Na+-K+-adenosine triphosphatase activities in gills of marine teleost fishes: Changes with depth,size and locomotory activity level

Activities of the primary enzyme responsible for monovalent ion regulation, Na⁺-K⁺-adenosine triphosphatase (Na⁺-K⁺-ATPase), were measured in gills of marine teleost fishes with different depths of occurrence (0 to 4800 m), body weights (a range of five orders of magnitude), and locomotory capacities. Specimens were collected off the coasts of California and Oregon in 1983–1989, and at the Galápagos Spreading Center and 13°N East Pacific Rise hydrothermal vent sites in 1987 and 1988, respectively. Except for two hydrothermal vent fishes, deep-sea species had much lower Na⁺-K⁺-ATPase activities g^–1 gill filament than shallow-living species, indicating that osmoregulatory costs, like total metabolic rate, are greatly reduced in most deep-living fishes. Within a species, the total branchial Na⁺-K⁺-ATPase activity per individual was dependent on size; the average allometric scaling exponent was 0.83. Using published values for oxygen consumption rates, and the total branchial Na⁺-K⁺-ATPase activities as an index of osmoregulatory costs, we estimated the maximal cost (as percent of ATP turnover) for osmoregulation in ten teleosts. Osmoregulatory costs averaged about 10% of total ATP turnover among these species, and maximal costs were no greater than about 20%. The percent costs of osmoregulation did not differ between shallow- and deep-living fishes. The reduced total ATP expenditure for osmoregulation in deep-living fishes is proposed to result from the sluggish locomotory habits of these fishes, not from selection for reduced osmotic coastper se. Thus, the reduced swimming abilities of these fishes lead to lower rates of water flow over the gills and less blood flow through the gills due to reduced demands for oxygen. Consequently, passive flux of water and ions through the gills is much lower than in more active fishes, and osmotic costs are thereby minimized. The relatively high activities of Na⁺-K⁺-ATPase in gills of the two hydrothermal vent fishes suggest that these fishes may be more active and have higher metabolic rates than other deep-sea fishes.     

Simultaneous behavior of skipjack (Katsuwonus pelamis), bigeye (Thunnus obsesus), and yellowfin (T. albacares) tunas, within large multi-species aggregations associated with drifting fish aggregating devices (FADs) in the equatorial eastern Pacific Ocean

Ten separate experiments monitoring the simultaneous behaviors of 26 skipjack (Katsuwonus pelamis), 26 bigeye (Thunnus obesus), and 33 yellowfin (T. albacares) tunas within large multi-species aggregations associated with drifting fish aggregating devices (FADs) were investigated using ultrasonic telemetry in the equatorial eastern Pacific Ocean. Experiments were conducted during a research cruise aboard a chartered purse seine vessel. Purse seine sets were made on the tuna aggregations associated with FADs at the termination of six of the ten experiments. Seventeen of the 44 tagged tunas were not recaptured indicating the transient nature of the associative behavior of tunas with FADs. Although there was considerable overlap in the depths of the three species, by day and night, there were some species-specific differences and diel differences within species. While we documented spatial and temporal differences in the schooling behavior of the three tuna species, the differences do not appear sufficient such that modifications in purse seine fishing practices could effectively avoid the capture of small bigeye and yellowfin tunas, while optimizing the capture of skipjack tuna in purse seine sets on FADs.     

Branchial and intestinal osmoregulatory acclimation in the four-eyed sleeper, <Emphasis Type="Italic">Bostrychus sinensis</Emphasis> (Lacepède), exposed to seawater

