Jan 14, 2025

Public workspaceForced Swim Test Protocol

DOI
dx.doi.org/10.17504/protocols.io.n2bvj9qnblk5/v1
  • 1University of Alabama at Birmingham
Ashley Adamson
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DOI: dx.doi.org/10.17504/protocols.io.n2bvj9qnblk5/v1
Protocol Citation: Ashley Adamson, Allie Smith, Briana R De Miranda 2025. Forced Swim Test Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.n2bvj9qnblk5/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: January 13, 2025
Last Modified: January 14, 2025
Protocol Integer ID: 118263
Funders Acknowledgements:
Briana De Miranda
Grant ID: R01ES034846
Briana De Miranda
Grant ID: TX220241
Ashley Adamson
Grant ID: F31 ES036890
Abstract
The following protocol is used to assess behavioral despair or learned helplessness using the University of Alabama at Birmingham's Neuroscience Behavioral Assessment Core.
Guidelines
The use of this protocol requires approval by the user's Institutional Animal Care and Use Committee (IACUC) or equivalent ethics committee.
Materials
  1. Chlorhexidine
  2. Paper towels
  3. Cylindrical tanks
  4. Clean water
  5. Ethovision version 17 software
  6. Recovery cages
  7. Heating pad
  8. Fish net
Before start
You will need a test mouse whose behavior is not included in the experiment to calibrate detection settings.
Set Up
Set Up
Bring animals into testing room for habituation (at least 30 minutes prior to start of experiment)
Set up test cylinders
  • Cylinder should be at least 50 cm tall and 20 cm wide for mice
Clean the cylinder (wipe out and spray with chlorohexidine)
Set up the number of cylinders you want to use (4 fit well under our camera setup)
Fill with clean water
  • Should be at 21° C
  • Fill water to level where subject cannot reach the bottom of the cylinder, but below level from which they could jump out
Make sure the EthoVision version 17 software dongle is in the computer
  • Shines a red light when working
Open EthoVision version 17 software
Start new experiment – New from Template
Name it
Tell it where to save
Experiment Settings
Select live tracking
  • Select the camera to be used
  • Center your arena under the camera
Select center point detection
Select the number of arenas that you will be using
Units should be cm, s, and deg
Arena settings
Grab a background image from the camera
  • Ensure the cylinders are in their intended area, moving them after this step will affect analysis
  • Mark placement with tape so the maze can be returned to the exact same location if accidentally between trials
Draw scale
  • Measure the width of one cylinder
  • Draw a scale bar over the area that you measured and indicate its length
Click first arena in the list on the right
  • Select the ellipse drawing tool
  • Trace one cylinder
  • Drag arena label inside ellipse
  • Repeat for every cylinder in the experiment
Validate setup
  • If there are any overlapping zones, they will be considered invalid and must be redrawn without overlapping
Trial Control Settings
Begin condition should be on the start button with the condition of center-point being in the arena
Stop track with the condition being after 6 minutes
Detection Settings
Click advanced settings
Dynamic response
Select appropriate setting to indicate if subject is darker or lighter than the background
Place a test subject in the maze
Click and drag around the subject to create a box around the subject
The software will create an automatic threshold around the subject that moves with it
Fine tune it with the scrub bar to make it fit around the subject perfectly
Trial list
Add the desired number of trials
Running the Experiment
Running the Experiment
Acquisition
Software will load the first trial
Put subjects in center of cylinders
Press record
  • The program will run for 6 minutes and automatically stop
You must watch subjects closely throughout the test. If the mouse dips below the water surface, remove this animal from the water, dry it, and place it in a recovery cage.
Critical
Remove subjects
Dry subject off with a clean towel and place in a clean cage over a heating pad set to high to avoid hypothermia
Remove all feces and bedding from water using a fish net
Repeat for all trials
Results and Analysis
Results and Analysis
Data Profile
Ensure that it is pulling data from all trials
Analysis Profile
Distance traveled
  • Arena
  • Statistics - total only
Velocity
  • Arena
  • Averaging interval - 1
  • Statistics - mean
Movement
  • Moving - cumulative duration
  • Not moving - cumulative duration
Mobility State
  • Highly mobile - cumulative duration
  • Mobile - cumulative duration
  • Immobile - cumulative duration
Results
Statistics and charts
Calculate to generate a file with the analysis parameters chosen
Export and save
Acknowledgements
We would like to thank the UAB Neuroscience Behavioral Assessment Core for use of their facilities.