Metallothionein-like proteins in the blue crab Callinectes sapidus: Effect of water salinity and ions

The effect of water salinity and ions on metallothionein-like proteins (MTLP) concentration was evaluated in the blue crab Callinectes sapidus. MTLP concentration was measured in tissues (hepatopancreas and gills) of crabs acclimated to salinity 30 ppt and abruptly subjected to a hypo-osmotic shock (salinity 2 ppt). It was also measured in isolated gills (anterior and posterior) of crabs acclimated to salinity 30 ppt. Gills were perfused with and incubated in an isosmotic saline solution (ISS) or perfused with ISS and incubated in a hypo-osmotic saline solution (HSS). The effect of each single water ion on gill MTLP concentration was also analyzed in isolated and perfused gills through experiments of ion substitution in the incubation medium. In vivo, MTLP concentration was higher in hepatopancreas than in gills, being not affected by the hypo-osmotic shock. However, MTLP concentration in posterior and anterior gills significantly increased after 2 and 24 h of hypo-osmotic shock, respectively. In vitro, it was also increased when anterior and posterior gills were perfused with ISS and incubated in HSS. In isolated and perfused posterior gills, MTLP concentration was inversely correlated with the calcium concentration in the ISS used to incubate gills. Together, these findings indicate that an increased gill MTLP concentration in low salinity is an adaptive response of the blue crab C. sapidus to the hypo-osmotic stress. This response is mediated, at least in part, by the calcium concentration in the gill bath medium. The data also suggest that the trigger for this increase is purely branchial and not systemic.     

The function of hemocyanin in respiration of the blue crab Callinectes sapidus.

Blood PO2 in the blue crab Callinectes sapidus, a very active species of tropical origin, is lower at 22 degrees C than that of larger crabs in colder waters. These low oxygen levels permit its hemocyanin to be highly oxygenated at the gill, and to deliver almost half of its oxygen to the tissues in resting animals. Sustained muscular activity results in conspicuous decreases in blood PO2, pH and hemocyanin oxygenation. Although the venous reserve is fully utilized, hemocyanin oxygenation at the gill decreases so much that there is no change in its total quantitative function. The large Bohr shift becomes functional during activity, but its quantitative importance is not clear.     

The effects of hypoxia and pH on phenoloxidase activity in the Atlantic blue crab, Callinectes sapidus

In its natural coastal and estuarine environments, the blue crab, Callinectes sapidus, often encounters hypoxia, accompanied by hypercapnia (increased CO2) and an associated decrease in water pH. Previous studies have shown that exposure to hypercapnic hypoxia (HH) impairs the crab's ability to remove culturable bacteria from its hemolymph. In the present study we demonstrate that the activity of phenoloxidase (PO), an enzyme critical to antibacterial immune defense in crustaceans, is decreased at the low levels of hemolymph O2 and pH that occur in the tissues of blue crabs exposed to HH. Hemocyte PO activity was measured at tissue O2 levels that occur in normoxic (5% and 15% O2, approximate venous and arterial hemolymph, respectively) and hypoxic (1% O2) crabs and compared to PO activity in air-saturated conditions (21% O2). PO activity decreased by 33%, 49% and 70% of activity in air at 15%, 5% and 1% O2, respectively. When O2 was held at 21% and pH lowered within physiological limits, PO activity decreased with pH, showing a 16% reduction at pH 7.0 as compared with a normoxic pH of 7.8. These results suggest that decreased PO activity at low tissue O2 and pH compromises the ability of crustaceans in HH to defend themselves against microbial pathogens.     

