Hidden in the sand: Alteration of burying behaviour in shore crabs and cuttlefish by antidepressant exposure.

Pharmaceuticals such as antidepressants are constantly released into the aquatic environment. Consequently, fluoxetine (FLX) and venlafaxine (VEN), the active molecules of Prozac© and Effexor©, are detected up to several µg.L-1 in freshwater and marine coastal waters. Both compounds act on the serotoninergic system, which may result in behavioural impairment, especially in juvenile animals presumed to be more susceptible to low concentrations than adults. The objective of this study was to determine whether environmental concentrations of FLX alone or combined with VEN modulate innate burying behaviour in two juvenile marine invertebrates, i.e. Sepia officinalis and Carcinus maenas. Juvenile cuttlefish were exposed from hatching to 30 days post-hatching to either FLX alone (i.e. 5 ng.L-1) or in mixture with VEN (i.e. either 2.5 ng.L-1 or 5 ng.L-1 of each antidepressant). Juvenile crabs (<2 cm carapace width) were exposed for a period of 22 days to 5 ng.L-1 of FLX and a mixture of 5 ng.L-1 of FLX and VEN each. Several parameters of sand-digging behaviour were analysed weekly in both species. The occurrence of sand-digging behaviour decreased in cuttlefish exposed to a mixture of FLX and VEN at the lowest concentration (2.5 ng.L-1 each). Because sand-digging behaviour improved in controls, this decrease was likely to be related to a modification of maturation and/or learning processes. At the mixture of 5 ng.L-1 VEN and FLX each, a better body covering was observed in juvenile crabs. In both species, innate behaviour was modified under exposure to mixtures of FLX and VEN at environmentally realistic concentrations. These alterations were observed at an early developmental stage, when animals are particularly prone to predation. Hence, modified maturation of behavioural traits and, putatively, learning processes by exposure to pseudo-persistent antidepressants may affect the survival of these two species in the long term.

Trace metal concentrations in post-hatching cuttlefish Sepia officinalis and consequences of dissolved zinc exposure.

In this study, we investigated the changes of 13 trace metal and metalloid concentrations (i.e. Ag, As, Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, Se, V, Zn) and their subcellular fractionation in juvenile cuttlefish Sepia officinalis reared in controlled conditions between hatching and 2 months post-hatching. In parallel, metallothionein concentrations were determined. Our results highlighted contrasting changes of studied metals. Indeed, As and Fe concentrations measured in hatchlings suggested a maternal transfer of these elements in cuttlefish. The non-essential elements Ag and Cd presented the highest accumulation during our study, correlated with the digestive gland maturation. During the 6 first weeks of study, soluble fractions of most of essential trace metals (i.e. Co, Cr, Cu, Fe, Se, Zn) slowly increased consistently with the progressive needs of cuttlefish metabolism during this period. In order to determine for the first time in a cephalopod how metal concentrations and their subcellular distributions are impacted when the animals are trace metal-exposed, we studied previously described parameters in juveniles exposed to dissolved Zn at environmental (i.e. 50 µg l(-1)) and sublethal (i.e. 200 µg l(-1)) levels. Moreover, oxidative stress (i.e. glutathione S-transferase (GST), superoxide dismutase (SOD) and catalase activities, and lipid peroxidation (LPO)) was assessed in digestive gland and gills after 1 and 2 months exposures. Our results highlighted no or low ability of this stage of life to regulate dissolved Zn accumulation during the studied period, consistently with high sensitivity of this organism. Notably, Zn exposures caused a concentration-dependent Mn depletion in juvenile cuttlefish, and an increase of soluble fraction of Ag, Cd, Cu without accumulation modifications, suggesting substitution of these elements (i.e. Mn, Ag, Cd, Cu) by Zn. In parallel, metallothionein concentrations decreased in individuals most exposed to Zn. Finally, no perturbations in oxidative stress management were detected in gills, whereas modifications of GST, SOD and catalase activity levels were recorded in digestive gland, resulting in an increase of LPO content after a 6-week exposure to low Zn concentration. Altogether, these perturbations are consistent with previously described high sensitivity of juvenile cuttlefish towards Zn. Our results underlined the need to study deeply contamination impact on this animal at this stage of life.

Cryptic and biochemical responses of young cuttlefish Sepia officinalis exposed to environmentally relevant concentrations of fluoxetine.

