The MazeEngineers tap-elicited swim test is used in free swimming zebrafish to evaluate the effects of EtOH, drugs and toxins on the learning process (non associative). Frequently measured behaviors include the “C-start” response, which increases in latency in early alcohol exposure. The MazeEngineers Tap Swim apparatus comes with 8 easy to use arrays that attaches to an automated tap array underneath. This array is controlled with the Conductor software (free of charge) to control interval taps and timing between interval taps.
Key variables that can be controlled include:
- Stimulus pattern: Number of taps within interval time
- Interstimulus interval
Integration with Conductor allows for access to Noldus Ethovision control of taps with the Noldus Baton. Set taps depending on zebrafish behavior.
Price & Dimensions
Tap Arenas (8)
- Number of Arenas: 2×4 array.
- Individual Arena: 6cm diameter, 9cm height.
- Separators (12cm height).
- Tap controlled by Conductor, 8 independent controls
Frequently Asked Questions
- Can I order more than 8 Arrays? Less?
- Yes, please let us know and we can customize an array for you
- Can I change the height and diameter of the arrays?
- Yes, please let us know. We need to ensure that the tap provides a consistent force, however, if you have a modification within 25% of the original sizing the tap force will be constant
The tap-elicited startle response in zebrafish is a reliable and easily amenable neurobehavioral assessment to perform toxicological and pharmacological studies (Eddins et al., 2010). The test enables the determination of effects on swimming activity related to muscle function and disease (Lebold et al., 2013, Sztal et al., 2016) and alterations in short-term memory (Levin and Cerutti, 2009).
The equipment typically consists of a 2 x 4 array of chambers with opaque separations to prevent visual contact between different tanks. Groups of eight fish can be tested simultaneously, with one fish being tested in each tank. An automated solenoid placed underneath the tanks is recommended in order to generate the tap(s). An additional automated system should be used to control number of taps and timing between trial blocks. Moreover, a video camera connected to a tracking software can be used to record swimming behavior.
The experimental protocol entails an initial acclimatization to the test room and the chambers. Assessment of baseline response and habituation to closely spaced or multiple taps is subsequently done. Test trials are performed on set timepoints, whose timing depends on the aim of the study. Responses to single or multiple taps can be studied.
The use of zebrafish enables the high-throughput study of neurodevelopmental defects induced by exposure to toxicants – such as alcohol (Carvan et al., 2004) – or neurological diseases, as well as drug screening. Compared to mammals commonly used in research, zebrafish present the advantages of easy reproduction in great numbers, fast development and little cost of experimentation. Furthermore, superior extrapolation is possible due to being a vertebrate
Apparatus and Equipment
We recommend testing fish individually in a test battery consisting of 2 x 4 array of cylindrical chambers/tanks. Dimensions can vary – Sztal et al. (2016) used 50-mm diameter tanks, whereas Eddins et al (2010) used 60 mm in diameter and 90 mm height, filled up to 20 mm. Tank water should consist of deionized water with sea salts. Opaque-white dividers should be used so that each fish is not able to see the other chambers (Eddins et al., 2010). An automated solenoid should be attached underneath the tanks to generate the tap(s). Moreover, the investigator should use an additional automated system to control number of taps and timing between trial blocks. Swimming behavior can be recorded with a video camera placed centrally above the array and connected to a tracking software for optimal data analysis. Placement of dividers should not create a visual barrier preventing successful video recording.
As with other neurobehavioral assessments, the experiment should be performed blinded to minimize experimental artifacts. In order to achieve reproducible and reliable results, time of the day, lighting conditions and water temperature should be standardized (Sztal et al., 2016). Fish should be brought to the test room in their home tanks. Constant filtration and aeration of home tanks should be ensured. Eddins et al. (2010) allowed 10 min of acclimatization to arenas before testing, whereas Lebold et al. (2013) allowed 24 hr.
Fish can be tested in groups up to 8 with one fish per chamber. Software should enable recording of fast swimming activity – Eddins et al. (2010) recorded fish location six times per second. Acceleration peaks should be observed immediately after taps (Sztal et al., 2016). In a study aiming to analyze the developmental effects of exposure to a toxic agent or drugs, Eddins et al. (2010) assessed startle responses in 10 trials with 1 min intertrial intervals to determine baseline response and habituation with repeated testing. Subsequent trials at 8, 32 and 128 min were done to measure reestablishment of the startle response without closely spaced trials. Conversely, in a study evaluating long-term effects of nutritional deficiencies, Lebold et al. (2013) performed three trials of 6 min without stimulus to evaluate baseline swimming behavior. Results from each fish were averaged and two further trials were then conducted; the first trial used single tap stimulation, whereas the second trial used multiple taps – one every 5 sec for 90 sec (18 taps total). Swimming speeds and distances were recorded. Fish were allowed to rest for 1 hr between single and multiple tap trials.
