Knowledge Base

Introduction to Trial Control

This Manual

This manual is meant for all users of EthoVision XT:

• For users of the Base version. You can find extensive information on the Trial Control conditions and analysis of Trial Control data. This information is not present in the EthoVision XT Help.
• For users of the Base version plus the Trial and Hardware Control module. You find all the information you need in this guide.
• If you use EthoVision XT in combination with hardware devices, you can find the relevant information in Section 7.

EthoVision XT Help

For extensive information on EthoVision XT or for troubleshooting, see the EthoVision XT Help. In EthoVision XT, choose Help > EthoVision XT Help or press F1.

Other Documentation

From the Windows Start menu choose All Apps > Noldus > EthoVision XT 18 Other documentation. There you find:

• EthoVision XT 18 - Application Manual for information on common tests including conditioning tests that make use of sub-rules in the Trial Control Settings.
• PhenoTyper - EthoVision XT 18 - Reference Manual for information on PhenoTyper and how to connect it to EthoVision XT.
• PhenoTyper - EthoVision XT 18 - Service Manual for information about PhenoTyper and devices such as the Noldus Pellet dispenser, the Noldus Lickometer and PhenoWheel.
• DanioVision DVOC-0041 - Reference Manual if you use Trial Control with the DanioVision Observation Chamber.

Why Use Trial and Hardware Control?

Trial Control allows you to automate your experiment. For example:

• You want to set a maximum duration for your trials.
• You want to automate the start and/or stop of data acquisition. For example:
  • Start recording when the rat is first detected in the open field.
  • Stop recording when the rat has reached the platform in the Morris water maze.
  • Start recording at exactly 12:30:00.
  • Stop recording after the animal has been in the closed arms of the plus maze for 5 minutes.
• You want to automate conditioning schedules, or operate hardware devices. For example:
  • When the animal presses a lever, have the food dispenser drop a food pellet.
  • When the animal leaves the shelter, use an External Command action to start a recording with Media Recorder.
  • When the animal enters the shelter, turn on a light.
  • Close the door of a radial maze when the animal has exited that arm.

To Use Trial and Hardware Control

1. Open the Trial Control screen.
2. Define the conditions that, when met during your trial, trigger specific actions. Organize conditions and actions in sequences.
3. Before starting data acquisition, make sure that those Trial Control Settings are active.

For information on how to manage Trial Control Settings, see also Settings and profiles, under File management in the EthoVision XT Help.

Conditions and Actions

A Condition is a statement that EthoVision evaluates. An Action is a command executed on a variable or a hardware device. You can therefore control your experiment by linking conditions with actions.

• Example 1: In a Morris water maze test, stop tracking when the rat reaches the platform (provided that the platform has been defined as a zone). Stop tracking is the action and Rat on the platform is the condition.
• Example 2: In a Fixed Ratio schedule, have EthoVision send a command to drop a pellet when the mouse has been detected in the Trigger zone. Drop one pellet is the action and Mouse in the Trigger zone is the condition.

You define and link conditions with actions in a graphical form. A condition is followed by an action. The condition checks that the animal is in the zone named "Trigger zone". The action "Drop a pellet" is taken when the condition is met.

You can copy, move, connect and delete Trial Control elements such as actions and conditions. For how to do so, see the EthoVision XT Help.

Trial Control Rules

A Trial Control rule is a set of conditions and actions organized in a logic sequence. It can be viewed as a set of instructions executed during a trial. There are two types of Trial Control rules:

• Start-Stop trial rule. For starting and stopping data acquisition. This rule cannot be repeated or deleted.
• Sub-rules (available only in EthoVision XT with the Trial and Hardware Control Module). Subroutines of the Start-Stop trial rule, which are meant to carry out specific actions. Sub-rules can start at specific times and be repeated according to conditions specified in a Sub-rule reference.

How Trial Control Instructions Are Executed

The instructions contained in the Trial Control Settings are carried out from the moment you start a trial, to the moment the trial is stopped. Only the instructions in the Trial Control Settings currently active (that is, highlighted in blue in the Experiment Explorer) are carried out.

The program evaluates the Trial Control sequence at each sample time. The rate at which this happens depends on your chosen sample rate, not on the video frame rate.

For the Start-Stop trial rule and each active sub-rule, the program remembers which Trial Control boxes were evaluated (active) in the previous sample. Depending on the type of this box:

• For a Condition box. EthoVision XT waits until the condition is met. When this happens (condition becomes true), the program passes control to the next box in the sequence. The condition becomes then inactive.
• For an Action box. EthoVision XT carries out the action, and passes control to the next box, which becomes active. Then, the Action box becomes inactive.
• For a Sub-rule Begin box. Control is passed to the box that follows. The Sub-rule Begin box becomes inactive.
• For a Sub-rule End box. This marks the end of a sub-rule. The sub-rule becomes inactive. Control is passed to the Reference box that had called that sub-rule. This Reference checks whether the sub-rule must be repeated. If this is not the case, control is passed to the box following the Reference box.
• For a Reference box. Control is passed to the Sub-rule Begin box. The Reference box makes the sub-rule active. The Reference box stays active until the last sub-rule repeat is completed. Next, the Reference box makes the sub-rule inactive and passes control to the box that follows. Next, the Reference box becomes inactive.

