The Tail Suspension Test (TST) is a key behavioral assay used in pharmacological research to study antidepressant-like effects in mice. This test helps researchers evaluate how potential antidepressant drugs influence behavior, offering insights into their efficacy. In this blog post, we’ll explore what the TST is, why it’s valuable, and how to conduct it properly in a pharmacy lab, including a practical protocol to ensure reliable results.
What is the Tail Suspension Test?The TST involves suspending a mouse by its tail in a controlled setting, observing its behavior as it responds to this mildly stressful situation. When suspended, a mouse typically alternates between struggling to escape and periods of immobility, where it hangs passively. This immobility is considered a model of “behavioral despair,” resembling depressive-like states. Antidepressant drugs often reduce the time spent immobile, suggesting potential therapeutic effects.The test is widely used because it’s straightforward, cost-effective, and provides consistent results for screening compounds. It’s particularly useful in pharmacy labs for testing new drugs or studying the mechanisms of existing antidepressants. However, the TST is a simplified model and doesn’t fully capture the complexity of human depression, so results should be interpreted alongside other tests.
Why Use the Tail Suspension Test?
- Screening Tool: The TST helps identify compounds with antidepressant potential by measuring reductions in immobility time.
- Efficiency: It allows testing multiple mice in a short time, ideal for high-throughput drug screening.
- Predictive Value: Many clinically effective antidepressants, like SSRIs or tricyclics, show activity in the TST.
- Minimal Equipment: The setup is simple, requiring basic lab tools and optional automated systems for scoring.
How to Perform the Tail Suspension Test in a Pharmacy LabConducting the TST requires careful planning to ensure ethical treatment of animals and reliable data. Below is a step-by-step guide to performing the test in a pharmacy lab, followed by a recommended protocol.Materials Needed
- Suspension Setup: A sturdy horizontal bar or shelf (about 60 cm long, 1 cm wide) elevated 20–30 cm above a padded surface to prevent injury if a mouse falls.
- Adhesive Tape: Medical-grade tape (1.5–2 cm wide) strong enough to hold a mouse but gentle for easy removal.
- Optional Cylinders: Small plastic tubes (4 cm long, 1 cm diameter) to prevent mice from climbing their tails, which some strains may do.
- Camera: A high-resolution video camera for recording behavior, ideally paired with software for automated analysis.
- Cleaning Supplies: A mild, unscented disinfectant (e.g., 1% chlorhexidine) to clean the apparatus between tests.
- Testing Area: A quiet, well-lit room with partitions or separate chambers to test multiple mice without visual or auditory interference.
- Prepare the Animals:
- Use adult mice (e.g., 6–12 weeks old) from a strain known to respond well in the TST, such as C57BL/6 or BALB/c. Avoid strains with excessive tail-climbing tendencies unless using cylinders.
- House mice in a controlled environment (12-hour light/dark cycle, stable temperature) and acclimate them to the testing room for at least 60 minutes to reduce stress.
- Divide mice into groups: a control group (receiving saline or vehicle) and test groups (receiving the drug being studied, administered 30–60 minutes before testing via injection or oral gavage).
- Set Up the Apparatus:
- Secure the horizontal bar at 20–30 cm above a soft surface. If testing multiple mice, use partitions (e.g., 15 cm wide x 50 cm high) to prevent them from seeing each other.
- Set up the camera to capture clear footage of the entire test area. Ensure consistent lighting (avoid shadows) for accurate video analysis.
- Cut adhesive tape into strips (15–20 cm long) and mark a 2 cm section for attaching to the mouse’s tail.
- Conduct the Test:
- Gently pick up a mouse by the base of its tail and attach the tip (2–3 mm) to the bar using adhesive tape. If needed, slide a plastic cylinder onto the tail base to prevent climbing.
- Suspend the mouse for 6 minutes. The first 2 minutes allow the mouse to adjust; focus scoring on the last 4 minutes for consistency.
- Test one mouse at a time or use partitions to test multiple mice simultaneously, ensuring they cannot interact.
- Record the session using a video camera for later analysis.
- Score the Behavior:
- Define immobility as periods when the mouse hangs passively without active struggling, excluding small twitches or pendulum-like swings caused by momentum.
- Measure the total immobility time (in seconds) during the 6-minute test or the last 4 minutes. Automated software is ideal for precision, but manual scoring can work with trained observers.
- Compare immobility times between control and test groups to evaluate drug effects. Lower immobility in the test group suggests antidepressant-like activity.
- Post-Test Procedures:
- Carefully remove the tape to avoid injuring the tail and return the mouse to its home cage.
- Clean the bar and any cylinders with disinfectant to eliminate odors that could affect subsequent tests.
- Review videos for anomalies (e.g., tape detachment or excessive climbing) and exclude invalid trials from analysis.
- Use the minimum number of mice necessary, following institutional guidelines (e.g., IACUC or equivalent).
- Limit testing to one session per mouse to avoid habituation or excessive stress.
- Ensure tape and suspension methods do not cause injury. Check tails post-test for signs of discomfort.
- Combine TST results with other behavioral tests (e.g., open-field test for locomotor activity) to confirm findings, as immobility alone may not fully reflect antidepressant effects.
- Strain Selection: Choose strains with consistent TST responses. For example, BALB/c mice are sensitive to SSRIs, while some strains may require cylinders to prevent climbing.
- Automation: Invest in software (e.g., video tracking systems) for objective scoring, especially for large studies.
- Consistency: Standardize handling, timing, and environmental conditions (e.g., noise, lighting) to reduce variability.
- Validation: Use a known antidepressant (e.g., fluoxetine) as a positive control to validate your setup.
- 6-Minute Test: Score immobility in the last 4 minutes to account for initial adaptation.
- Single Use per Mouse: Avoid retesting to prevent behavioral changes.
- Automated Scoring: Use video analysis software for objectivity, with manual scoring as a backup.
- Controlled Environment: Minimize noise, standardize lighting, and use partitions for multiple mice.
- Ethical Standards: Follow animal welfare guidelines and monitor mice for stress or injury.