Technique
A thigmotaxis-based method of recapturing and transporting small mammals in the laboratory Rafał Stryjek, PhD & Klaudia Modlińska, MSc
© 2013 Nature America, Inc. All rights reserved.
One of the challenges associated with breeding and testing of wild animals in the laboratory is their drive to escape. Wild rats attempt to escape their cages as soon as they are opened, and recapturing is often difficult and dangerous to laboratory personnel. The authors describe a method of recapturing wild small mammals in a laboratory setting that takes advantage of thigmotaxis, or the natural tendency to move close to walls and other solid objects. They describe a simple device that is easy to construct and can be used to recapture escaped rats and return them to their home cages. The recapture technique is efficient and benefits animals’ well-being by minimizing their distress.
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Breeding and testing wild animals in a laboratory setting poses a number of technical challenges. Most difficulties are associated with manipulating undomesticated animals during care activities and experimental procedures. Wild rats demonstrate significantly higher levels of anxiety, aggression and defensive behavior than laboratory rats1–4. One of the difficulties encountered when breeding wild animals is their powerful drive to escape2,3,5. This complicates transportation and makes introducing or removing a rat to or from an experimental arena a real challenge. As soon as its cage is opened, a rat will attempt to escape and run away from laboratory personnel, which are perceived as a threat. If a rat manages to escape, recapturing it is very difficult and often can result in injury such as bites. This can pose risks of zoonosis to laboratory personnel, particularly when handling recently captured rats6,7. Navigating a novel, threatening environment may also be stressful for the rats. In our laboratory, we have bred wild rats of the Warsaw Wild Captive Pisula Stryjek (WWCPS)8 strain for many years. During this time, we have developed a number of practical techniques for handling these animals9,10. All rodents, rats included, demonstrate thigmotaxis, which is a tendency to move close to solid objects such as walls. Thigmotaxis may be adaptive5: it protects rats from predatory attacks (by birds
in particular). Although wild rats avoid novel objects and rarely enter traps in a familiar setting11, a rat looking for an escape route in a novel environment, such as the floor of a laboratory, will move along the walls and hide in small, unlit spaces. Taking advantage of this tendency, we devised a simple method of recapturing and transporting wild rats in a laboratory setting. We constructed a box of dark plexiglass that provides the rat with a place to hide. The boxes are placed adjacent to the wall of the room, leaving unobstructed passage along the walls, to provide access to the box’s opening as the rat travels along the wall. This procedure helps us to recapture the rat quickly and harmlessly so that we can return it to its cage. Our device is not only effective for recapturing escaped animals but, when appropriate in size, can also be used to transport rats between cages and experimental arenas and setups. When captured using this technique, rats likely experience less stress than when caught using traditional methods, because the technique takes advantage of the rats’ natural and spontaneous behavior. Stress levels also may be mitigated by the brief time needed to complete the entire procedure. TECHNIQUE All procedures described in this paper were approved by the 4th Local Ethics Committee for Animal
Institute of Psychology, Polish Academy of Sciences, Warsaw, Poland. Correspondence should be addressed to R.S. (
[email protected]).
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FIGURE 1 | The recapture device. A transparent and flexible cover can be inserted through a slot (1) in the back end of the box. A detachable flap can be mounted obliquely on the sides of the box (2) opposite the entrance. Two grooves (3) guide the cover to close over the opening in the front of the box.
