High-Throughput Screening of Psychotropic Compounds: Impacts on Swimming Behaviours in Artemia franciscana.

Animal behaviour is becoming increasingly popular as an endpoint in ecotoxicology due to its increased sensitivity and speed compared to traditional endpoints. However, the widespread use of animal behaviours in environmental risk assessment is currently hindered by a lack of optimisation and standardisation of behavioural assays for model species. In this study, assays to assess swimming speed were developed for a model crustacean species, the brine shrimp Artemia franciscana. Preliminary works were performed to determine optimal arena size for this species, and weather lux used in the experiments had an impact on the animals phototactic response. Swimming speed was significantly lower in the smallest arena, whilst no difference was observed between the two larger arenas, suggesting that the small arena was limiting swimming ability. No significant difference was observed in attraction to light between high and low light intensities. Arena size had a significant impact on phototaxis behaviours. Large arenas resulted in animals spending more time in the light side of the arena compared to medium and small, irrespective of light intensity. The swimming speed assay was then used to expose specimens to a range of psychotropic compounds with varying modes of action. Results indicate that swimming speed provides a valid measure of the impacts of behaviour modulating compounds on A. franciscana. The psychotropic compounds tested varied in their impacts on animal behaviour. Fluoxetine resulted in increased swimming speed as has been found in other crustacean species, whilst oxazepam, venlafaxine and amitriptyline had no significant impacts on the behaviours measured. The results from this study suggest a simple, fast, high throughput assay for A. franciscana and gains insight on the impacts of a range of psychotropic compounds on the swimming behaviours of a model crustacean species used in ecotoxicology studies.

Species-specific behaviours in amphipods highlight the need for understanding baseline behaviours in ecotoxicology.

Behavioural studies in ecotoxicology are increasing with techniques and endpoints used in pharmacology being translated to other vertebrate and invertebrate species. Despite this, data on the baseline behaviours of model organisms, and inter-species variability in behaviour are currently under-studied. This study assessed a range of behaviours associated with anxiety including swimming speed, phototaxis and thigmotaxis in a marine and freshwater amphipod (Echinogammarus marinus and Gammarus pulex). Differences in sensitivity to these assays were observed between species with E. marinus showing a greater sensitivity to the phototaxis assay than G. pulex, while in thigmotaxis assays G. pulex appeared better suited than E. marinus for measuring differences in the use of central zones. Significant inter-species differences were also observed in swimming patterns when breaking the data into ten second time bins but not when data was broken into two-minute time bins. The results of this study provide evidence of phototactic and thigmotactic behaviours in two model crustacean species with potential for use in behavioural ecotoxicology. Inter-species variability in sensitivity to behavioural assays highlights the importance of systematic assessment of baseline responses for all model species used in behavioural studies. Careful analysis of data is also required when performing behavioural studies so as not to lose sensitivity in your data.

Shape and size of the arenas affect amphipod behaviours: implications for ecotoxicology.

The use of behaviour in ecotoxicology is expanding, however the lack of standardisation and validation of these assays currently presents a major drawback in moving forward in the development of behavioural assays. Furthermore, there is a current paucity of control data on test species, particularly invertebrate models. In this study we assessed a range of behaviours associated with spatial distribution and locomotion in relation to arena size and shape in two species of amphipod crustacean (Echinogammarus marinus and Gammarus pulex). Arena shape had significant effects on almost all behavioural parameters analysed. Increasing arena size resulted in an increased mean velocity and activity plus increased proportional use of the central zones. These results indicate that 'ceiling effects' may occur in some ecotoxicological studies resulting in potentially 'false' negative effects if careful consideration is not paid to experimental design. Differences in behaviours were observed between the two species of amphipod. For example, G. pulex spend approximately five times (∼20%) more of the available time crossing the central zones of the arenas compared to E. marinus (∼4%) which could have implications on assessing anxiolytic behaviours. The results of this study highlight several behaviours with potential for use in behavioural ecotoxicology with crustaceans but also underscore the need for careful consideration when designing these behavioural assays.