A sex-specific metabolite identified in a marine invertebrate utilizing phosphorus-31 nuclear magnetic resonance

Hormone level differences are generally accepted as the primary cause for sexual dimorphism in animal and human development. Levels of low molecular weight metabolites also differ between men and women in circulating amino acids, lipids and carbohydrates and within brain tissue. While investigating the metabolism of blue crab tissues using Phosphorus-31 Nuclear Magnetic Resonance, we discovered that only the male blue crab (Callinectes sapidus) contained a phosphorus compound with a chemical shift well separated from the expected phosphate compounds. Spectra obtained from male gills were readily differentiated from female gill spectra. Analysis from six years of data from male and female crabs documented that the sex-specificity of this metabolite was normal for this species. Microscopic analysis of male and female gills found no differences in their gill anatomy or the presence of parasites or bacteria that might produce this phosphorus compound. Analysis of a rare gynandromorph blue crab (laterally, half male and half female) proved that this sex-specificity was an intrinsic biochemical process and was not caused by any variations in the diet or habitat of male versus female crabs. The existence of a sex-specific metabolite is a previously unrecognized, but potentially significant biochemical phenomenon. An entire enzyme system has been synthesized and activated only in one sex. Unless blue crabs are a unique species, sex-specific metabolites are likely to be present in other animals. Would the presence or absence of a sex-specific metabolite affect an animal's development, anatomy and biochemistry?     

Salinity dependent Na+-K+ATPase activity in gills of the euryhaline crab Chasmagnathus granulata

The occurrence and response of Na+-K+ATPase specific activity to environmental salinity changes were studied in gill extracts of all of the gills of the euryhaline crab Chasmagnathus granulata from Mar Chiquita coastal lagoon (Buenos Aires Province, Argentina). All of the gills exhibited a salinity dependent Na+-K+ATPase activity, although the pattern of response to environmental salinity was different among gills. As described in other euryhaline crabs highest Na+-K+ATPase specific activity was found in posterior gills (6 to 8), which, with exception of gill 6, increased upon acclimation to reduced salinity. However, a high increase of activity also occurred in anterior gills (1 to 5) in diluted media. Furthermore, both short and long term differential changes of Na+-K+ATPase activity occurred among the gills after the transfer of crabs to reduced salinity. The fact that variations of Na+-K+ATPase activity in the gills were concomitant with the transition from osmoconformity to ionoregulation suggests that this enzyme is a component of the branchial ionoregulatory mechanisms at the biochemical level in this crab.     

CO2 Excretion Across Isolated Perfused Crab Gills: Facilitation by Carbonic Anhydrase

Crab gill carbonic anhydrase is shown to facilitate the excretionof carbondioxide across isolated perfused gills. A techniquefor perfusing crab gills and assessing the metabolic viabilityof perfused gills is also described in detail. The techniqueis used to follow the disappearance of ¹⁴C label as HCO₃^–and CO₂ from internal perfusate passing through the gill. Theexcretion of the label increases with the flow rate of the externalperfusate across the outside of the gills. The addition of carbonican hydrase to the internal perfusate results in a two- to fourfoldincrease in the excretion of label while Diamox (acetazolamide)treatment decreases the excretion of label by half. It is alsosuggested that carbonic anhydrase, present in muscle tissuesof crabs, minimizes the disequilibrium of the hemolymph CO₂system as metabolically produced CO₂ leaves the tissues andenters the hemolymph. Parallels are drawn between the presenceof carbonic anhydrase in the crab gill system and the presenceof this enzyme in the respiratory organs of both aquatic andterrestrial animals.     

NaCl absorption across split gill lamellae of hyperregulating crabs: Transport mechanisms and their regulation

The method of mounting split lamellae of crab gills in modified Ussing chambers offers the advantage that active ion transport can be measured as short-circuit current and/or flux of radioactive tracers in relation to the epithelial surface. Moreover, further modern techniques like microelectrode impalements and current-noise analysis can be applied. The epithelium of posterior gills of Chinese crabs (Eriocheir sinensis) acclimated to fresh water actively absorbs Na⁺ and Cl⁻ independent of each other. The epithelium of the gills of shore crabs (Carcinus maenas) acclimated to brackish water actively absorbs NaCl in a coupled mode. The different osmotic gradients maintained by the two crab species are reflected in the characteristics of their gill epithelia. Chinese crabs, migrating to fresh water, have a tight gill epithelium. The gill epithelium of shore crabs, living in brackish water of at least 6–8‰ salinity, is an intermediate between tight and leaky. Regulation of NaCl absorption across the gill epithelium of Chinese crabs is achieved in a hormone-independent way by the haemolymph side osmolarity (autoregulation). Moreover, NaCl absorption is regulated by a hormonal factor of so far unknown chemical nature in the eyestalk extract which stimulates the transport rates via a cAMP-dependent signal transduction pathway, activating apical V-ATPase activity and increasing the number of open apical Na⁺ channels.     