Experimentation, Warsaw, Poland. All rats were cared for in accordance with the Regulation of the Polish Minister of Agriculture and Rural Development of 10 March 2006 on laboratory animal care12. Animals We used eight male and twelve female wild-type rats of the WWCPS strain, aged 3–12 months, in this study. We derived the WWCPS strain in 2006 from genetic material obtained from five independent colonies of wild rats8. In order to prevent domestication of the rat population, we systematically add to the colony new rats that have been captured from various locations in the area of Warsaw, Poland. The rats included in this study were from generations F2, F3 and F4. The rats were housed in groups of three to five rats per cage in Eurostandard type IV cages (Tecniplast, Buguggiate, Italy) and had ad libitum access to water and standard laboratory feed (Labofeed H, WP Morawski, Kcynia, Poland). The room in which they were housed was maintained on a 12-h:12-h light:dark cycle. Equipment We constructed a box (Fig. 1) of 5-mm-thick dark plexiglass (polymethyl methacrylate, HEKO, Oświęcim, Poland). We designed the device to be used in two ways. First, we equipped the box with a transparent, flexible, insertable cover made of 1.5-mm-thick copolyester (Eastman Tritan, Kingsport, TN) that can be manually closed after a rat has entered the box (Fig. 2a,b). Second, we mounted a 1.5-mm-thick copolyester flap obliquely opposite the entrance of the box (Fig. 2c) so that a rat can enter the box by pushing up the flap (Fig. 2d); the flap then closes behind it, preventing it from exiting the box (Fig. 2e). The transparency of the cover allows viewing of the captured 322 Volume 42, No. 9 | SEPTEMBER 2013
animal while it is inside the box. The materials that we used to construct the device can be sterilized in an autoclave. Experimental arena To test our capture technique, we replicated the typical conditions associated with attempted recapture of an escaped animal in an experimental arena. The arena floor had an area of 200 cm × 200 cm and was covered with ceramic tiles. The arena was surrounded by 1-m-high walls that were covered with galvanized metal sheeting (Fig. 3). A centrally located lamp provided consistent lighting throughout the arena of ~160 lux. At one wall of the arena, we placed two of the boxes back-to-back, with a distance of 15 cm between them owing to the two insertable covers sticking out in open positions. Procedure We removed each rat individually from its breeding cage using a special transporting device10 and introduced it onto the floor of the experimental arena facing the wall (Fig. 3). The animal was free to move around the experimental area, with the walls preventing it from escaping. To simulate normal conditions in the laboratory during breeding manipulations,
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FIGURE 2 | Two methods of recapturing a rat using the device. First, after the rat enters the box (a), the entrance can be manually closed by pushing the copolyester cover forward along grooves (a) so that it covers the entrance (b), trapping the rat in the box. Second, an oblique copolyester flap can be mounted inside the box at a 35° angle (c), making a one-way entrance. A rat enters the box by lifting the flap (d), which then closes behind it, preventing it from exiting the box (e). www.labanimal.com
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the experimental arena was cleaned carefully with Virkon disinfectant (DuPont Chemicals, Wilmington, DE) in order to remove any odors from previous rats.
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FIGURE 3 | The experimental arena. Two boxes were placed back-to-back against one wall of the arena. Arrows indicate the openings of the boxes. Location of the rat indicates where rats were introduced to the arena.
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laboratory assistants remained close to the experimental arena while talking and making moderate noise (~45–50 dB). Each trial ended when the rat completely entered one of the boxes placed in the arena by pushing up the copolyester flap on the front of the box (Fig. 4a). For each rat, we recorded the amount of time that passed from when the rat was introduced into the experimental arena to when it entered one of the boxes. Then the box with the captured animal inside was removed from the experimental arena. Finally, the rat was placed in a clean breeding cage by putting the box vertically into the cage and slightly pulling back the copolyester cover (Fig. 4b). After each trial,
OUTCOME Once in the experimental area, rats on average spent 85 ± 10% (72–100%) of time in close proximity to the walls. There were no significant sex differences in the proportion of time spent near the walls (Student’s t-test; P > 0.05). All rats in the experiment were captured within 74 s (min, 4 s; mean ± s.d., 22 ± 20 s) of being introduced into the experimental arena. There were no significant sex differences in latency period (student’s t-test; P > 0.05). CONCLUSIONS Using the technique described, we quickly and easily recaptured escaped rats by taking advantage of the rats’ natural thigmotaxis behavior. Our method is highly effective, safer for the laboratory staff and beneficial for the animals’ well-being. Our recapture device is superior to live traps currently available on the market (e.g., Sherman traps) for use in laboratory conditions for several reasons. Live traps are often made of galvanized steel, which is difficult to sterilize. Alternatively, they may be made of wire mesh, providing a well-lit interior during daytime that reduces the appeal to animals looking for a hiding space. They are often equipped with a trap door that slams shut, causing distress to the animals. Moreover, commercially available traps are often too big to be placed in barely accessible laboratory spaces or in an
