Differential effects of amphetamine on the attack and defense components of play fighting in rats

Physiology& Behavior,Vol. 56, No. 2, pp. 325-330, 1994 Copyright © 1994ElsevierScienceLtd Printedin the USA. All rights reserved 0031-9384/94$6.00 + .00

Pergamon 0031-9384(94)E0086-J

Differential Effects of Amphetamine on the Attack and Defense Components of Play Fighting in Rats EVELYN

F. F I E L D A N D S E R G I O M. P E L L I S l

Department of Psychology, University of Lethbridge, Lethbridge, Alberta TI K 3M4, Canada R e c e i v e d 24 J u n e 1993 FIELD, E. F. AND S. M. PELLIS. Differentialeffects of amphetamine on the attack and defense components of playfighting in rats. PHYSIOL BEHAV 56(2) 325-330, 1994.--Treatment with d-amphetamine has been shown to cause a decrease in play fighting by juvenile rats. Previous studies, however, did not determine if all behavioral components of play were equally diminished. In this study, the effects of amphetamine on both the attack and defense patterns of play fighting were analyzed. Experiment 1 shows that a 0.5 mg/ kg dose, injected subcutaneously in the nape, decreases both attack and defense. In contrast, Experiment 2 shows that the same dose, injected subcutaneously in the hip, decreases the level of attack to a similar level, but does not significantly affect defense. This suggests that while the 0.5 mg/kg dose of amphetamine is primarily affecting the anackcomponents of play, via its action on the central nervous systems, the reduced likelihood of defense for those rats injected in the nape probably results from a local anesthetic effect, which numbs the area of the body defended during play fighting. Further doses (0.15 & 1.0 mg/kg), injected in the hip, were also tested. The highest dose decreased both components of play. The lowest dose had no effect on either attack or defense. It is suggested that the attack and defensive components of play fighting may be mediated by different neural systems. Amphetamine

Play fighting

Attack

Defense

d-Amphetamine

MANY species of mammals, as juveniles, engage in a form of social interaction that has been called play fighting or rough and tumble play (1,5). The species of choice for laboratory studies of such play has usually been the rat (12,28), which engages in both play fighting (7,9) and locomotor play (15,22). Play fighting involves a combination of attack and defense. Pellis and Pellis (16) have shown that during play fighting, juvenile rats direct their contacts to the partner's nape. If successful, this playful contact usually involves one juvenile rubbing its snout into its partner's nape area. If the recipient of this contact does not respond, the attacking animal briefly rubs its snout into the nape of its partner before jumping away (13). More commonly, the recipient adopts one of several defensive maneuvers to avoid, or extricate itself from such contact, thus producing a play fight. Previous studies have shown that the psychomotor stimulant, d-amphetamine, decreases play behavior in juvenile rats in a dose-dependent manner (2,3,26,27). These studies looked at the effects of amphetamine by recording the time spent in play or scoring the number of pins exhibited during a specified time period. Pins involve one of the pairmates attacking the other, usually at the nape, and the recipient rotating, over onto its back, and ending with the attacking rat standing over its supine partner (10). A pin then, is a composite measure that needs both an attack and defense component to be achieved. Changes in the frequency of pins or in the amount of time spent in play may arise from an

Juvenile rat

increase or decrease in the frequency of attack, defense, or some combination of both (18). Qualitative changes in the types of defensive maneuvers used and in the target areas attacked may also produce quantitative changes in the frequency or duration of play episodes. Therefore, previous studies have not documented which components of play have been changed by amphetamine. A second difficulty in understanding the effects of amphetamine on play behavior is that in most studies both pairmates are injected. This makes it impossible to evaluate the effects of the drug on defense. If the attack component is altered or decreased in one animal, the defensive responses of the other animal will also be altered. This suggests that a normal pairmate is needed to analyze the effects of a pharmacological manipulation on the defensive components of play exhibited by the injected animal (21). Two previous studies have used the one pairmate treatment paradigm, but they did not evaluate the attack and defensive components of play separately (7,26). In this paper, the effects of damphetamine on these components of play fighting, using one injected pairmate, were studied. GENERAL METHOD

Subjects Seventy-two female L o n g - E v a n s hooded rats were used. They were born and raised in the animal colony of the Depart-

KTo whom requests for reprints should be addressed. 325

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ment of Psychology at the University of Lethbridge. At weaning (21 days), they were housed as pairs in hanging wire mesh cages (37 × 15 × 25 cm). They were kept in a 12L:12D cycle, lights off at 1900 h. Testing occurred during the light part of the cycle. Temperature was kept constant between (21-23°C). They were provided with Purina Rodent Chow and water ad lib. All animals, in their pairs, were given 20 min exposure a day, for 3 consecutive days, to the test terrarium. The testing terrarium (46 × 46 × 50 cm), consisted of two Plexiglas and two plywood sides. The floor of the terrarium was covered with a 2.5 cm base of processed corn cobs. The white and black pelage patterns were recorded for individual identification.