Bacteria associated with crabs from cold waters with emphasis on the occurrence of potential human pathogens.

A diverse array of bacterial species, including several potential human pathogens, was isolated from edible crabs collected in cold waters. Crabs collected near Kodiak Island, Alaska, contained higher levels of bacteria than crabs collected away from regions of human habitation. The bacteria associated with the crabs collected near Kodiak included Yersinia enterocolitica, Klebsiella pneumoniae, and coagulase-negative Staphylococcus species; the pathogenicity of these isolates was demonstrated in mice. Although coliforms were not found, the bacterial species associated with the tissues of crabs collected near Kodiak indicate possible fecal contamination that may have occurred through contact with sewage. Compared with surrounding waters and sediments, the crab tissues contained much higher proportions of gram-positive cocci. As revealed by indirect plate counts and direct scanning electron microscopic observations, muscle and hemolymph tissues contained much lower levels of bacteria than shell and gill tissues. After the death of a crab, however, the numbers of bacteria associated with hemolymph and muscle tissues increased significantly. Microcosm studies showed that certain bacterial populations, e.g., Vibrio cholerae, can be bioaccumulated in crab gill tissues. The results of this study indicate the need for careful review of waste disposal practices where edible crabs may be contaminated with microorganisms that are potential human pathogens and the need for surveillance of shellfish for pathogenic microorganisms that naturally occur in marine ecosystems.     

Effect of salinity on osmoregulatory patch epithelia in gills of the blue crab Callinectes sapidus

In euryhaline crabs, ion-transporting cells are clustered into osmoregulatory patches on the lamellae of the posterior gills. To examine changes in the branchial osmoregulatory patch in the blue crab Callinectes sapidus in response to change in salinity and to correlate these changes with other osmoregulatory responses, crabs were acclimated to a range of salinities between 10 and 35 ppt. When crabs that had been acclimated to 35 ppt were subsequently transferred to 10 ppt, both the size of the osmoregulatory patch on individual gill lamellae and the specific activity of Na+, K+-ATPase in whole-gill homogenates increased only after the first 24 h of exposure to dilute seawater. Enzyme activity and size of patch area increased gradually and reached their maxima (increasing by 200% and 60%, respectively) 6 days following transfer to 10 ppt seawater and then remained at these levels. Patch size at acclimation varied inversely with the salinity for seawater dilutions below 26 ppt (the isosmotic point of the crab), although it did not vary in salinities at or above 26 ppt. Thus, the size of the patch clearly is modulated with acclimation salinity, but it increases only in those salinities in which the crab hyperosmoregulates. An increase in the total RNA/DNA ratio in gill homogenates, the lack of mitotic figures in the lamellae, and the lack of incorporation of bromodeoxyuridine into nuclei of lamellar epithelial cells during acclimation to dilute seawater were interpreted as evidence that no cell proliferation had occurred and that increases in the size of the osmoregulatory patch occurred through differentiation of existing gas exchange cells or of undifferentiated epithelial cells into ion-transporting cells.     

