleum hydrocarbons (PHC) were investigated. After long-term exposure to 100 gl^-1 of water-accomodated fractions (WAF) of crude oil, PHC were found to accumulate most rapidly in the digestive gland and then in the gills, with saturation levels being reached within 100 d of exposure in both cases. PHC accumulation, both in the mantle and muscle tissues, was more gradual and consistent throughout the whole exposure period. After 150 d of exposure, the digestive cells of the digestive gland were significantly reduced in height (atrophy) and exhibited reduced lysosomal membrane stability. After 144 h of exposure to higher concentration of PHC (820 and 420 gl^-1), several cytological effects were recorded, including an increase in cell volume and activity of gill mucocytes as well as in the number of haemocytes in gill blood sinuses. There was also evidence of damage to the epithelial lining of the foot, stomach and style sac and marked atrophy of the digestive cells of the digestive gland. The significance of such responses is discussed.     

Tissue and sub-cellular distribution of Fe,Cu, Zn and 210Po in the abalone Haliotis rubra

The tissue and sub-cellular distribution of Fe, Cu, Zn and the naturally occurring radionuclide polonium-210 was determined in the gastropod mollusc Haliotis rubra collected from Western Port Bay, Australia, between March and July 1988. The highest concentrations of the metals, with the exception of Cu, were found in the digestive gland. Copper was more uniformly distributed, with tissues that are more vasculated having higher concentrations. Ultrastructural examination of the digestive gland, gill and kidney showed dense membrane-bound granules within the cytoplasm. Elemental analysis of the granules by electron probe x-ray microanalysis indicated that the granules in the digestive gland and gill contained high concentrations of iron, with small amounts of copper and zinc. In contrast, the metal-containing granules in the kidney were predominantly composed of iron, copper and phosphorus, with variable contributions of sodium potassium, and calcium. Homogenisation and fractionation of the digestive gland by differential centrifugation confirmed that approximately 80, 10, 90 and 50% of the total homogenate Fe, Cu, Zn and ²¹⁰Po, respectively, sedimented at 1200xg. In the haemolymph, all the elements studied were associated with the soluble high molecular weight component of the serum, not with the amoebocytes. ²¹⁰Po was present in the mucus-secreting hypobranchial glands at about half the concentration found in gill tissue.     

Biochemical and histopathological changes of Biomphalaria alexandrina snails exposed to bisphenol A

The effects of sub-lethal concentrations of bisphenol A on some biochemical parameters in Biomphalaria alexandrina snail's hemolymph and tissues as well as on histological changes of the hermaphrodite gland were evaluated. Adult snails were exposed to sub-lethal concentrations for four weeks and samples of snails were investigated after two and four weeks. Total protein and albumin levels in hemolymph, as well as lipid peroxidation and glutathione contents of snails’ tissues, were measured. After two weeks of exposure, total protein content and albumin levels decreased in all groups. Elevation of lipid peroxidation levels was correlated with decreasing glutathione content in the treated snails. Histological examination of the hermaphrodite gland revealed deformations in eggs and affected sperm production.     

Trace metal concentrations and contents in the tissues of the shore crab<Emphasis Type="Italic"> Carcinus maenas</Emphasis>: effects of size and tissue hydration

Water content in the tissues, tissue proportions, and content and concentrations of copper, zinc, iron, cadmium, manganese, and cobalt were determined in male shore crabs Carcinus maenas of different sizes. Average wet to dry weight ratios in midgut gland, gills, muscle, exoskeleton, stomach, heart, testes, hypodermis, and haemolymph were 5.3, 9.5, 4.1, 1.5, 4.8, 6.1, 3.8, 4.7, and 13, respectively. The water content in the soft tissues generally increased with increasing size of the crabs (except in testes). Dry midgut gland, gills, muscle, exoskeleton, stomach, heart, testes, hypodermis, and haemolymph constituted 4, 0.9, 20, 68, 0.5, 0.12, 0.5, 0.7 and 5.3%, respectively, of the body dry weight. Dry midgut gland, gills, stomach, hypodermis, and haemolymph constituted a larger proportion of the body weight in small than in large crabs, whereas the opposite was the case for exoskeleton. Protein concentrations in the haemolymph decreased with size whereas haemolymph volume increased. Seventy percent of the copper body burden was located in the haemolymph. The majority of zinc (61%) occurred in muscle, whereas the midgut gland was the major site of cadmium storage (89%). The exoskeleton contained the major part of the body burden of iron (71%) and manganese (95%). Most of the cobalt body burden was located in the exoskeleton (45%) and haemolymph (44%). As tissue hydration and tissue proportions changed with size, tissue metal concentrations did not necessarily vary identically with size when expressed on a dry weight compared to a wet weight basis. Tissue potassium concentrations were negatively correlated with tissue water contents, indicating that water in tissues with a high wet to dry weight ratio was of extracellular origin. However, comparison of tissue and haemolymph copper levels showed that this extracellular water could not be pure haemolymph as tissue copper concentrations would have been higher than the values actually found.     

Characterisation and expression of four mRNA sequences encoding glutathione S-transferases pi,mu, omega and sigma classes in the Pacific oyster<Emphasis Type="Italic"> Crassostrea gigas</Emphasis> exposed to hydrocarbons and pesticides

Hydrocarbon and pesticide pollution in coastal ecosystems can disturb marine bivalve metabolism. In this study, we characterised four full-length cDNA sequences encoding glutathione S-transferases (GSTs) in the Pacific oyster Crassostrea gigas. A BLAST X search showed that these four sequences encode GSTs from four different classes: GST pi, sigma, mu and omega. A phylogenetic analysis of GST was made to determine the position of oyster GST compared to invertebrate and vertebrate sequences. We developed a semi-quantitative, multiplex RT-PCR to follow the expression of these four GSTs in tissues of oysters exposed to hydrocarbons and two pesticide treatments (glyphosate and a mixture composed of atrazine, diuron and isoproturon) under experimental conditions. Our results showed strong differential expression of these four GSTs that was both tissue specific as well as time and treatment dependent. We observed that expression levels were higher in digestive gland than in gill tissues in pesticide-exposed oysters. Furthermore, omega and mu class GST mRNA expression in the digestive gland might be useful as a possible marker of hydrocarbon exposure, while pi and sigma class GST mRNA expression in the digestive gland may be similarly useful as a marker of pesticide exposure in monitoring programmes.Communicated by S.A. Poulet, Roscoff     

Modelling of paralytic shellfish toxin biotransformations in the course of Crassostrea gigas detoxification kinetics

The aim of this study was to evaluate the importance of biotransformation of paralytic shellfish toxins during the detoxification process in contaminated oysters. Mathematical models based upon the detoxification patterns of digestive gland and other tissues were developed. It was demonstrated that biotransformations do not seem to play an important role in digestive gland or other tissue detoxification kinetics with our data set. Moreover, different toxin transfers from digestive gland toward other tissues were investigated. No significant transfer was highlighted in our data set. These first conclusions were drawn after comparing the results obtained from 13 biotransformations and identifiable transfer scenarios. Finally, to determine a more robust model, all 12 states corresponding to toxic compounds and tissues were aggregated into a single state model. The best adjustment was obtained with a simple one-compartment model based on total flesh toxicity with elimination rate expressed by a function depending on initial concentrations of GTX3 and GTX2 (i.e. the two major toxic compounds found in contaminated oysters).