Bostrychus sinensis is a facultative air breather that inhabits waters of a wide range of salinities. This study aimed to elucidate whether branchial and intestinal osmoregulatory acclimation occurred in B. sinensis transferred from 5‰ water through a progressive increase in salinities to seawater. Our results indicate that B. sinensis acted as a hyperosmotic regulator in 5‰ water, but exhibited hypoosmotic hypoionic regulation in seawater. During short- (1 day) and medium- (10 days) term acclimation to seawater, there were only minor perturbations in plasma osmolality and [Na⁺], which returned to control levels after 45 days of exposure to seawater. Branchial Na⁺/K⁺-ATPase activity was unaffected by 1, 10 or 45 days of exposure to seawater. However, prolonged (45 days) acclimation to seawater led to a significant increase in Na⁺/K⁺-ATPase α-subunit protein abundance. Taken together, these results indicate that there could be changes in the expression of Na⁺/K⁺-ATPase isoforms and/or post-translational modification of Na⁺/K⁺-ATPase in the gills of fish exposed to seawater. Immunofluorescence microscopy revealed that acclimation to seawater for 10 days only resulted in no change in branchial Na⁺/K⁺-ATPase protein expression, but there were increases in protein expression of cystic fibrosis transmembrane regulator (CFTR)-like chloride channel and Na⁺:K⁺:2Cl⁻ cotransporter (NKCC; probably NKCC1). Indeed, NKCC was undetectable in gills of fish kept in 5‰ water by Western blotting, but it became weakly detectable in fish exposed to seawater for 10 days and prominently expressed in fish exposed to seawater for 45 days. Therefore, our results indicate that branchial CFTR-like chloride channel and NKCC1 were the determining factors in the transition between hyperosmotic regulation and hypoosmotic hypoionic regulation in B. sinensis. Furthermore, the intestine of B. sinensis also served as an important osmoregulatory organ, since there were significant increases in both the activity and protein abundance of intestinal Na⁺/K⁺-ATPase in fish acclimated to seawater for 45 days. The effectiveness of branchial and intestinal osmoregulatory acclimation in B. sinensis during seawater acclimation led to only a minor increase in plasma osmolality, and thus resulted in relatively unchanged free amino acid contents in muscle and liver.     

Acclimation of Solea senegalensis to different ambient temperatures: implications for thyroidal status and osmoregulation

We have investigated the regulation of thyroidal status and osmoregulatory capacities in juveniles from the teleost Solea senegalensis acclimated to different ambient temperatures. Juveniles, raised in seawater at 19°C, were acclimated for 3 weeks to temperatures of 12, 19 and 26°C. Since our preliminary observations showed that at 12°C feed intake was suppressed, our experimental design controlled for this factor. The concentration of branchial Na⁺,K⁺-ATPase, estimated by measurements of enzyme activity at the optimum temperature of this enzyme (37°C), did not change. In contrast, an increase in Na⁺,K⁺-ATPase activity (measured at 37°C), was observed in the kidney of 12°C-acclimated fish. In fish acclimated to 12°C, the hepatosomatic index had increased, which correlated with increased plasma levels of triglycerides and non-esterified fatty acids. Plasma cortisol levels did not differ significantly between the experimental groups. In liver and gills, the amount of iodothyronine deiodinases that exhibit thyroid hormone outer ring deiodination was up-regulated only when fish did not feed. When assayed at the acclimation temperature, kidney deiodinase activities were similar, indicating a temperature-compensation strategy. 3,5,3′-triiodothyronine (T3) tissue concentrations in gills and kidney did not differ significantly between experimental groups. However, at 12°C, lower T3 tissue levels were measured in plasma and liver. We conclude that S. senegalensis adjusts its osmoregulatory system to compensate for the effects of temperature on electrolyte transport capacity. The organ-specific changes in thyroid hormone metabolism at different temperatures indicate the involvement of thyroid hormones in temperature acclimation.     

Environmental preferences of yellowfin tuna (Thunnus albacares) at the northern extent of its range

We used acoustic telemetry to examine the small-scale movement patterns of yellowfin tuna (Thunnus albacares) in the California Bight at the northern extent of their range. Oceanographic profiles of temperature, oxygen, currents and fluorometry were used to determine the relationship between movements and environmental features. Three yellowfin tuna (8 to 16 kg) were tracked for 2 to 3 d. All three fish spent the majority of their time above the thermocline (18 to 45 m in depth) in water temperatures >17.5 °C. In the California Bight, yellowfin tuna have a limited vertical distribution due to the restriction imposed by temperature. The three fish made periodic short dives below the thermocline (60 to 80 m), encountering cooler temperatures (>11 °C). When swimming in northern latitudes, the depth of the mixed layer largely defines the spatial distribution of yellowfin tuna within the water column. Yellowfin prefer to spend most of their time just above the top of the thermocline. Oxygen profiles indicated that the tunas encountered oceanic water masses that ranged most often from 6.8 to 8.6 mg O₂ l⁻¹, indicating no limitation due to oxygen concentrations. The yellowfin tuna traveled at speeds ranging from 0.46 to 0.90 m s⁻¹ (0.9 to 1.8 knots h⁻¹) and frequently exhibited an oscillatory diving pattern previously suggested to be a possible strategy for conserving energy during swimming. Received: 14 February 1997 / Accepted: 14 April 1997     