The events marked in italics above are those you can define in the Analysis profile and analyze or visualize in a chart. When a box becomes active, the previous one becomes inactive.

Schematic of Trial Control Instruction Execution

1. Tracking starts, either manually or because a previous condition has been met.
2. Control passes to a Condition box (for example "Is mouse on top of Shelter?"), which becomes active. The condition is evaluated.
3. The Condition is met (becomes true).
4. Control passes to the next box. In this case, it is an Action. Actions are taken immediately.
5. The Action box becomes inactive, and the next box becomes active.

For clarity, steps 3 and 4 are placed separately. In reality, the software is so fast that when a condition becomes true it also becomes inactive at the same sample time, and control passes to the next box.

When plotted against time, the time that the animal is in the zone is indicated by a horizontal bar. When tracking starts, the Action "Start track" becomes active and immediately inactive as control passes to the next box. The condition "On top of Shelter?" becomes active and immediately false since the animal is not in that zone. When the animal enters the zone, the condition becomes true and becomes immediately inactive as control passes to the next box. The action becomes active and immediately inactive as the command "drop a pellet" is sent to the pellet dispenser.

Evaluation of Trial Control Boxes

• Boxes combined in parallel using operators are evaluated at about the same time, in random order. This means that you cannot establish which condition is evaluated or which action is taken first.
• When your trial control is complex, for example when it includes sub-rules or conditions combined with operators, it is possible that two or more Trial Control boxes are evaluated at the same sample time.
• When a condition based on a dependent variable is evaluated (and therefore may or may not be met) also depends on the statistic used.
• If two or more boxes are evaluated at the same time, an action on a hardware device is taken only after all active boxes that must be evaluated have been evaluated.
• If the next box to be evaluated contains a condition that is fulfilled immediately, the program passes control to the next box. Therefore, within one sample time the program can pass control to two or more boxes to the right.
• When you stop the trial or the Maximum trial duration has been reached, all Trial Control boxes are deactivated.
• When the Rule End box of the Start/Stop trial rule is evaluated for an arena, data recording stops for that arena. From that moment, Trial Control is deactivated for that arena, even in those sub-rules that were ongoing in the meantime.
• A trial stops when trial control is deactivated in all arenas.
• Actions on hardware devices. A command is sent to a device after all boxes that must be evaluated have been evaluated. However, it may take some time for the outcome to be seen. For example, when a pellet dispenser is almost empty it may take a few seconds before a pellet is released.
• Sub-rules. A sub-rule is completed when the Rule End box has been evaluated, or when it is interrupted.
• If two or more sub-rules are evaluated at the same time, it is not possible to establish which sub-rule is evaluated first at a specific sample.

Important: When two or more actions on hardware devices follow each other, it is possible that they are evaluated at the same sample time. This happens because the computer is fast enough to analyze multiple Trial Control boxes in the sample interval. To make sure that actions are taken in the correct sequence and do not interfere with each other, add a Time condition between the action boxes, so after the first action is carried out EthoVision XT waits for a short time (for example, 0.1 s) before carrying out the next one.

Trial Control in Multiple Arenas

If your experimental setup includes two or more arenas, Trial Control is applied to each arena separately. This means that, if a condition is met in one arena, EthoVision XT takes the corresponding action in that arena, not the others.

Example: The setup includes four cages, each defined as an arena. A Trial Control In zone condition has been defined so that tracking starts when the animal is first detected in the arena. When you first put an animal in Arena 2, the condition is met in this arena and tracking starts for that arena. When you release the second animal in Arena 4, 2 seconds later, tracking in that arena starts 2 seconds later than in Arena 2.

The advantage of Trial Control in multiple arenas is that you can put one animal at a time into the arenas, and EthoVision XT will start tracking in each arena at the appropriate moment.

If your setup includes multiple arenas, you cannot define a condition or action specific to one arena. This means that the zones and hardware devices specified in the condition or action must be present in all arenas, and have the same name.

• If a zone or hardware device is not present in an arena, and a condition is based on that zone or device, Trial Control cannot progress for that arena. Therefore, tracking does not stop unless you set a maximum trial duration or tracking reaches the end of the video.
• Trial Control executes the instructions for each arena separately; however one cannot establish which arena is evaluated first at a specific sample time.

Your EthoVision License and Trial Control

Your EthoVision XT license determines which type of Trial Control you can set up.

• EthoVision XT Base license. You can define a rule to start and stop data recording (Start-Stop trial rule), not sub-rules. You can also use an External command action. You cannot control hardware devices.
• EthoVision XT Base + Trial and Hardware Control Module. You can define a Start/Stop trial rule, and in addition sub-rules that serve as subroutines of the Start-Stop trial rule. Moreover, you can control hardware devices.

Note: To acquire data in an experiment created with the Trial and Hardware Control Module, you must have a license that includes this module.

For more information on your EthoVision XT license, see the EthoVision XT Help.

Analysis of Trial Control Data

The Trial Control function also allows you to easily analyze events that occurred during the trial. For example, in a conditioning experiment:

• Number of rewards. Calculate the number of pellets dropped during a trial.
• Latency between action and reward. Calculate the time interval between the time the animal pressed the lever and the time the animal approached the pellet dispenser.
• Number of failures. Calculate the number of times the animal approached the lever during the period between dropping the pellet and the next opportunity.

Source: EthoVision XT 19 - Trial and Hardware Control Reference Manual