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FIGURE 4 | Recapture procedure. (a) A wild rat enters the device. (b) The recaptured wild rat is reintroduced to its home cage. LAB ANIMAL
FIGURE 5 | Using the device to capture a rat from its home cage. (a) A wild rat enters the device that has been placed inside its home cage. (b) A female wild rat inside the device. A wire-covered floor allows the rat to climb up into the device from its home cage. Volume 42, No. 9 | SEPTEMBER 2013 323
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experimental arena. Safe removal of an aggressive or wild animal from a trap is difficult and can lead to re-escape. Some commercially available live traps have a triggering mechanism on the outside of the trap, enabling both humans and animals to shut the trap accidentally. In our laboratory, the technique also proved to be extremely effective in the recapture of several other animal species, such as wild house mice (Mus musculus), striped field mice (Apodemus agrarius) and common shrew (Sorex araneus). Therefore, we are confident that a similar method can be used with other small wild or laboratory species that demonstrate thigmotaxis behavior (e.g., laboratory mice13 and opossums14). Although laboratory animals generally have a much weaker tendency to escape and lower stress levels, our technique may be useful when dealing with distressed animals, such as those subjected to medical examination or experimental procedures involving administration of drugs or surgical interventions. It should be noted that this method cannot be used with excessive regularity. Our observations have shown that some rats captured multiple times using this method learn to avoid the box, which makes them even more difficult to catch. In addition to recapturing escaped small laboratory animals, our device also may be used as a capturing instrument (Fig. 5a) for transporting highly aroused or very aggressive rats. In this case, the box can be equipped with detachable wire floor allowing rats to climb up easily (Fig. 5b). A comb-like device8,10 can be used to reduce the cage size and push the rat toward the transporter to transport animals and deposit them in breeding cages. The device can also serve as a relatively safe place from which animals may start examining a new experi mental area. According to the free-exploratory paradigm15, animals can regulate their own approach to novelty only when they are given a choice of whether or not to explore a given environment. Such an approach to transporting the rats encourages this by bringing the rats into an experimental chamber in a relatively familiarized transportation device instead of directly placing them in the novel environment. The recapture device could be also used as a chamber for preliminary anesthesia: cotton wool
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moistened with an inhalation anesthetic could be inserted into the device through the opening by slightly elevating the cover. It should be noted that the device is not hermetic, however, so it is advisable to keep the box under an extractor hood when used for anesthetic purposes. COMPETING FINANCIAL INTERESTS The authors declare no competing financial interests. Received 22 October 2012; accepted 17 May 2013 Published online at http://www.labanimal.com/ 1. Barnett, S.A., Dickson, R.G. & Hocking, W.E. Genotype and environment in the social interactions of wild and domestic ″Norway″ rats. Aggr. Behav. 5, 105–119 (1979). 2. Blanchard, D.C. et al. Defensive reactions of “wild-type” and “domesticated” wild rats to approach and contact by a threat stimulus. Aggr. Behav. 20, 387–397 (1994). 3. Blanchard, R.J., Flannelly, K.J. & Blanchard, D.C. Defensive behavior of laboratory and wild Rattus norvegicus. J. Comp. Psychol. 100, 101–107 (1986). 4. Lockard, R.B. The albino rat: a defensible choice or a bad habit? Am. Psychol. 23, 734–742 (1968). 5. Barnett, S.A. A Study in Behaviour (Methuen, London, 1963). 6. Heyman, P. et al. Seoul hantavirus in Europe: first demonstration of the virus genome in wild Rattus norvegicus captured in France. Eur. J. Clin. Microbiol. Infect. Dis. 23, 711–717 (2004). 7. Webster, J.P. & Macdonald, D.W. Parasites of wild brown rats (Rattus norvegicus) on UK farms. Parasitology 111, 247–255 (1995). 8. Stryjek, R. & Pisula, W. Warsaw Wild Captive Pisula Stryjek rat—establishing a breeding colony of Norway rat in captivity. Pol. Psychol. Bull. 39, 67–70 (2008). 9. Stryjek, R. Devices for handling small mammals in laboratory conditions. Acta Neurobiol. Exp. (Wars.) 68, 407–413 (2008). 10. Stryjek, R. A transportation device for rats. Lab Anim. (NY ) 39, 279–281 (2010). 11. Stryjek, R., Modlin´ska, K. & Pisula, W. Species specific behavioural patterns (digging and swimming) and reaction to novel objects in wild type, Wistar, Sprague-Dawley and Brown Norway rats. PLoS ONE 7, e40642 (2012). 12. Official Journal of Laws 2006, No. 50, item 368, as amended. 13. Kvist, S.B. & Selander, R.K. Maze-running and thigmotaxis in mice: applicability of models across the sexes. Scand. J. Psychol. 33, 378–384 (1992). 14. Wesierska, M. & Turlejski, K. Spontaneous behavior of the gray short-tailed opossum (Monodelphis domestica) in the elevated plus-maze: comparison with Long-Evans rats. Acta Neurobiol. Exp. (Wars.) 60, 479–487 (2000). 15. Griebel, G., Belzung, C., Misslin, R. & Vogel, E. The freeexploratory paradigm: an effective method for measuring neophobic behaviour in mice and testing potential neophobia-reducing drugs. Behav. Pharmacol. 4, 637–644 (1993).
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