Stable isotopic evidence for trophic groupings and bio-regionalization of predators and their prey in oceanic waters off eastern Australia

Muscle tissue was collected for stable isotope analysis (SIA) from the main fish predators and their fish and cephalopod prey from oceanic waters off eastern Australia between 2004 and 2006. SIA of δ¹⁵N and δ¹³C revealed that the species examined could be divided into three main trophic groups. A “top predator” group consisted mainly of large billfish (Xiphias gladius and Tetrapturus audax), yellowfin (Thunnus albacares), bigeye (T. obesus) and southern bluefin (T. maccoyii) tunas and sharks; with mako (Isurus oxyrinchus) the highest. Below this tier was a second group composed of mid-trophic level fishes including albacore tuna (Thunnus alalunga), lancet fish (Alepisaurus ferox), mahi mahi (Coryphaena hippuris) and ommastrephid squid. Underlying both groups was a grouping of small fishes including myctophids, small scombrids and nomeids as well as surface fishes including macrorhamphosids. These groupings were based largely on mean animal size which showed a positive linear relation to δ¹⁵N (r ² = 0.58). Some species showed significant ontogenetic variation in either δ¹⁵N (swordfish, lancet fish, yellowfin and albacore tuna) or δ¹³C (mako shark). We also noted a consistent latitudinal change in δ¹⁵N and δ¹³C at ~28°S for the top predator species, particularly albacore and yellowfin tuna. The differences were consistent with a change from oligotrophic Coral Sea to nutrient rich Tasman Sea waters. These differences suggest that predatory fishes may have extended residence time in distinct regions off eastern Australia.     

Biomarkers as tools to assess the chronic toxicity of ammonia in the juvenile Mugil cephalus

With juvenile fish as the subject, the effects of low concentration ammonia on antioxidant system were studied using Mugil cephalus. Samples of gill and liver tissue were obtained from 0.35, 0.70, 1.5 and 3?mg/L ammonia groups at 0, 5, 10, 15 and 20 days of exposure, at which times the biomarkers were measured. Results showed that gill malondialdehyde (MDA) content exhibited an initial significant increase (p?≤?0.05) at unionised ammonia concentrations of 0.70, 1.5 and 3.0?mg/L on day 5, followed by subsequent declines, while liver MDA levels exhibited significant increases (p?≤?0.05) at unionised ammonia concentrations of 1.5?mg/L starting on day 10 and at 3.0?mg/L starting on day 5. With exposure to ammonia at different concentrations, Na⁺-K⁺-ATPase activity in liver and gill decreased over time. The Na⁺-K⁺-ATPase activity was negatively related to ammonia concentration from 0.70 to 3.0?mg/L. Overall, our results show that MDA and Na⁺-K⁺-ATPase, evaluated here as potential biomarkers of ammonia exposure, exhibited responses to sublethal concentrations of ammonia that were concentration dependent.     

食物源nC_(60)损害斑马鱼脑、鳃、肾和肝胰腺正常机能

水体中稳定存在的富勒烯纳米晶体(nC_(60))可被浮游动物滤食,并通过食物链传递到更高营养级生物。为探究食物源nC_(60)的生物效应,本试验选取携带nC_(60)的大型溞喂养斑马鱼21 d,考察了食物源nC_(60)对斑马鱼脑、鳃、肾和肝胰腺4个器官中ROS、Na~+K~+-ATPase、Ca2+-ATPase、酸性磷酸酶(ACP)、碱性磷酸酶(AKP)、谷丙转氨酶(GPT)和谷草转氨酶(GOT)活性等指标,用以评价食物源nC_(60)对斑马鱼的机能影响。暴露于食物源nC_(60)下的结果表明:斑马鱼脑ROS随时间增加而增加,暴露21 d后增加了79.17%。鳃、肾Na~+K~+-ATPase活性随暴露时间增加而降低,暴露21 d后分别降低了47.09%和51.07%;鳃、肾Ca2+-ATPase活性随暴露时间增加而减少,暴露21 d后分别降低了28.28%和35.13%。鳃、肾、肝胰腺AKP活性随时间增加而增加,暴露21 d后分别增加45.97%、26.68%和83.01%;鳃、肾、肝胰腺ACP活性随时间增加而增加,暴露21 d后分别增加38.85%、84.12%和55.77%。肝胰腺GPT和GOT活性随时间增加而降低,暴露21 d后各降低了50.05%和76.50%。本研究不但阐述了食物源nC_(60)降低高一级水生动物(斑马鱼)脑、鳃、肾和肝胰腺的正常机能,而且为进一步研究食物源nC_(60)对水生生物的生态毒理提供了部分基础数据。     

