# Other analyses with zones

What do you want to do?

▪Calculate path length and speed for specific zones

▪Calculate path length and speed for a combination of zones

▪Calculate the distance traveled before reaching the platform in a Morris water maze

▪Calculate the distance traveled or time before entering a zone

▪Calculate the distance traveled or time before a specific zone visit

▪Find where a behavior occurred

▪Find out when the subject disappears from the arena

Calculate path length and speed for specific zones

In this case you calculate dependent variables for track segments. First, you specify the track segments When in zone with a Data profile and then you choose the dependent variables in an Analysis profile.

1.In the Data profile, in the Result box, click the Settings button and then select Results per zone. Specify the zones you are interested in.

2.In the Analysis profile, click the button next to the dependent variable you require, e.g Distance moved or Velocity. In the Trial Statistics tab, select the statistic (e.g., Total for distance moved and Mean for velocity).

3.Click Statistics and Charts, then Calculate.

Calculate path length and speed for a combination of zones

1.In the Data profile, in the Components pane, under Nesting click the button next to In zone. Specify the zones you are interested in. Select the option that you require:

▪When in any of selected zones to analyze when the animal was in one or another zone.

▪When in all selected zones to analyze when the animal was in all the zones simultaneously. This assumes that the zones are at least partially overlapping: for instance, a quadrant and a border zone in a water maze. If zones are not overlapping, data are not selected since the subject’s body points cannot be in two non-overlapping zones at the same time.

For more information, see Nesting over In zone.

2.In the Analysis profile, click the button next to the dependent variable (e.g. Velocity).

3.Click Statistics and Charts, then Calculate.

Calculate the distance traveled before reaching the platform in a Morris water maze

Use the function Free Interval in the Data profile to select the interval from the track start to when the animal stays on the platform for say five seconds.

1.In the Data profile, in the Components pane, under Nesting click the button next to Free interval. Specify the interval:

▪Start criterion: Time (Track start).

▪Stop criterion: Dependent Variable In zone. Statistic: Current duration >=5 s”.

▪Select Ignore last interval if incomplete. This will consider only cases when the animal reaches the platform.

2.In the Analysis profile, click the button next to the Distance moved. In the Trial Statistics tab, select Total.

3.Click Statistics and Charts, then Calculate.

EthoVision XT only gives results for those trials where the animal stayed on the platform for five seconds (here below, Trial 1). In all other cases, including when the animal swims over the platform, but does not stop there, EthoVision XT shows “-”; see Trial 2.

Calculate the distance traveled or time before entering a zone

For example, in a Barnes maze, you want to calculate the Total distance moved before the animal enters a zone.

For distance traveled

1.In the Data profile, in the Components pane, under Nesting click the button next to Latency to zone. Specify 1st visit, the zone and keep Interval before zone visit selected.

2.In the Analysis profile, click the button next to the Distance moved. In the Trial Statistics tab, select Total.

3.Click Statistics and Charts, then Calculate.

For time

If you want to know the time before the animal enters a zone, you do not need a Data profile.

1.In the Analysis profile, click the button next to the In zone. Choose the zone and in the Trial Statistics tab, select Latency to first.

note Latency is always calculated from the start of the track, even when you define time bins and nesting intervals.

2.Click Statistics and Charts, then Calculate.

Calculate the distance traveled or time before a specific zone visit

In this example, we want to know the distance traveled or the time taken to stay in a zone for more than a specific time.

For distance traveled:

1.In the Data profile, in the Components pane, under Nesting click the button next to Free interval. Specify the interval “From Time (track start) to [dependent variable]. For example,

“From track start to Current duration >=20 s”. This will pick out the interval until the animal stays in a zone for at least 20 s.

2.In the Analysis profile, click the button next to the Distance moved. In the Trial Statistics tab, select Total.

3.Click Statistics and Charts, then Calculate.

For time:

1.In the Analysis profile, under Custom Variables click the button next to the Free Interval. Define the interval. In the Trial Statistics tab, select Cumulative duration.

2.Click Statistics and Charts, then Calculate.

Find where a behavior occurred

The term “Behavior” here indicates a state like Movement, Mobility, Activity, Body elongation, behaviors scored manually, or behaviors detected automatically.

Quantitative analysis

If you are interested in quantitative results (for example, the number of occurrences per zone). Specify the zones in a Data profile and select the behavior in an Analysis profile.

1.In the Data profile, in the Result box, click the Settings button and then select Results per zone. Select the zones you are interested in.

2.In the Analysis profile, select the behavior you are interested in. In the Trial Statistics tab, select Frequency.

3.Click Statistics and Charts, then Calculate.

Qualitative analysis

If you are interested in a visualization. Select the behavior and optionally the zones of interest in a Data profile and then create a heatmap of the frequency of visits to the zones.

1.In the Data profile, in the Components pane, under Nesting, click the button next to the behavior you want to visualize.

optional If you are interested in particular zones, in the Result box, click the Settings button and then select Results per zone. Specify the zones you are interested in.

2.Click Heatmap Visualization, then Plot Heatmaps.

Find out when the subject disappears from view

There are situations when you want to know when the subject is no longer found in the arena. In the wire hanging test, used to evaluate abnormalities in muscle strength, the time that the subject remains hanging on the wire needs to be recorded. A way to do this is to measure the time until the subject falls from the wire and disappears from view.

1.Make sure that the camera view does not include the bottom of the experimental cage. So when the animal falls, the software does not see it anymore.

2.In the Arena Settings, define a hidden zone at the bottom of the arena. Define an entry zone that should occupy the lower half of the arena. Make sure that the entry zone partly overlaps with the hidden zone. See Shelters and other hidden zones

3.Acquire the tracks. When the subject falls, its center point is first found in the entry zone, then it disappears, so it is automatically assigned to the hidden zone.

4.In the Analysis profile, click the button next to In Zone. Select the hidden zone and on the Trial statistics tab choose Latency to first.

5.Click Statistics and Charts, then Calculate.