Salinity adaptation of HCO3--dependent ATPase activity in the gills of blue crab (Callinectes sapidus)

A bicarbonate-dependent ATPase (EC 3.6.1.3) was found in microsomal preparations from blue crab gills. When the crabs were transferred to low salinity (200 mosmolal) from seawater (1000 mosmolal), the HCO3- dependent ATPase increased in all gill pairs, reaching its new steady state in 2 weeks. The greatest increase occurred in the sixth and seventh gill pairs (approx. 2.5-fold). Maximal enzyme activity was observed at an Mg2+ concentration of 2 mM and an optimal pH of 7.8. The apparent Ka for HCO3- was found to be 8.9 mM. Kinetic analysis showed that low-salinity adaptation increased the Vmax without altering the Km for ATP. When the microsomes from high-salinity crab gills were treated with detergent or assayed at different temperatures, the total enzyme activity did not reach the activity levels after adaptation to low salinity. These results suggest that the alteration of HCO3- -ATPase activity may be due to synthesis, rather than modulation of membranes or of the existing enzyme activity.     

Effect of microcystin on ion regulation and antioxidant system in gills of the estuarine crab Chasmagnathus granulatus (Decapoda,Grapsidae)

The objective of this work was to evaluate mechanisms of microcystin toxicity on crustacean species. Adult male crabs of Chasmagnathus granulatus (13.97+/-0.35 g) acclimated to low salinity (2 per thousand ) were injected with saline (control) or Microcystis aeruginosa aqueous extract (39.2 microg/l) at 24 h intervals for 48 h. After the exposure period, the anterior and posterior gills were dissected, measuring Na(+),K(+)-ATPase and glutathione-S-transferase (GST) activity. Total oxyradical scavenging capacity (TOSC) and lipid peroxides (LPO) content were also determined. Na(+),K(+)-ATPase activity in anterior gills was significantly lower in crabs injected with toxin than in control crabs, while no significant difference in the enzyme activity was detected in posterior gills. Both sodium and chloride concentration in the hemolymph were not affected by toxin exposure. Significant changes in GST activity were detected in posterior gills, with higher values being observed in the toxin-injected crabs. Crabs exposed to microcystin also showed a significant increase in the TOSC value against peroxyl radicals, for both anterior and posterior gills. Lipid peroxides level did not change in both gill types after exposure to the toxin. The increased levels of TOSC suggest the occurrence of a crab response against oxidative stress induced by toxin injection, which prevents lipid peroxidation.     

Oxygen, gills, and embryo behavior: mechanisms of adaptive plasticity in hatching.

Many species alter the timing of hatching in response to egg or larval predators, pathogens, or physical risks. This plasticity depends on separation between the onset of hatching competence and physiological limits to embryonic development. I present a framework based on heterokairy to categorize developmental mechanisms and identify traits contributing to and limiting hatching plasticity, then apply it to a case of predator-induced hatching. Red-eyed treefrogs have arboreal eggs, and tadpoles fall into ponds upon hatching. Egg and tadpole predators select for earlier and later hatching, respectively. Embryos hatch up to 30% early in predator attacks, and later if undisturbed. They maintain large external gills throughout the plastic hatching period, delaying gill regression while development otherwise continues. Rapid gill regression occurs upon hatching. Prolonged embryonic development depends on external gills; inducing gill regression causes hatching. External hypoxia retards development, kills eggs, and induces hatching. Nonetheless, embryos develop synchronously and without hatching prematurely across a broad range of perivitelline PO2, from 0.5-12.5 kPa. Embryos exploit spatial variation of PO2 within eggs by positioning gills against patches of air-exposed surface. Respiratory plasticity and oxygen-sensitive behavior appear critical for the hatching plasticity that balances a predation risk trade-off across life stages.     

Bacterial diseases of crabs: a review

Bacterial diseases of crabs are manifested as bacteremias caused by organisms such as Vibrio, Aeromonas, and a Rhodobacteriales-like organism or tissue and organ tropic organisms such as chitinoclastic bacteria, Rickettsia intracellular organisms, Chlamydia-like organism, and Spiroplasma. This paper provides general information about bacterial diseases of both marine and freshwater crabs. Some bacteria pathogens such as Vibrio cholerae and Vibrio vulnificus occur commonly in blue crab haemolymph and should be paid much attention to because they may represent potential health hazards to human beings because they can cause serious diseases when the crab is consumed as raw sea food. With the development of aquaculture, new diseases associated with novel pathogens such as spiroplasmas and Rhodobacteriales-like organisms have appeared in commercially exploited crab species in recent years. Many potential approaches to control bacterial diseases of crab will be helpful and practicable in aquaculture.     