Different inter-block intervals can be used in the same study. Aiming at assessing short-term memory, Levin and Cerutti (2009) adjusted stimulus rates, with 20 taps presented with an inter-block interval of 10 sec, followed immediately by 20 additional taps with an inter-block interval of 20 sec.
Besides analyzing startle response through distance swam, touch-evoked tests can also be used to assess swimming kinematics, such as shape and speed of the body wave during the swimming motion (Muller and van Leeuwen, 2004), to obtain a quantitative measurement of the locomotor behavior.
Data analysis on the tap-elicited swim test typically consists in comparisons of distance swam (Eddins et al., 2010). Results can be shown as variations throughout the trials (A), distance swam in specific trials during or after (B) the trial block, or average of the trials within the same block (C). Although distance units are commonly cm/5 sec (Eddins et al., 2010), total distance swam per minute post-tap can also be used (Lebold et al., 2013). Results are shown as mean ± standard error of the mean:
Strengths & Limitations
Tap-elicited swim test enables toxicological and pharmacological studies, including developmental assessments, and tests of muscle function and short-term memory (Levin and Cerutti, 2009, Sztal et al., 2016). Comparisons of different genetic backgrounds or studies of the effects of genetic manipulation are also possible.
The use of zebrafish enables the study of neurodevelopmental processes with the benefit of optical clarity, while affording superior extrapolation as a vertebrate. This model increases experimental throughput, while it decreases cost of experimentation in comparison to mammals, as fish are easily bred in great numbers and do not require a large housing space. In addition, development is extremely fast, as spontaneous muscle contraction is observed by 24 hours post-fertilization, whereas controlled swimming behaviors are exhibited by 48 hours post-fertilization (Sztal et al., 2016)
Caution should be taken when studying the effects of drugs in zebrafish delivered via immersion, as issues with water solubility and pharmacokinetics concerning translational validity have been raised (Bailey et al., 2015).
Furthermore, in studies assessing alterations in memory, neurobehavioral studies in zebrafish do not enable the elucidation of the role of discrete central nervous system areas, although existent structures in the zebrafish brain appear to be biochemically and functionally similar to the mammalian brain to permit neuropharmacological manipulations (Bailey et al., 2015). Special attention should be paid to experimental design, so that it is clear if the differences reported result from changes in cognition or from motor effects (Bailey et al., 2015).
Summary and Key Points
- Tap-elicited swimming test is an easy to use test for non-associative learning
- The equipment required typically consists of a 2 x 4 array of chambers with opaque dividers preventing visual contact between different tanks. An automated solenoid is used underneath the tanks to generate the tap(s) and an additional automated system is recommended to control number of taps and timing between trial blocks. Swimming behavior can be recorded with a video camera connected to a tracking software.
- The experimental protocol typically consists of an initial analysis to obtain a baseline response and assess habituation to closely spaced taps. Subsequent trials are performed on set timepoints depending on the aim of the study. Furthermore, responses to single or multiple taps can be compared.
- Results are typically reported as distance swam/5 sec after tap(s), although variations have been reported, including distance swam per minute for each minute of the trial block. Swimming speed or kinematics have also been analyzed in tap-elicited tests.
Eddins D, Cerutti D, Williams P, Linney E, Levin ED (2010) Zebrafish provide a sensitive model of persisting neurobehavioral effects of developmental chlorpyrifos exposure: comparison with nicotine and pilocarpine effects and relationship to dopamine deficits. Neurotoxicology and teratology 32:99-108.
Lebold KM, Lohr CV, Barton CL, Miller GW, Labut EM, Tanguay RL, Traber MG (2013) Chronic vitamin E deficiency promotes vitamin C deficiency in zebrafish leading to degenerative myopathy and impaired swimming behavior. Comparative biochemistry and physiology Toxicology & pharmacology : CBP 157:382-389.
Sztal TE, Ruparelia AA, Williams C, Bryson-Richardson RJ (2016) Using Touch-evoked Response and Locomotion Assays to Assess Muscle Performance and Function in Zebrafish. Journal of visualized experiments : JoVE.