Procedures Prior to the test trials, the pairmates were separated for 24 h, into individual hanging wire mesh cages (37 x 24 × 18 cm). This enhanced the occurrence of playful activity during testing (10,11 ). Each trial consisted of placing a pair of isolated rats into the test terrarium for 10 min. Sessions were filmed using a Sony High-8 video camera, with settings for artificial light and high sensitivity, at a shutter speed of 250/s. The camera was placed 140 cm above the terrarium perpendicular to the base. Light was provided by a 100 W red bulb placed 60 cm above the terrarium. For each pair, one animal was assigned to the treatment condition and injected 20 min before each trial. Testing was carried out between 30 and 45 days of age. The pairs were tested for two trials on alternate days. Trial one was the pretest trial in which neither animal received an injection. Trial two was the treatment condition in which one of the two animals received an injection of either amphetamine or saline. For all trials, the treated rat was paired with the same nondrugged partner.

Behavioral Analysis Play fighting was divided into two functional components; attack, which involved one rat bringing its snout towards its partner's nape, and defense, which involved the recipient of an attack performing some maneuver to withdraw the nape. In addition, several distinctive types of such defense were distinguishable (20). These behavior patterns were identified and scored using a High-8 video recorder allowing frame by frame, slow motion, and normal speed viewing, as follows: Attack. This was scored when one of the animals brought its snout toward or made contact with the nape of the other animal (15,16,18). Because recipients of such contact could respond before the snout contacted the nape (20), all body contacts by the snout were scored as playful attacks unless specified otherwise. This was calculated as the number of attacks per animal per 10 min. Defense. Withdrawal of the nape area from the attacking partner was scored as a defense (17,18). Because different rats received differing frequencies of attacks, defense was calculated as the probability of responding to an attack (17,18). Defense was first scored as an overall measure. This measure was then broken down into one of several forms of defense (19). These included: a) Evade--this occurs when the recipient of an attack leaped or swerved away to avoid contact with the nape [see Fig. 1A in (21)]. b) Facing defense--in this type of defense the recipient of a nape attack withdrew its nape by changing its body position so as to face the attacking animal. This was classified into the following three types. 1) Complete rotation: this involved the recipient rotating supine, cephalocaudally, along its longitudinal axis [see Fig. 1B in (21)]. 2) Partial rotation: the recipient rotated, cephalocaudally, along its longitudinal axis, but stopped at the pelvis, thus maintaining a standing position on the hind feet [see

14EI~D ANi~ i'EH,IS

Fig. IC in (21)]. 3) Other: in this lorm o! delensc th~ recipient adopted an alternate form of turning to face the attacking animal [see Fig. I in (20)]. When comparing small groups of rats, significant dilterenccs can arise even without treatment (14). Because the baseline for each test group could differ from other test groups, crossgroup comparisons were not meaningful. Therefore, the amount of change between the pretest and treatment trials for all treated animals was calculated. For each animal, the treatment score was subtracted from the pretest score. The derived value was then divided into the pretest score to determine the percentage of change from baseline. These scores were then assigned a positive or negative value, depending on whether the difference represented an increase or decrease in the behavior being measured. This allowed a one-way analysis of variance to be used to evaluate the drug effects for each component of play analyzed. EXPERIMENT

I

Previous experiments have shown that a dose of 0.5 mg/kg of d-amphetamine significantly reduces the occurrence of juvenile play (2,3,26,27). However, as noted above, these studies did not show whether this decrease was due to the attack and/or defense components of play being affected. The aim of this experiment was to determine if the various components of play were affected differentially by d-amphetamine when a treated animal is allowed to interact with a nondrugged partner.

Method Twenty-four juvenile rats were used. Animals were divided into two groups of 12 pairs each; one animal from each pair received either an injection of 0.5 mg/kg d-amphetamine or a 0.9% saline injection of equal volume subcutaneously into the nape.

