Knowledge Base

Experiment Settings

Choose Setup > Experiment Settings.

Under Video Source and Tracked Features, make sure that the options selected correspond to your needs. If you want to analyze exploration in detail, choose Center-point, Nose-point and Tail-base detection.

If you want to use the Deep Learning technique to track the subject's nose, under Body Point Detection Technique choose Deep Learning. Note that in order to use this technique there are additional requirements. Among other things, you need a powerful graphics card. See Deep Learning: Requirements and Limitations in the EthoVision XT Help.

Manual Scoring Settings

If your EthoVision XT license does not include the Behavior Recognition module, EthoVision XT cannot detect sniffing or rearing automatically. However, you can still score those and other behaviors with keystrokes.

Choose Setup > Manual Scoring Settings.

If you used the template for an open field experiment, four behaviors have been defined:

• Sniffing R and Sniffing O, to record manually sniffing at the receiver (unfamiliar mouse) and object (either familiar mouse or empty cage, depending on the test), respectively.
• Rearing R and Rearing O, to record manually rearing at the receiver (unfamiliar mouse) and the object cage (either familiar mouse or empty cage, depending on the test), respectively.

You can also rename the behaviors if you like.

Arena Settings

Choose Setup > Arena Settings.

If you work with a Sociability test cage with backlight unit, you can reduce the intensity of the backlight before adjusting the arena settings. This makes the edges of the floor plate and walls more visible.

If the separation walls show reflections of the animal, this may hinder tracking. In the Arena Settings, click the Arena layer and draw rectangles around the separation walls.

Detection Settings

Choose Setup > Detection Settings.

Check in the Video Section that the sample rate is set to:

• Center-point tracking: 5–8 samples/second (for rats) and 12.5–15 samples/second (for mice).
• Nose-tail tracking (rats and mice): 25–30 samples/second.

Notes

If detection of the subject is not optimal after using the Automated Setup function, see Configure Detection Settings in the EthoVision XT Help for details on the advanced detection settings. If you use hooded animals, use the detection method Differencing.

If the animal moves from one compartment to the other, it may not be detected well. To improve detection, open the Advanced section in the Detection Settings pane. Under Subject Contour, select one or more pixels for the Dilation filter and the second Erosion filter. Leave the first Erosion filter set to zero. Select more pixels for Dilation than for Erosion (for example, 3 vs. 1) until the entire animal's contour is detected.

Trial List

Choose Setup > Trial List. Click Add Variables and enter your independent variables such as the experimental phases and the ID code of your test subject.

Acquiring Data

Protocol

Sociability Test

For each test animal, three trials are carried out:

1. The first trial (for example, 5 min) aims at habituating the test individual to the chamber. Tracking is required when one wishes to know whether the animal explores one chamber more than the other.
2. A second trial (5 min) is carried out with the empty tubes in the test chambers. The aim of this trial is to quantify the time spent in the two side chambers and near the tubes.
3. In the third trial (next 5 min), an empty tube is removed and replaced by a female animal restrained in an identical tube. This session is designed to observe differences between the time spent sniffing the social versus non-social object.

Social Discrimination Test

For each test animal, three trials are carried out:

1. The first trial (10 min) is aimed at habituating the test individual to the chamber and empty tubes.
2. In the sociability test (next 10 min), an empty tube is removed and replaced by a female animal restrained in an identical tube. This session is designed to observe differences between the time spent sniffing the social versus non-social object.
3. In the social discrimination test (10 min), the other empty tube is removed and replaced by a novel female. This session is designed to test the ability of the test animal to distinguish between the two females.

Score Behaviors Manually

To score sniffing and other behaviors defined under Manual Scoring Settings, open the Manual Scoring tab on the Acquisition screen. There you will find the key codes for the behaviors.

To score an instance of the behavior or its end, either press the key or click the corresponding button on the screen.

Notes

• Important: If you are doing tracking from video, make sure you deselect DDS in the Playback Control window.
• You can also score behaviors after tracking. See Acquire Data > Score Behaviors Manually in the EthoVision XT Help.

Source: EthoVision XT 18 - Application Manual, Noldus Information Technology