Antidepressants released in the environment have the potential to generate neural disrupting effects in non-target organisms, yet their putative effects on behaviors have never been studied in cephalopod molluscs. This study assessed the impact of the antidepressant fluoxetine (FLX) on the efficiency of cryptic behaviors (body patterns on uniform, checkerboard and sandy substrates), locomotor activity, and brain chemistry in young cuttlefish exposed to environmental concentrations (1 and 100ngL(-1) of FLX) during the perinatal period. Behavioral responses of cuttlefish were monitored at hatching and two weeks later, and brain monoamine contents were quantified at one month of age. FLX significantly altered the camouflage efficiencies on uniform and sandy backgrounds only at the lowest concentration, but not at 100ngL(-1). Hatchlings exposed to 1ngL(-1) of FLX exhibited a duration exposure-dependent decrease in the uniform camouflage. They also showed a significant increase of the frequency of sand digging behaviors which might make them highly visible to predators in nature. When tested again two weeks later, cuttlefish seemed to have recovered and no more behavioral alterations were observed showing a transitory effect of the antidepressant. FLX did not affect the levels of serotonin, norepinephrine and their metabolites; however, it seemed to influence dopaminergic activity between the two FLX-exposed groups. The results show for the time that environmentally realistic concentrations of a single SSRI significantly impair the cryptic performances of newly hatched cuttlefish, and may ultimately reduce their chance for survival.

Effects of perinatal exposure to waterborne fluoxetine on memory processing in the cuttlefish Sepia officinalis.

Recent ecotoxicological studies highlight the increasing presence of pharmaceuticals discharged in the aquatic environment. Amongst them is the antidepressant fluoxetine (FLX), a selective serotonin reuptake inhibitor, primarily indicated for treatment of depression. The effect of chronic exposure to FLX on memory processing in 1-month-old cuttlefish Sepia officinalis was evaluated. Three groups of new-borns were reared in different conditions: one control group (no FLX) and two groups exposed to environmental concentrations of FLX (1 and 100ng/L) from 15 days pre-hatching to 1 month post-hatching. Acquisition and retention performances were assessed using the 'prawn-in-the-tube' procedure. Perinatal exposure to fluoxetine led to significant changes in memory processing of the animals. The lowest observed effect concentration of this antidepressant on learning and retention was 1ng/L which is under the range of environmental contamination. Cuttlefish exposed at low FLX concentration had impaired acquisition capabilities and animals exposed at high FLX concentration displayed a deficit of memory retention compared to the control group that had nonimpaired initial acquisition and retention performances. The results subsequently suggested that FLX-induced changes in cognitive capacities could potentially lead to inappropriate predatory behaviors in the natural environment. The study provides the basis for future studies on how pharmaceutical contaminants disrupt cognition in ecologically and economically relevant marine invertebrates.

Sexually mature cuttlefish are attracted to the eggs of conspecifics.

Cuttlefish typically are solitary, but form aggregations to spawn. We tested the hypothesis that bioactive factors in the eggs of conspecifics may facilitate the formation of spawning groups of Sepia officinalis. Cuttlefish detected odors from cuttlefish eggs, resulting in an increased ventilation rate. Extracts from female ovaries induced the largest increase in ventilation rate, suggesting that this organ could be a potential source of the bioactive chemicals. In y-maze assays, sexually mature, but not subadult, cuttlefish, were attracted to odors of cuttlefish eggs. These data suggest that Sepia eggs could be a source of reproductive pheromones.

Metal concentrations in digestive gland and mantle of Sepia officinalis from two coastal lagoons of Portugal.

Concentrations of both essential (Fe, Cu, Zn) and non essential (Cd, Hg and Pb) metals were measured in the digestive gland and mantle of female cephalopods Sepia officinalis captured in two distinct lagoons in Portugal: Aveiro Lagoon, with a history of anthropogenic and industrial pollution, and Formosa Lagoon receiving urban effluents. We provide evidence for the following: (1) the digestive gland is the main target organ for both essential and non essential metals, frequently containing concentrations few orders of magnitude higher as compared to mantle; the sole exception from this was the Hg that is equally distributed in the two tissues; (2) unexpectedly, the higher levels of metals were found in animals captured in the less polluted lagoon, except for Cd whose bioavailability in Aveiro lagoon might be related to industrial sources, while the influence of Cd speciation in local pray composition should not be ruled out (3) size influenced metal concentration in different way: smaller individuals accumulated significantly more Cu, while Hg concentrations showed the opposite trend; (4) Cd is positively correlated to Zn and Cu in digestive gland of specimens collected in spring in Aveiro Lagoon, and no relationship was found in Formosa Lagoon; (5) the molar ratios Cd:Zn and Cd:Cu in digestive gland increased with body weight in specimens from Aveiro area, both ratios becoming particularly higher in older individuals. Metal-specific accumulation patterns in both mantle and digestive gland at the two sites are discussed in the light of their toxicological implications.