Results and Discussion Compared to the saline-treated rats, the rats injected with amphetamine showed a significant decrease in the number of attacks initiated, F(I, 11) = 28.95, p < 0.01 (Fig. 1A). In addition, the amphetamine-treated group was significantly less likely to defend against playful attacks, F(1, 1t) = 8.08, p < 0.05 (Fig, 1B). These results suggest that amphetamine, at a dose of 0.5 mg/kg, significantly affects both playful attack and defense. This would support the idea that play fighting, as a whole, decreases after an injection with amphetamine (2,3,26,27). However, frame-by-frame inspection of the videotapes suggested an alternative interpretation. Defensive responses seemed to diminish when contact occurred closer to the site of injection. To quantify this, the nape was divided into the dorsal and lateral aspects and contacts for each were scored separately. Because the rats were filmed from above, the lateral edges of the nape were defined as the sides of the neck and the flat visible surface was defined as the dorsal aspect of the nape. A significant decrease was found in the probability of response to playful attacks directed at the dorsal areas of the nape, F(1, 11) = 14.63, p < 0.05 (Fig. 1C), but not for the lateral areas of the nape (Fig. 1D). This was not the case in the saline condition. Also, no significant differences were found between the saline and amphetamine condition for the various types of defense that were measured. No significant differences were found, on the above measures, for the untreated partners. Because amphetamine has been shown to have small analgesic effects in both experimental animals and man (6) it is possible that the differences in the probability of defense between

AMPHETAMINE AND PLAY FIGHTS

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FIG. I. Mean number of playful attacks (A) and mean probability of playful defense (B), are shown for both saline- and amphetamine-treatedgroups when injected in the nape. The mean probability of defense is contrasted for the dorsal (C) and lateral (D) surfaces of the neck. the dorsal and lateral surfaces of the nape were due to the drug's local anesthetic effect. A greater effect at the site of injection and a reduced effect further from this site is consistent with this possibility. Milder reductions in the effect on the lateral surfaces may reflect a partial diffusion of the anesthesia beyond the site of injection. The nape is the main target area for attack and defense during play fighting (16), and local anesthesia of this area has been shown to diminish play fighting by reduced likelihood of defense (25). Therefore, in the second experiment, an injection site further from the nape was used. EXPERIMENT 2 In this experiment, the hip was chosen as the site of injection because it is not directly involved as a target for playful attack and defense (16). The behaviors of both the treated animal and its untreated partner were analyzed to determine the effects of amphetamine on play. This was done to examine the effects playful interaction has on both the treated animal, as in Experiment

1, and the response of an untreated animal to the altered behaviors presented by the treated animal. In addition, the doses of amphetamine administered were varied so as to determine if different components of play were affected at different doses (2,3,26,27). Method

Forty-eight female rats were used. They were divided into four groups of 12 animals. In the treatment trial, one animal from each pair received either a SC injection of 0.15, 0.5, or 1.0 mg/ kg of d-amphetamine or a 0.9% saline injection of equal volume, in the hip. Results and Discussion

Playful attack was significantly different among conditions, F(3, 23) = 6.40, p < 0.01. Post hoc analyses (Fisher PLSD) revealed that both the 0.5 and 1.0 mg/kg conditions differed significantly from saline (p < 0.05) (Fig. 2A). This is consistent

328

FIELD A N i ) PEI I r i s

PR~TEST [] TREATMENT •

Defensive responses to nape contacts revealed signilicant dilferences between groups for both dorsal, F(3, 23) -: 50.99, p : 0.001, and lateral, F(3, 23) = 3.95, p < (l.05, attacks on the nape (Fig. 3A,B). Post hoc analyses showed that for both dorsal and lateral defense, the 1.0 mg/kg condition differed significantly from all other conditions (p < 0.05). The 0.5 mg/kg dose did not alter the area of the nape defended, as it did in Experiment 1. This further supports that the decline in defense in Experiment 1 was due to the anesthetic effect of amphetamine on the nape area. The significant decreases found at the 1.0 mg/kg dose for both areas of the nape is likely due to the overall decline in

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FIG. 2. Mean frequency for nape attacks and mean probability of defense are shown for all treatment conditions using hip-injected animals.

with the findings of Experiment 1, suggesting that a 0.5 mg/kg dose of amphetamine is decreasing attack through a centrally mediated mechanism. Playful defense was also significantly different among conditions, F(3, 23) = 69.88, p < 0.001. Post hoc analysis showed that the 1.0 mg/kg condition differed significantly from all other groups, (p < 0.05) (Fig. 2B). At the 0.5 mg/kg dose, however, no significant differences were found, suggesting that the significant difference found in defense in Experiment 1 was likely due to the peripheral anesthetic effects of amphetamine on the nape.

1

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FIG. 3. Mean probability of defense to attacks directed toward the dorsal or lateral areas of the nape is shown for all hip-injected treatment conditions.