Glucose and glycerol uptake by isolated zooxanthellae fromCassiopea xamachana: Transport mechanisms and regulation by host homogenate fractions

We investigated the mechanisms by which glycerol and glucose enter freshly isolated zooxanthellae (FIZ),Symbiodinium microadriaticum Freudenthal, of the mangrove jellyfish,Cassiopea xamachana Bigelow, and the specific sites of host factor interaction. Glucose entry into FIZ is accomplished by a Na⁺-dependent symport system driven by an electrochemical gradient generated via a Na⁺-K⁺ ATPase. Inhibition of glucose uptake by a low molecular weight fraction of host homogenates [mol. wt<2 kilodaltons (kD)] occurs through the interaction of a putative host factor with the carrier protein and not the ATPase. Glycerol entry is apparently accomplished by simple or facilitated diffusion and is not affected by host homogenate fractions.     

ATP content and adenylate energy charge of the mussel Mytilus edulis during the annual reproductive cycle in Lindåspollene,Western Norway

The oxygen consumption rates ( VO₂) of 6 specimens (6 to 13 kg) of the albacore tuna Thunnus alalunga were measured at sea, using specimens collected 300 km west of San Diego, California (USA) during July and August, 1981. Fish were tested in a closed continuous-flow respirometer, where they swam at about 1.3 body lengths s^-1 velocity in 15° to 19°C water. The albacore tuna is a temperate pelagic species experiencing water temperatures from about 10° to 20°C and attaining a maximum weight of 45 kg. The VO₂ ranged from 1 249 to 3 336 ml h^-1 (the mean VO₂ for the 6 fish was 2 228 ml h^-1); such values approach those of mammals of a similar size and are 3 to 4 times those of most active fishes (e.g. sockeye salmon). Among fishes, the only higher VO₂ values yet recorded were for the skipjack tuna Katsuwonus pelamis, a tropical species. The remarkably high metabolic rates of tunas are presumably correlated with their continuous swimming activity and the maintenance of endothermy. The exponent relating VO₂ to body weight (1.18), although large, is not statistically different from the exponents for most other active vertebrates.     

三甲基氯化锡对斑马鱼(Danio rerio)生理生化特性的影响

为初步探讨三甲基氯化锡(trimethyltin chloride,TMT)对鱼类的毒性效应以及评价环境中TMT的安全性,采用静态鱼类急性毒性试验法测定了TMT对斑马鱼(Danio rerio)生理生化指标的影响;参考TMT的96 h-LC50值,设定3个浓度(0.39、0.78和1.17 mg·L-1)处理斑马鱼,测...     

Evidence that the rotifer<Emphasis Type="Italic"> Brachionus plicatilis</Emphasis> is not an osmoconformer

The rotifer Brachionus plicatilis is euryhaline (growing between 2 and 97 ppt) and has previously been considered an osmoconformer. We suggest that B. plicatilis is an osmoregulator, exhibiting a pattern of Na⁺/K⁺ ATPase activity in response to salinity consistent with that of other osmoregulating euryhaline invertebrates. To examine salinity tolerance, growth rates between 5 and 60 ppt were determined. The activity of Na⁺/K⁺ ATPase was examined, over the same range of salinities, by measuring ATPase activity in rotifer homogenates in the presence and absence of a Na⁺/K⁺ ATPase inhibitor. Maximum specific growth rate (0.95 day^–1) occurred at 16 ppt, highest mean amictic eggs per female (1.41) occurred at 20 ppt, and both parameters decreased rapidly as salinity increased. Egg development time was constant with salinity at 0.92 days. The activity of Na⁺/K⁺ ATPase per milligram protein increased from 3.9 µmol h^–1 at 5 ppt to 6.8 µmol h^–1 at 50 ppt and accounted for 15 and 30% of total ATPase activity, respectively. We suggest that these observations are consistent with increasing stress at high salinities and the occurrence of a hypo-osmoregulatory response. Given the high ATP consumption of Na⁺/K⁺ ATPase at high salinities, it is possible that a proportion of the corresponding decreases in growth rate and egg production are a direct cost of regulation.Communicated by J.P. Thorpe, Port Erin     