Effects of hypercapnic hypoxia on the clearance of Vibrio campbellii in the Atlantic blue crab, Callinectes sapidus Rathbun

Callinectes sapidus, the Atlantic blue crab, encounters hypoxia, hypercapnia (elevated CO(2)), and bacterial pathogens in its natural environment. We tested the hypothesis that acute exposure to hypercapnic hypoxia (HH) alters the crab's ability to clear a pathogenic bacterium, Vibrio campbellii 90-69B3, from the hemolymph. Adult male crabs were held in normoxia (well-aerated seawater) or HH (seawater with PO(2) = 4 kPa; PCO(2) = 1.8 kPa; and pH = 6.7-7.1) and were injected with 2.5 x 10(4) Vibrio g(-1) body weight. The animals were held in normoxia or in HH for 45, 75, or 210-240 min before being injected with Vibrio, and were maintained in their respective treatment conditions for the 120-min duration of the experiment. Vibrio colony-forming units (CFU) ml(-1) hemolymph were quantified before injection, and at 10, 20, and 40 min afterward. Total hemocytes (THC) ml(-1) of hemolymph were counted 24 h before (-24 h), and at 10 and 120 min after injection. Sham injections of saline produced no change in the bacterial or hemocyte counts in any treatment group. Among the groups that received bacterial injections, Vibrio was almost completely cleared within 1 h, but at 10-min postinjection, Vibrio CFU ml(-1) hemolymph was significantly higher in animals held in HH for 75 and 210-240 min than in those held in normoxia. Within 10 min after crabs were injected with bacteria, THC ml(-1) significantly decreased in control and HH45 treatments, but not in the HH75 and HH210-240 treatments. By 120 min after injection of bacteria, hemocyte counts decreased in all but the HH45 group. These data demonstrate that HH significantly impairs the ability of blue crabs to clear Vibrio from the hemolymph. These results also suggest that HH alters the normal role of circulating hemocytes in the removal of an invading pathogen.     

Early carbonic anhydrase induction in the gills of the blue crab, Callinectes sapidus, during low salinity acclimation is independent of ornithine decarboxylase activity

Carbonic anhydrase (CA) induction in the gills of the euryhaline blue crab, Callinectes sapidus, was measured in response to lowered environmental salinity. Simultaneous measurements of ornithine decarboxylase (ODC) activity were made in gills and nonbranchial tissues to determine whether ODC activity and the resultant synthesis of polyamines played a role in the initiation and regulation of CA induction. CA induction in the seventh gill pair (G7) was proportional to the decrease in ambient salinity, but activity in the third gill pair (G3) remained unchanged. Induction began by 24 hr after low salinity transfer, much earlier than previously reported, and peaked after 4 days. The magnitude of salinity change affected the magnitude of CA induction only, not the time course. A general cell volume regulatory response, as measured by the appearance of total ninhydrin-positive substances (TNPS) in the hemolymph, was initiated within 4 hr of low salinity transfer and was complete by 24 hr post-transfer. General cell swelling may be the initial signal in the pathway of CA induction. ODC activity in the gills of acclimated animals was not influenced by salinity. For crabs transferred from 35 to 25 ppt, ODC activity did not change significantly over the time course of acclimation. There was an early but transient increase in ODC activity in all tissues for crabs acclimated to 28 ppt and transferred to 15 ppt. Induction of ODC activity does not appear to be a precursor for CA induction; therefore, it does not appear that polyamines are substantially involved in the up-regulation of transport enzyme activity in low salinity. ODC, and resultant polyamine synthesis, may, however, have a role in cell volume regulation.     