AMPHETAMINE AND PLAY FIGHTS

defense that was found for this dose. However, this could be due to a direct effect on the neural Systems mediating playful defense, or to some indirect effects of amphetamine on behavior at this higher dose. For example, Schmidt (23,24) demonstrated that the degree of stimulation of the central DA receptors may affect an animal's ability to limit responses to irrelevant external stimuli and focus on salient stimuli. This may then be the case with the response of the 1.0 mg/kg-treated animals. The stimulation of the DA receptors by amphetamine may be limiting the animals' ability to ignore the stimuli provided by its surroundings and focus on the salient stimuli of the attacking animals. Thus, a significant decrease in defense by the treated animal is exhibited. In the juvenile phase, the two most common forms of defense are complete rotations and evasion (21). These two forms of defense were compared to determine if the differences found in defensiveness for the lateral and dorsal areas of the nape at the 1.0 mg/kg dose could be due to a change in the probability of the occurrence of these two forms of defense. Complete rotations differed significantly among conditions, F(3, 23) = 5.75, p < 0.01. Post hoc analyses revealed that the 1.0 mg/kg dose was significantly different from saline (p < 0.05) and that the 0.15 mg/kg dose differed significantly from the 0.5 and 1.0 mg/kg doses (p < 0.05) (Fig. 4A). Although evasion did not differ significantly between conditions, as the dose of amphetamine increased, so did the likelihood of evasions (Fig. 4B). This suggests that there may be an inverse relationship between these two components of defense; as the probability of complete rotations decreased, evasions were likely to increase as a proportion of overall defense. The remaining two forms of defense that were functionally defined earlier, partial rotations and other, showed no significant differences between the saline and amphetamine conditions. The various components of juvenile play were also analyzed in the untreated partners for each treatment condition. No significant differences were found for any of the behavioral components analyzed. A trend, however, was found for the defensive components labeled complete rotation and evasion. As the dose of amphetamine was increased, the probability of complete rotations decreased and the probability of evasion increased. This is similar to the inverse relationship seen between these two components for the treated animals and suggests that interaction with an amphetamine treated animal may have a subtle effect on the type of playful defense exhibited by its untreated partners. GENERAL DISCUSSION Play fighting, as measured by pinning or time spent in play, has been shown to decrease following injections of moderate doses (0.25-1.0 mg/kg) of amphetamine (2,3,26,27). The present study, although confirming these results, suggests that amphetamine has a differential effect on different components of play. Playful attack was reduced at lower doses than was playful defense. Similarly, haloperidol (0.15 and 0.3 mgkg) has been shown to decrease the likelihood of playful attack, but not defense (8), whereas apomorphine, which has been shown to increase play fighting (2), appears to do so by increasing the occurrence of playful attack at doses of 0.05 and 0.3 mg/kg (Pellis and McKenna, unpublished observations). This suggests that playful attack is pharmacologically disassociable from playful defense and, thus, involves separate neural control systems (18). Although the probability of defense was only significantly influenced by the highest dose, lower doses did tend to alter the normai frequency of defensive maneuvers used. Therefore, amphetamine does have an effect on playful defense. Previous work with amphetamine has shown that the effects of amphetamine on play are centrally mediated, because 4 OH-

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FIG. 4. Mean probability of complete rotations and evasion for the four treatment conditions in Experiment 2 are shown. amphetamine, which does not easily cross the blood-brain barrier, does not decrease play fighting (3). Even so, at present there is no evidence that amphetamine affects play fighting through its action as an indirect catecholamine agonist (4). The findings from Experiment 1, however, showed that amphetamine can affect play fighting via its peripheral effects. The nape is the target of attack and defense during play fighting by rats (16). Anesthetizing the nape with xylocaine reduces playfulness (25). Similarly, injection with amphetamine into the nape appears to have a similar effect because playful defense is decreased at a dose that does not do so when injected at another site (Experiment 1 vs. 2). These findings underscore the impor-

FIEIA) ANt) PEII,IS

330

tance of not using the nape as the site of injection in pharmacological studies of play fighting in rats (12). This study suggests that detailed analyses of the c o m p o nents of a b e h a v i o r such as play fighting are n e c e s s a r y for d e t e r m i n i n g the effects o f p h a r m a c o l o g i c a l agents. Such analyses are n e e d e d to d e t e r m i n e w h e t h e r the agent b e i n g tested is h a v i n g a u n i f o r m effect on all aspects of a b e h a v i o r or

w h e t h e r certain aspects of a b e h a v i o r are being differenlially affected at different doses, ACKNOWLEDGEMENTS We wish to thank Vivien C. Pellis for her comments on earlier drafts. This research was supported in part by a grant from the Natural Sciences and Engineering Research Council of Canada to S. Pellis.

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