A comparison of transport-related gill enzyme activities and tissue-specific free amino acid concentrations of Baltic Sea (brackish water) and freshwater threespine sticklebacks, Gasterosteus aculeatus, after salinity and temperature acclimation

The osmoregulatory abilities of one freshwater and two brackish water (Baltic Sea) populations of the euryhaline teleost fish Gasterosteus aculeatus were studied with respect to evolutionary physiology. Plasma osmolality, activities of Na⁺K⁺-ATPase, citrate synthase, creatine kinase in the gill and free amino acids in liver, axial muscle and pectoral fin muscle were measured. After transfer from 10 to 35 ppt at 15 °C, time-course changes of plasma osmolality and gill Na⁺K⁺-ATPase showed no significant fundamental differences between the freshwater and one of the Baltic Sea populations. In a multi-factorial experiment, each population was exposed to four different abiotic regimes. Both brackish water populations had high mortality in freshwater at 4 °C, which is discussed as a failure of osmotic regulation (reduced taurine concentrations). Freshwater specimens had higher levels of glycine in the axial and pectoral fin muscles compared to the brackish water populations. This is interpreted as a genetically based effect. In brackish (20 ppt) water of 15 °C, the freshwater population had high activities of Na⁺K⁺-ATPase, but low activities of creatine kinase, whereas both brackish water populations behaved in the opposite way. A fundamental difference between the freshwater and brackish water populations on the level of the osmoregulatory machinery was not observed. Received: 10 December 1998 / Accepted: 22 September 1999     

Blood oxygen-binding characteristics of bigeye tuna (Thunnus obesus), a high-energy-demand teleost that is tolerant of low ambient oxygen

 We found blood from bigeye tuna (Thunnus obesus) to have a significantly higher O₂ affinity than blood from other tunas. Its P₅₀ (partial pressure of oxygen, P_O2 required to reach 50% saturation) was 1.6 to 2.0 kPa (12 to 15 mmHg) when equilibrated with 0.5% CO₂. Previous studies employing similar methodologies found blood from yellowfin tuna (T. albacares), skipjack tuna (Katsuwonus pelamis), and kawakawa (Euthynnus affinis) to have a P₅₀ of 2.8 to 3.1 kPa (21 to 23 mmHg). These observations suggest that bigeye tuna are more tolerant of low ambient oxygen than other tuna species, and support similar conclusions derived from laboratory whole-animal studies, depth-of-capture data, and directly-recorded vertical movements of fish in the open ocean. We also found the O₂ affinity of bigeye tuna blood to be essentially unaffected by a 10 C° open-system temperature change (as is the blood of all tuna species studied to date). The O₂ affinity of bigeye tuna blood was, however, more affected by a 10 C° closed-system temperature change than the blood of any tuna species yet examined. In other words, bigeye tuna blood displayed a significantly enhanced Bohr effect (change in log P₅₀ per unit change in plasma pH at P₅₀) when subjected to the inevitable changes in partial pressure of carbon dioxide (P_CO2) and plasma pH that accompany closed-system temperature shifts, than when subjected to changes in plasma pH accomplished by changing P_CO2 alone. In vivo, the resultant large decrease in O₂ affinity (i.e. the increase in P₅₀) that occurs as the blood of bigeye tuna is warmed during its passage through the vascular counter-current heat exchangers ensures adequate rates of O₂ off-loading in the swimming muscles of this high-energy-demand teleost. Received: 12 March 1999 / Accepted: 18 December 1999     

Analysis of the eastern Pacific yellowfin tuna fishery based on multiple management objectives

Vector optimization techniques were used to generate arbitrary segments of a policy frontier for a dynamic yellowfin tuna (Thunnus albacares) fishery model assuming fixed technology and considering four policy objectives: minimizing dolphin mortality, minimizing incidental catch (all species except dolphins), maximizing sustainable yield, and minimizing biological risk for the yellowfin tuna stock. Results show that along the policy frontier: (1) reducing incidental dolphin mortality increases the incidental catch of other species in a nonlinear way; (2) yield increases (subject to a biomass precautionary level) can only be obtained at the expense of higher levels of dolphin mortality and incidental catch; (3) biological risk increases as the level of tunas caught increases, but this increase depends on the type of fishery (longline fishing and three different modes of purse-seining: log-sets, dolphin-sets or school-sets) that dominates the fishing effort; (4) there is an indirect relationship between the dolphin mortality levels and those of biological risk; (5) there is a direct relationship between the incidental catch levels and biological risk. Catch obtained with dolphin-sets dominates the Pareto-optimal solutions with highest dolphin mortality levels but is associated with lower biological risk, whereas catch obtained with log-sets dominates in Pareto-optimal solutions with higher incidental catch and higher biological risk. In general, trade-offs or shadow prices among objectives are not linear, indicating that marginal costs vary along the policy frontier. Results of the trade-off analysis may provide useful information for decision-makers and other policy actors. Complete information about the preferences of the decision-makers regarding the objectives is necessary to recommend a specific management policy.     

