Maaike Nieveen

PCBs and the energy cost of migration in the European eel (Anguilla anguilla L.)

The effect of polychlorinated biphenyls (PCBs) on the energy consumption of fasting silver European eel (Anguilla anguilla L.) was studied over a 27-day period during which the animals were at rest or were swimming 800 km in Blazka swim tunnels. Three-year-old female hatchery eels (silver stage) between 73 and 80 cm long weighing around 1 kg were dosed intraperitoneally with PCBs at a nominal dosage of 10x the consumption standard as a mixture representative for planar (7 microg PCB126/kg eel), non-planar (5 mg PCB153/kg eel) and metabolizable PCBs (50 microg PCB77/kg eel) found in wild eel, or only with the vehicle (corn oil, 10 ml/kg eel). Four major observations were made: (1) PCB-exposed animals lose less weight compared to their unexposed controls; (2) PCB-concentrations on a lipid basis are 2.8-14 times higher in swimming compared to resting animals; (3) the standard metabolic rate is significantly lower in the PCB-exposed animals than in unexposed controls. In addition, PCB-exposure significantly reduces oxygen consumption during swimming, and starting at 400 km (18 days) this effect increases with time; (4) the relative spleen and liver weight significantly increased in the PCB-swim animals but not in the PCB-rest animals. The swimming animals lost about 75% more weight compared to resting animals and had about 50% lower plasma fat content. Hematocrit, haemoglobin, plasma pH, ion levels (sodium and potassium), and plasma lactate were not affected by PCB-exposure or swimming. Apparently, the current levels of PCBs and other dioxin-like compounds may seriously impair the reproduction of the European eel.

Energy status and free fatty acid patterns in tissues of common carp (Cyprinus carpio, L.) and rainbow trout (Oncorhynchus mykiss, L.) during severe oxygen restriction

Common carp and rainbow trout were exposed to a severe level of oxygen restriction up to a near lethal value, to study the occurrence of tissue damage. Rainbow trout lost equilibrium at a PO2 of 3.2 kPa, whereas carp were able to survive 1.5 hr of anoxia. In both species, the anaerobic metabolism was significantly activated and the energy status (PCr, ATP and energy charge) was significantly depressed in brain, liver, and red and white muscle. No marked release of PUFA to the FFA pool was observed, while membrane leakage was not increased as evidenced by plasma LDH-activity. These results indicate the absence of a marked hydrolysis of membrane lipids. Thus, even after a near lethal exposure to hypoxia or anoxia, no tissue damage occurs in fish liver and skeletal muscles. The changes of the FFA patterns in the skeletal muscles and liver of both species after oxygen deprivation may be related to changes in desaturase activities, a reduction of lipolytic activity and PUFA metabolism.

Neurotransmitter levels and energy status in brain of fish species with and without the survival strategy of metabolic depression

Abstract The effects of anoxia were studied in the whole brain of three fish species, each with a specific metabolic strategy for anoxic survival. Goldfish (Carassius auratus) combine a lactate to ethanol conversion with a metabolic depression, tilapia (Oreochromis mossambicus) use an anaerobic glycolysis with the strategy of metabolic depression, and carp (Cyprinus carpio) use an increased anaerobic glycolysis for energy production. Tilapia and carp were exposed to anoxia until they lost equilibrium and exhibited escape reactions, this occurred after 2 hours of anoxia for tilapia and 30 minutes of anoxia for carp. Goldfish were exposed to a selected period of 8 hours anoxia. The energy status and neurotransmitter (amino acid) levels in whole brain tissue were measured after anoxia exposure. The energy status was affected in all three groups exposed to anoxia. Lactic acid levels increased five- to six-fold in all three groups. No direct correlation was observed between energy status and survival strategy. Remarkably, the changes in the amino acid patterns in whole fish brains show the greatest changes in the anoxia-tolerant goldfish, an intermediate pattern in tilapia, and nearly no changes in the anoxia-intolerant carp. The changes in amino acid are probably dependent on the period of anoxia exposure. For goldfish, the lactate-ethanol conversion primarily determines anoxic survival, but the strategy of metabolic depression observed in goldfish and tilapia may contribute secondarily to anoxic tolerance. It is hypothesized that a decrease of excitatory neurotransmitters (mainly glutamate), in combination with an increase of inhibitory neurotransmitters (mainly GABA), may contribute to the process of metabolic depression and prolong survival.

