Open Field Test in Rodent Models of Stroke

At a glance

The open field test is a common measure of exploratory behavior and general activity in both mice and rats, where both the quality and quantity of the activity can be measured. The open field test has been widely used to measure locomotion and some aspects of anxiety-related behavior in rodent models of ischemic and hemorrhagic stroke.

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Introduction to The Open Field Test

While the original studies were in rats, the open field test has also been extensively used in mice. Basically, a wooden or plastic open field maze with an enclosure, generally square, rectangular, or circular with surrounding walls is used in this test. Mice are placed in the open field and allowed to explore for 2-10 minutes. Exploratory behavior is recorded and analyzed by video, focusing on the distance traveled by the mice to measure locomotor activity. More recent approaches, including video tracking and tracking by the number of infrared beam breaks, allow for much higher throughput and longer periods of monitoring to visualize and analyze the routes of mice. In addition, resting time and standing behavior can be collected and analyzed to provide a comprehensive assessment of locomotor ability.

In addition, the open field test is widely used to assess emotional behavior. Defecation and urination can indicate mood, but these metrics are affected by factors such as food and water intake, making them difficult to standardize. In contrast, the amount of time an animal spends in specific areas of the open field has been used to analyze anxiety levels. Rodents generally prefer corners of the open field; therefore, the shorter the time an animal spends in the center of the open field, the higher the level of anxiety.

Fig.1. Methods for monitoring open field behavior. Fig.1. Approaches to track open field behavior. (Ruan, et al., 2020)

Advantages of Open Field Test

The open field test is a straightforward procedure that doesn't require advanced equipment. For stroke models that impair locomotion, a dark environment may encourage exploration since light can reduce rodent activity. However, recording in low-light conditions necessitates higher-quality video equipment, complicating the testing process.

Open-field-dependent Parameters

Movement

Distance moved (either actual or relative distance)
Time spent moving
Rearing (vertical activity)
Freezing
Grooming
Other stereotypic behaviors

Location

Time spent in the center
Crosses into center

Autonomic nervous system

Defecation (number of fecal boli)
Urination

Common Problems When Conducting the Open Field Test

Problems	Solutions
Low (or High) Amount of Time in Center	Stress confirmation: Ensure that mice have not experienced stress prior to testing. Lighting adjustments: Reduce light levels, as mice tend to spend more time in the center in lower light conditions. Testing under red light is also an option, as certain wavelengths are not visible to rodents. Center zone size: Adjust the size of the center zone, which typically comprises 25–50% of the total area of the open field. Field Size Variation: Use different sizes of open fields, as larger areas are more sensitive to thigmotaxis (the tendency to stay near the walls).
Large Standard Error Bars	Uniform testing history: Confirm that all experimental animals have the same testing history and are of similar ages, as age differences can affect results. Sex consideration: Be aware of potential sex effects on activity levels. Adequate sample size: Ensure that the study is sufficiently powered; while a group size of 8–10 mice may sometimes be adequate, larger groups (often double that size) are frequently needed for behavioral experiments.
Lack of Exploratory Activity	Strain effects: Different mouse strains exhibit distinct behaviors, both unmedicated and medicated. For instance, BALB/c and A/J mice have low baseline activity and do not exhibit hyperlocomotion when treated with D-amphetamine. Timing of testing: Activity levels can vary depending on the time of day, with increased activity typically observed during the dark phase of the light-dark cycle, which is when rodents are most active. Lighting conditions: Ensure consistent lighting across all areas of the open field. Lower light levels and the use of red light can generally enhance activity levels. Ambient environment: Check for environmental stressors, such as fluctuations in humidity or temperature, and ensure that humans are not within the mice line of sight, as these factors can impact behavior.

Reference

Gould, T. D., et al. (2009). The open field test. Mood and anxiety related phenotypes in mice: Characterization using behavioral tests, 1-20.