Effect of MS-222 on response to light and rate of metabolism of the little skate Raja erinacea

The effect of tricaine methanesulfonate (MS-222) on the standard (SMR) and routine (RMR) metabolic rates of Raja erinacea was estimated from oxygen-consumption measurements. Data were gathered from a computerized, flow-through respirometry system. Individual trials were run for 96 h at 10 °C on anesthetized and untreated fish. The resulting rates, SMR = 20.1 ± 1.99 SE and RMR = 48.3± 2.5 SE mg O₂ kg⁻¹ h⁻¹, for an unanesthetized skate of 0.5 kg standardized weight, are the lowest reported for any elasmobranch. Periodogram analysis revealed a significant light-response component to the oxygen consumption of these fish, indicating a pattern of nocturnal and crepuscular activity. This activity pattern was disrupted in skates subjected to anesthesia. The use of low-dosage MS-222 in conjunction with the respirometry trials provides positive preliminary evidence that this technique may be useful in giving quick, accurate estimates of SMR in the more intractable elasmobranch fishes. Received: 15 January 1997 / Accepted: 6 February 1997     

Modulation of branchial ion transport protein expression by salinity in glass eels (<Emphasis Type="Italic">Anguilla anguilla</Emphasis> L.)

The Anguillid juvenile glass eel must deal with the osmoregulatory consequences of highly variable environmental salinities on its recruitment migration from coastal to fresh waters. Changes in ionoregulatory parameters and branchial ion transport protein [Na⁺/K⁺-ATPase, Na⁺:K⁺:2Cl⁻ cotransporter (NKCC), cystic fibrosis transmembrane regulator (CFTR) anion channel, V-type proton ATPase] expression (activities, protein and/or mRNA level expression and/or cellular localization) in response to acclimation to a broad range of ionic strengths [distilled water (DW) to hypersaline water (HSW; 150%) sea water (SW 32‰)] was studied. The estuarine glass eels were very euryhaline and successfully acclimated to acute changes in environmental ionic strength from 50% SW, with high mortality only observed in HSW (51%) and sublethal osmoregulatory indicators (whole body water content and sodium levels) disturbed at the extremes (DW and HSW). Central to a high salinity acclimation were elevated branchial Na⁺/K⁺-ATPase, NKCC and CFTR expression. At lower salinity, Na⁺/K⁺-ATPase expression was maintained and NKCC and CFTR expressions were reduced. Branchial chloride cells increased in size up to SW but decreased in HSW. During hypotonic disturbance (DW), no compensatory elevation in V-ATPase or Na⁺/K⁺-ATPase expression was observed.     

Protein kinase C and ion transport in the posterior gills of the Chinese crabEriocheir sinensis

The possible involvement of protein kinase C in control of ion transport was investigated on a preparation of isolated, perfused posterior gills of the Chinese crabsEriocheir sinensis (collected in 1989 from lakes near Emden, northern Germany) acclimated to fresh water. 1-oleyl-2-acetyl-sn-glycerol (OAG) and phorbol 12-myristate 13-acetate (PMA), two activators of protein kinase C, when added to the perfusion saline, induced depolarisation of the transepithelial potential difference (PD) and an increase in transepithelial Na⁺ influx. The observed increase was proportional to OAG concentration up to 250µM, with a 2.5× accelerated Na⁺ influx. OAG and PMA remained without effect on Cl^– fluxes. The observed effects were in agreement with an activation, via protein kinase C, of the Na⁺/K⁺ ATPase located on the serosal side of the epithelium.     