Temporal expression of hepatic estrogen receptor 1, vitellogenin1 and vitellogenin2 in European silver eels

Because European silver eels have never been caught during or after their 6000-km reproductive migration to the Sargasso Sea, all existing knowledge on their sexual maturation comes from hormonal stimulation. Silver eels that start their oceanic migration are still immature with pre-vitellogenic oocytes. Hence we assumed that vitellogenesis should start with the expression of the estrogen receptor in the liver before the circulating 17beta-estradiol (E2) can have any effect. In this study we followed the hepatic vitellogenesis upon 4 weekly injections with carp pituitary extracts (CPE). New molecular primers for the expression of the estrogen receptor 1 (esr1), vitellogenin1 (vtg1) and vitellogenin2 (vtg2) in the liver were developed. Sequences of vtg2 and esr1 were not previously described in Anguilla anguilla. All eels showed weekly increase of the eye size and pectoral fin length, which are signs of early maturation. The same occurred with the gonadosomatic index, the oocyte stage and diameter, and number of deposited fat droplets. Early vitellogenesis appeared as a 3-step process (1) E2-levels and esr1 expression were significantly increased already after one injection, (2) vtg1 and vtg2 expression were significantly increased after one and two injections, respectively, and (3) vtg1 and vtg2 expression increased further after three and four injections. Then also plasma calcium (corresponds with plasma vitellogenin) increased and yolk globuli appeared in the oocytes. These results show that esr1 is the first of the three genes examined that is expressed during the onset of hepatic vitellogenesis. Furthermore, ovarian vitellogenesis (appearance of yolk globuli in oocytes) occurs 1-2 weeks later than the onset of hepatic vitellogenesis.

Male silver eels mature by swimming

BackgroundIf European silver eels are prevented from reproductive migration, they remain in a prepubertal stage by dopaminergic inhibition of pituitary activity. Because this inhibition is likely a requirement for an extended female growth stage, we tested if it is sex-specific by subjecting both sexes to stimulation by GnRHa (Gonadotropin-Releasing Hormone agonist) – injection or 3-months swimming in seawater.ResultsIn contrast to females, males showed a two- to three-fold higher LHβ (luteinising hormone β subunit) – expression, a three- to five-fold higher GSI (Gonadosomatic index) and induced spermatogenesis when compared with the untreated control group.ConclusionDopaminergic inhibition is thus not effective in males and swimming results in natural maturation, probably via GnRH-release.

Influence of Hypoxia Exposure On the Energy Metabolism of Common Carp (Cyprinus Carpio L.)

Hypoxic conditions is a common adverse environmental condition in an aquatic environment. To study the responses of fish to this phenomenon common carp (Cyprinus carpio L.) were exposed to a graded hypoxia load and the oxygen consumption was measured continuously. At 30%AS (air saturation) 20%AS, 10%AS, 5%AS and 3%AS blood, liver and white muscle samples were collected. In the blood, haematological parameters, substrates (FFA, lactate and glucose), as well as the stress hormone cortisol were measured. High-energy phosphorylated compounds and lactate were measured in liver and white muscle tissue. During hypoxia, ATP concentrations and the adenylate energy charge (AEC) remained constant in white muscle, whereas both declined in liver tissue. The critical oxygen tension, which reflects the onset of a physiological or biochemical response at a certain hypoxia load, indicates that rapid changes were recorded in the blood, followed by the liver while white muscle (except for phosphocreatine (PCr)) is rather insensitive to environmental hypoxia. It is concluded that the impaired oxidative phosphorylation is compensated by the creatine kinase equilibrium reaction (depletion PCr pool) and the anaerobic glycolysis (lactate production).

