A common question asked by behavioral researchers is how to clean the testing apparatus in between subjects and at the conclusion of an experiment. Maybe you have wondered what the best cleaning agent is, or whether your current regimen is up to par. Perhaps you once cleaned an area of the maze with that antibacterial spray that was conveniently sitting on the shelf in the vivarium, but then you noticed that your mice began avoiding that part of the maze. Maybe you observed during a test that one of your mice followed the exact path traveled by the previous mouse tested, and you wondered whether pheromonal cues might be affecting your results.
Why is Cleaning a Behavioral Testing Apparatus Important?
Unlike people, rodents have an incredibly sensitive olfactory system. Smells that are undetectable to us can have profound effects on rodent behavior[1]. The proper cleaning of behavioral equipment is a critical step to minimize the impact of these residual odorant molecules on test results[2].
Rodents leave behind a variety of odorant cues in their environment. During behavioral testing, the excretion of pheromone-containing urine and feces is a common event. These substances are released due to the relatively stressful testing environment, which includes stressors such as bright light, a novel environment, and human handling. Rodents can also secrete pheromones from their whisker region and in fluids from their foot pads[3, 4]. Odorant molecules in this fluid can be detected by other rodents, who can use the scent to track the animal’s spatial trajectory[5].
In behavioral testing, these cues can cause an animal to favor the path of a previously tested subject. Conversely, the release of alarm pheromones can act as a deterrent[6]. For these reasons, it is incredibly important to thoroughly clean an apparatus after every test subject.
Unlike people, rodents have an incredibly sensitive olfactory system. Smells that are undetectable to us can have profound effects on rodent behavior[1]. The proper cleaning of behavioral equipment is a critical step to minimize the impact of these residual odorant molecules on test results[2].
Rodents leave behind a variety of odorant cues in their environment. During behavioral testing, the excretion of pheromone-containing urine and feces is a common event. These substances are released due to the relatively stressful testing environment, which includes stressors such as bright light, a novel environment, and human handling. Rodents can also secrete pheromones from their whisker region and in fluids from their foot pads[3, 4]. Odorant molecules in this fluid can be detected by other rodents, who can use the scent to track the animal’s spatial trajectory[5].
In behavioral testing, these cues can cause an animal to favor the path of a previously tested subject. Conversely, the release of alarm pheromones can act as a deterrent[6]. For these reasons, it is incredibly important to thoroughly clean an apparatus after every test subject.
What is the Best Method for Cleaning a Behavioral Testing Apparatus?
Laboratories use a variety of agents to clean testing apparatuses, but some have advantages over others. Water is commonly used because it has no odor itself, but this may not be the most effective option for removing odors and microbes.
A number of published protocols recommend the use of disinfectant agents such as Quatricide[7]; however, their intrinsic odor can act as a deterrent to rodents. These agents, which are commonly found in vivariums, may be better reserved for cleaning behavioral testing equipment at the conclusion of testing sessions.
Seventy percent (70%) ethanol is another common disinfectant used to clean testing equipment[8], but care must be taken to ensure that it has evaporated completely before placing an animal in the apparatus. Like many disinfectants, alcohol itself has an odor that can influence rodent behavior.
Another validated option for cleaning the testing apparatus is super oxidized or super hypochlorous water, which has been demonstrated to be an effective odor removal agent with a weak intrinsic odor[8, 9].
The ideal testing apparatus will be composed of a material that is easy to clean and devoid of narrow spaces that are difficult to reach. All areas of the apparatus should be wiped clean with the agent of choice after each animal is tested. It is also critical to thoroughly dry the apparatus before placing an animal inside, as moisture can also influence rodent behavior.
The next time you run a behavioral test, make sure you have the appropriate disinfectant and clean paper towels along with your other testing supplies. Starting each mouse with a clean slate is the best way to begin a behavioral test, and taking care to perform this important step will be reflected in your results.
Note: Caution should be taken with harsh cleaners such as hydrogen peroxide which can damage acrylic and other plastics
References
- Bind RH, Minney SM, Rosenfeld S, Hallock, RM. (2013) The Role of Pheromonal Responses in Rodent Behavior: Future Directions for the Development of Laboratory Protocols. J Am Assoc Lab Anim Sci. 52(2):124–129.
- Hanell A and Marklund N. (2014) Structured evaluation of rodent behavioral tests used in drug discovery research. Front Behav Neurosci. 8:252.
- Kiyokawa Y, Kikusui T, Takeuchi Y, Mori Y. (2004) Alarm pheromones with different functions are released from different regions of the body surface of male rats. Chem Senses. 29(1):35-40.
- Quatrale RP and Laden K. (1968) Solute and water secretion by the eccrine sweat glands of the rat. J Invest Dermatol. 51(6):502-4.
- Wallace DG, Gorny B, Whishaw IQ. (2002) Rats can track odors, other rats, and themselves: implications for the study of spatial behavior. Behav Brain Res. 131(1-2):185-92.
- Shorey, HH. (1976) Animal Communication by Pheromones. Academic Press.
- Walf AA and Frye CA. (2007) The use of the elevated plus maze as an assay of anxiety-related behavior in rodents. Nat Protoc 2(2):322-328.
- Leo LM and Pamplona FA. (2014). Elevated Plus Maze Test to Assess Anxiety-like Behavior in the Mouse. Bio-protocol 4(16):e1211
- Komada M, Takao K, Miyakawa T. (2008) Elevated Plus Maze for Mice. J Vis Exp. (22):1088.
- Gunaydin M, Esen S, Karadag A, Unal N, Yanik K, Odabasi H, Birinci A. (2014) In vitro antimicrobial activity of Medilox® super-oxidized water. Ann Clin Microbiol Antimicrob. 13:29.