Inhibition of Na+/K+-ATPase and of active ion-transport functions in the gills of the shore crab Carcinus maenas induced by cadmium

Inhibition of Na⁺/K⁺-ATPase from gill plasma membranes of the shore crab Carcinus maenas by cadmium was investigated and compared with inhibitory effects by known antagonists (ouabain and Ca²⁺). For comparative considerations the Cd²⁺-inhibition of the enzyme from dog kidney was also tested. Na⁺/K⁺-ATPase from dog kidney and from crab gill differed greatly in sensitivity against ouabain. The inhibition constant K _i of the dog enzyme amounted to 9.1 × 10⁻⁷ mol l⁻¹, i.e. more than 300-fold smaller than the K _i of 2.9 × 10⁻⁴ mol l⁻¹ determined for the crab enzyme. Ca²⁺ inhibited the activity of Na⁺/K⁺-ATPase from crab gill plasma membranes with a K _i of 4.3 × 10⁻⁴ mol l⁻¹. The Na⁺/K⁺-ATPase from crab gill was inhibited by Cd²⁺ with a K _i of 9.1 × 10⁻⁵ mol l⁻¹. Cd²⁺ inhibited the Na⁺/K⁺-ATPase from dog kidney with a K _i (6.4 × 10⁻⁵ mol l⁻¹) comparable to that observed in the crab gill enzyme. Under experimental conditions Cd²⁺-inhibition of Na⁺/K⁺-ATPase was irreversible. Repeated washing, centrifugation and homogenization of the plasma membranes (four times) with Cd²⁺-free buffer did not restore any activity lost in the presence of 1 × 10⁻³ mol l⁻¹ Cd²⁺. Since ouabain-insensitive (nonspecific) ATPases in the plasma membrane fraction of crab gills were inhibited by Cd²⁺ in the same way as Na⁺/K⁺-ATPase, the heavy metal is considered as an unspecific ATPase inhibitor. Comparing these results with literature data on Cd²⁺-binding to electrophoretically separated proteins suggests that Na⁺/K⁺-ATPase is a Cd²⁺-binding enzyme. The results obtained on Na⁺/K⁺-ATPase were reflected by Cd²⁺-inhibition of the branchial ion-transport functions depending on this enzyme. The transepithelial short-circuit current of isolated gill half lamellae, a direct measure of area-specific active ion uptake, and the transepithelial potential difference of isolated, perfused whole gills, also indicative of active ion uptake, were inhibited by the heavy metal in a time- and dose-dependent mode. Remarkably these inhibitions were also irreversible. These findings are ecologically and biomedically significant: even when the actual environmental or tissue concentrations measured are low, biological microstructures such as Na⁺/K⁺-ATPase may accumulate the heavy metal by tight binding over prolonged periods until the first inhibitory effects occur. Received: 25 June 1997 / Accepted: 25 August 1997     

溴氰菊酯对菲律宾蛤仔体内酶活性和组织损伤的初步探索

采用不同质量浓度的溴氰菊酯(0.0070 mg·L~(-1)、0.014 mg·L~(-1)、0.020 mg·L~(-1)、0.027 mg·L~(-1))对菲律宾蛤仔进行20 d半静置染毒,测定不同时间淋巴液中乙酰胆碱酯酶(ACh E)和钠离子-钾离子-三磷酸腺苷酶(Na~+-K~+-ATPase)活性、鳃和肝脏中谷胱甘肽转硫酶(GST)活性的变化,并观察染毒20 d后鳃丝组织和消化盲囊组织的损伤情况。酶活性分析结果显示,与对照组相比,低浓度组(0.0070 mg·L~(-1))试验期间酶活性均无显著差异(P0.05);中浓度组(0.014 mg·L~(-1)、0.020 mg·L~(-1))淋巴液中ACh E和Na~+-K~+-ATPase均呈先激活后抑制的变化规律(P0.05),鳃和肝脏中GST活性均呈上升趋势(P0.05);高浓度组(0.027mg·L~(-1))淋巴液中ACh E和Na~+-K~+-ATPase、肝脏中GST活性在试验期间持续下降(P0.01),而鳃中GST活性呈先抑制后升高的趋势(P0.05)。研究表明,低中浓度的溴氰菊酯对菲律宾蛤仔体内的酶活性表现为先诱导后抑制,具有明显的时间、剂量效应;高浓度的溴氰菊酯对菲律宾蛤仔体内酶活持续抑制,且染毒浓度越高,组织细胞变异越显著,表现为鳃丝上皮细胞纤毛层萎缩、纤毛脱落,消化盲囊上皮细胞膨胀,出现包涵体样结构。