Swimming suppresses hepatic vitellogenesis in European female silver eels as shown by expression of the estrogen receptor 1, vitellogenin1 and vitellogenin2 in the liver

BackgroundWhen European silver eels (Anguilla anguilla) venture into the Atlantic Ocean for their 6,000 km semelparous spawning run to the Sargasso Sea, they are still in a prepubertal stage. Further sexual development appears to be blocked by dopaminergic inhibition of hypothalamus and pituitary activity. Recently, we found that swimming for several weeks in freshwater stimulated the incorporation of fat droplets in the oocytes. So, it was hypothesized that long term swimming in seawater would release the inhibition further and would also stimulate the production of vitellogenin by the liver.MethodsFor this study a swim-flume was constructed to allow simulated migration of migratory female silver eels for 3 months (1,420 km) in natural seawater at 20 degrees C. Primers were designed for polymerase chain reactions to measure the mRNA expression of estrogen receptor 1 (esr1), vitellogenin1 (vtg1) and vitellogenin2 (vtg2) genes in the liver of European female silver eels.ResultsIn comparison to resting eels, swimming eels showed a diminished expression of esr1, vtg1 and vtg2 in the liver. They also had lower plasma calcium (Ca; indicative of vitellogenin) levels in their blood. This showed that vitellogenesis is more strongly suppressed in swimming than in resting eels. However, when eels were subsequently stimulated by 3 weekly carp pituitary extract injections, the expression of the same genes and plasma levels of Ca strongly increased in both groups to similar levels, thus equalizing the initial differences between resting and swimming.ConclusionsIt is concluded that vitellogenesis remains suppressed during resting and even more during swimming. The fact that swimming stimulates fat deposition in the oocytes but suppresses vitellogenesis indicates that these events are separated in nature and occur sequentially. Swimming-suppressed vitellogenesis may imply that in nature eels undergo vitellogenesis and final maturation near or at the spawning grounds.

Hypoxia-induced changes in hemoglobins of Lake Victoria cichlids

ABSTRACT In a previous study, broods of the Lake Victoria cichlid Haplochromis ishmaeli raised under hypoxic or normoxic conditions showed striking differences in isohemoglobin (isoHb) pattern that were not observed in two other cichlids that do not belong to the Lake Victoria species flock. We therefore hypothesized that the adaptive mechanism seen in H. ishmaeli in response to hypoxia constitutes a trait that the Lake Victoria species flock inherited from ancestors that lived in hypoxic environments. We tested this hypothesis by designing split-brood experiments with three other representative species from the same species flock: the insectivorous Haplochromis thereuterion, the mollusk-shelling Platytaeniodus degeni and the zooplanktivorous Haplochromis piceatus, while keeping H. ishmaeli as a reference. Split broods were raised, under either normoxia or hypoxia. All hypoxia-raised (HR) individuals of each of the four species exhibited a distinctly different isoHb pattern compared with their normoxia-raised (NR) siblings. The hemoglobin of HR H. thereuterion showed higher O2 affinity compared with NR siblings particularly in the presence of ATP and GTP, indicating that blood of HR juveniles has significantly improved O2-binding affinity under hypoxic conditions. We also tested the capacity to acclimate at greater age in two species by reversing the O2 condition after 7 (H. thereuterion) and 4 (H. ishmaeli) months. After reacclimation for 1 and 2 months, respectively, we found incomplete reversal with intermediate isoHb patterns. As three of the four species do not encounter hypoxic conditions in their environment, this unique trait seems to be a relic inherited from predecessors that lived in hypoxic environments. Summary: Four cichlids of the Lake Victoria species flock raised under hypoxia exhibit a remarkable change in isohemoglobin pattern, corresponding with improved hemoglobin O2-binding affinity.