Effects of eyestalk ablation on carbonic anhydrase activity in the euryhaline blue crab Callinectes sapidus: neuroendocrine control of enzyme expression

Carbonic anhydrase (CA) activity in the gills of the euryhaline blue crab, Callinectes sapidus, was measured in response to acute low-salinity transfer and treatment with eyestalk ablation (ESA) in an attempt to elucidate potential regulatory mechanisms of salinity-mediated CA induction. ESA alone resulted in an approximate doubling of CA activity in the posterior, ion-transporting gills of crabs acclimated to 35 ppt. Transfer of intact crabs to 28 ppt, a salinity at which the blue crab is still an osmotic and ionic conformer, had no effect on CA activity, but treatment with ESA prior to transfer resulted in a 5-fold increase. Hemolymph osmolality was unaffected by ESA. There was a 7-fold induction of CA activity in posterior gills of intact crabs transferred from 35 to 15 ppt, and this was potentiated by about 100% by ESA. Hemolymph osmolality was slightly elevated in the ESA-treated crabs. CA activity in anterior gills did not increase in response to any treatment. Hemolymph concentrations of methyl farnesoate (MF) were measured for all experimental animals. MF concentrations were undetectable in all intact crabs, regardless of salinity. Treatment with ESA resulted in elevated levels of hemolymph MF, but these levels were still relatively low and unrelated to salinity. These results suggest that CA induction is under the control of a regulatory substance located in the eyestalk. This substance appears to be a CA repressor, keeping CA expression at low levels in the gills of crabs acclimated to high salinity. Exposure to low salinity, or treatment with ESA, removes the effects of this putative repressor and allows CA induction to occur.     

Ultrastructural changes in the gill epithelium of the green crab Carcinus maenas in relation to the external salinity

Observation of semi-thin and ultrathin sections performed in the gills of green crabs (Carcinus maenas) kept in 100% and in dilute 30% sea water respectively reveals marked differences between the six anterior and the three posterior pairs of gills. The anterior gill lamellae are almost entirely lined by a thin pavement epithelium (0.9 to 3 mum thick) which does not undergo any noticeable change when crabs are acclimated from full to dilute sea water. This supports the view it is chiefly involved in the respiratory function. In addition to the pavement epithelium, the posterior gills exhibit small areas corresponding to a thick prismatic epithelium (10 mum) the ultrastructure of which is similar to that of most of the so-called 'salt transporting epithelia'. When submitted to reduced external salinity, this epithelium undergoes structural changes consisting of elaboration of an extensive apical plasma membrane infolding system, enlargement of the subcuticular compartment and close association of mitochondria with basolateral membrane infoldings. Pilaster cells exhibit ultrastructural features of either thin (respiratory) or thick (salt transporting) epithelial differentiation according to their localization within the gill. Their peculiar organization suggests they ensure, in addition, mechanical reinforcement of the gill lamellae against blood hydrostatic pressure. The fact that salt-transporting epithelium areas do not exceed, at most, 30% of the total lamellar surface is probably related to the weak osmoregulatory capabilities of the shore crab.     

Pathological and physiological changes in the South African freshwater crab Potamonautes warreni calman induced by microbial gill infestations.

The impact of microbial gill infestations on the pathology and physiology of the freshwater crab Potamonautes warreni was investigated by comparison of infested and uninfested crab populations from, respectively, a polluted and an unpolluted site along the Mooi River, North West Province, South Africa. Heavy gill infestations by bacteria (70%), peritrichous ciliates such as Lagenophrys sp. (15%), Zoothamnium sp. (10%), and Epistylis sp. (5%), and motile protozoans resulted in species-specific lesions in the gill epithelia of P. warreni and physiological changes in crabs from the polluted site. Bacterial colonies enmeshed in polysaccharide-like films produced indentations of the gill cuticular surfaces and dissociation of microvillous membranes at the basal zone of epithelial cells of gill lamellae of P. warreni. Lagenophrys sp. induced large subcuticular spaces with an unfolding or resorption of the plasma membrane in the gill epithelia. The attachment of stalks of Zoothamnium and Epistylis resulted in dilation of lamellar tissues, the formation of vacuoles, and an increase in subcuticular spaces in the epithelia. Physiological changes in infested crabs included significant differences (P = 0.001) in increments of wet body mass and a reduced growth rate over time compared with uninfested crabs. The specific oxygen consumption (M(O2)) in rested infested crabs significantly increased (31.29 +/- 5.8 micromol O2/kg/min) compared with the M(O2) in uninfested crabs (27.92 +/- 5.6 micromol O2/kg/min; P = 0.009). The heart rate of infested rested P. warreni was significantly lower (40.77 +/- 13.79 beats/min; P < 0.02) than that in uninfested crabs (61.09 +/- 29.02 beats/min) but the heart rate of infested crabs increased significantly with body mass (r = 0.53, P = 0.02). These findings suggest an interrelationship among organic pollution, microbial gill infestations, and specific pathological and physiological responses in the crab host. The role of P. warreni and its microbial gill communities as bioindicators of pollution are discussed.     

Response to environmental salinity of Na-KATPase activity in individual gills of the euryhaline crab Cyrtograpsus angulatus

The occurrence, localization and response to environmental salinity changes of Na⁺-K⁺ATPase activity were studied in each of the individual gills 4-8 of the euryhaline crab Cyrtograpsus angulatus from Mar Chiquita coastal lagoon (Buenos Aires Province, Argentina). Na⁺-K⁺ATPase activity appeared to be differentially sensitive to environmental salinity among gills. Upon an abrupt change to low salinity, a differential response of Na⁺-K⁺ATPase activity occurred in each individual gill which could suggest a differential role of this enzyme in ion transport process in the different gills of C. angulatus. With the exception of gill 8, a short-term increase of Na⁺-K⁺ATPase specific activity was observed in posterior gills, which is similar to adaptative variations of this activity described in other euryhaline crabs. However, and conversely to that described in other hyperregulating crabs, the highest increase of activity occurred in anterior gills 4 by 1 day after the change to dilute media which could suggest also a role for these gills in ion transport processes in C. angulatus. The fact that variations of Na⁺-K⁺ATPase activity in anterior and posterior gills were concomitant with the transition to hyperregulation indicate that this enzyme could be a component of the branchial ionoregulatory mechanisms at the biochemical level in this crab. The results suggest a differential participation of branchial Na⁺-K⁺ATPase activity in ionoregulatory mechanisms of C. angulatus. The possible existence of functional differences as well as distinct regulation mechanisms operating in individual gills is discussed.     

Uptake of cadmium through isolated perfused gills of the Chinese mitten crab, Eriocheir sinensis

Using the perfusion method, we compared cadmium accumulation and influx across the gills of the euryhaline Chinese mitten crab Eriocheir sinensis, exposed to 4.8 microM cadmium in the incubation medium (OUT). Cadmium influx was not observed across posterior gills while it ranged from 0.15 to 6.82 nmol Cd g(-1) gill w.w. h(-1) across anterior ones. For these respiratory gills, a strong increase (40 times) was observed when calcium was removed in both incubation and perfusion media while the lack of sodium in the perfusion medium resulted in a 46 times decrease. For crabs acclimated 15 days to artificial seawater, cadmium influx across anterior gills showed a 21 times decrease when compared with freshwater acclimated ones. On the other hand, after 3 h of perfusion, we detected cadmium accumulation in both types of gills, ranging from 3.8 to 68 nmol Cd g(-1) gill w.w. in anterior gills and from 2.1 to 39 nmol Cd g(-1) gill w.w. in posterior ones. Such accumulations represent between 61.3 and 100% of the total uptake of cadmium through the gills. From these results, we suggest that cadmium can penetrate more easily into the hemolymph space through the 'respiratory' type epithelium present in the anterior gills but absent in the posterior ones. This metal uptake is likely to occur at least in part through the same pathways as calcium. On the contrary, cadmium seems to be sequestered inside the posterior gills, perhaps in the cuticle of the salt-transporting type epithelium.