Effect of Postnatal Litter Size on Adult Aggression in the Laboratory Mouse VIRGINIA RYAN FRANCINE WEHMER Department of Psychology Wayne State Utziversity Detroit, Michigan
Growth, emotionality, food competition, and aggression were examined in mice nursed in litters of 3 or 9 and rearcd in isolation until testing. Animals from large litters were lighter at weaning and in adulthood and were more emotional in the open field than subjects from small litters. They did not win more food competition tests than subjects from small litters although their consummatory behavior during food competition tests was greater. Subjects from large litters were more aggressive in initial encounters, but over repeated encounters became more submissive. In a 2nd open-field test, emotionality of large-litter subjects was reduced more than that of subjects from small litters. When later placed in group-living cages, subjects from small litters sustained less long term physical assault than subjects from large litters. High correlations were found between the 4 measures of brief aggression.
The effects of postpartum nutrition during the preweaning period have been studied by fostering rodent pups at birth so as to constitute litters ranging in size from 3 to 18, thus increasing or decreasing the supply of milk available to each of the neonates (Chase, Dorsey, & McKhann, 1967; LaBarba & White, 1971; Winick & Noble, 1967). An alternate procedure for providing differential nutrition to nursing pups is to maintain nursing mothers on diets which differ with respect t o percent of protein calories (Fra'ikovi, 1972; Levitsky & Barnes, 1972). Rat subjects from large litters are more active (Fraskovi, 1972) and more successful in food competition (Seitz, 1954) than subjects from small litters. Although variations in the size of mouse litters affects components of emotionality (Egan & Royce, 1973; LaBarba &White, 1971), the possibility that differential litter size affects adult aggressive behavior has not been investigated. The outcomes of aggression tests between mice, varied with respect to litter size, might be influenced by several variables. Bronfenbrenner (1968) has hypothesized that drive deprivation during infancy increases general drive level in adulthood and specifically increases drive related to the previous deprivation, particularly when the animal is again placed in the deprived state. Aggression is also related t o dominance over conspecifics. The dominance hierarchies of rodents established under one set of testing conditions are not necessarily representative
Received for publication 22 July 1974 Revised for publication 20 November 1974 DevelopmentalPsychobiology, 8(4): 363-370 (1975) @ 1975 by John Wiley & Sons, Inc.
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of the hierarchies established by the same subjects when parameters of size of encounter area and presence of food or water are manipulated (Baenninger, 1970; Lindzey, Manosevitz, & Winston, 1966). In addition, rodent hierarchies are initially unstable, but tend to increase in stability after repeated testing (Candland & Bloomquist, 1965; Wilson, 1968). In the present study we investigated the effects of differential litter size on the aggressive behavior of mice. We also assessed the possible influences of presence of food, size of testing area, and number of encounters on the outcome of dominance tests. Further, we examined the representativeness of hierarchies established on the basis of dyadic test encounters, and compared these hierarchies to those established in group living conditions.
Method S u biect s Fifteen primiparous pregnant female mice (Mus rnusculus) of the Swiss-Webster strain were procured through the Animal Resources Department of Wayne State University and individually caged in plastic “shoe box” type cages (31.5 x 15.3 x 20.4 cm). On the day following parturition, half of the litters were randomly culled to 9 or 3 pups. Fostering was not required in that all litters were originally larger than 9. Pups remained with their mothers and littennates until 33 days of age. Purina Mouse Chow and water were available to the pups ad lib w h l e they remained in the maternal cages. At 3 3 days of age, pups were individually housed in cages identical to those described above. (We extended the weaning age to insure similar maternal care for pup groups maturing at different rates due to differential nutritional conditions [Simonson & Chow, 19691 . However, overnutrition which might result from extended maternal contact in small litters has not been shown to produce animals which demonstrate superior adult behaviors [Frayikovli, 19701 .) Those animals maintained in large litters prior to weaning were designated Large Litter subjects (LL‘s) and those animals from small litters were designated Small Litter subjects (SL‘s). Ten male LL‘s an 10 male SL‘s, no more than 2 from any given litter, were randomly selected as experimental subjects. Sixteeen additional LL’s and SL‘s from an earlier pilot study were used in the home cage dominance phase of the study, with n o more than 2 males from a litter used in the study.
Apparatus Open field. Measures of emotionality and activity were taken in an open box, 94.5 cm square with sides 20.4 cm, constructed of wood and painted gray. Black lines were painted on the floor to form a grid. The lines, placed every 7.6 cm, were sequentially numbered on the interior sides of the box. Large-field free aggression. These tests were run in the same apparatus described above. The box was slotted on 2 sides to allow for the insertion of a partition constructed of #16 gauge, 2 x 2 (each square, 1.3 x 1.3 cm) hardware cloth. When this partition was in place, the box was bisected into equal sections measuring 94.5 x 46.8 x 20.4 cm.
LITTER SIZE AND AGGRESSlON
365
Large-field food aggression. These tests occurred in a box similar to that of large-field free aggression but with a 1.9-cm hole in the middle of the field. A cup of Purina Mouse Chow was placed beneath the hole in the floor of the field. The food was accessible to only 1 animal at a time and was reached by the animal placing its head through the hole into the cup. A hardware cloth partition similar to that described above was also provided for this apparatus. When the partition was in place, bisecting the box, it also bisected the food hole and prevented all access to the food. Small-field ,free aggression. These tests took place in a box measuring 8.9 x 17.8 x 15.2 cm. A hardware cloth partition as described for the large-field free aggression apparatus was provided and, when placed in position, this partition bisected the box into equal 8.9-cm-square compartments. A #16 gauge, 2 x 2 hardware cloth cover was provided for the top. Smal1,ft’eld ,food aggression. The apparatus for these tests was similar to that described for small-field free aggression. However, like the apparatus described for larg-e-fieldfood aggression, it was provided with a hole in the center and a food cup from which the animals could feed.
Procedure When the subjects were 33 days old, they were weighed and placed in individual cages. Food and water were available ad lib. At 70 days of age subjects were again weighed, coded with indelible ink of various colors painted on different parts of the body, and given 5 2-min testing sessions in the open field. Animals were placed in one corner of the apparatus and their latencies to move were noted. These latencies constituted their emotionality scores. The numbers of lines the animals crossed as they moved around the apparatus were also noted and constituted their activity scores. After these initial tests, when the subjects were 75 days old, they were placed on a mild restricted feeding schedule of 4.5 g of Purina Mouse Chow per day until the termination of the brief period of dominance testing. Average weight loss on this schedule over the 30 days during which it was enforced was 5%. After this schedule was implemented, subjects were placed alone in each of the apparatuses for a 5-min period 3 days before formal testing began. In the apparatus containing food cups the 5-min period did not commence until the animal had approached the food. When 80 days old, every animal in the LL group was tested against every animal in the SL group in each type of encounter apparatus. Thus each animal met every animal that was not in his litter size group 4 times, In this way each animal had 40 dominance tests. Tests were arranged so that any given pair of animals encountered each other only after they had encountered 9 other animals since their last encounter. Each animal received a unique sequence of tests and no animal was given the same test twice in a row. The mice were tested over a 3-week period and were given an average of 4 tests per day at I-hr intervals, 3 days each week. Large-field food aggression. Two animals were placed simultaneously in this apparatus, separated from each other by the partition. The partition was removed after 30 sec. Animals remained in this apparatus for 2 min or until one animal attacked the other, whichever came first. A record of which animal attacked was made. The number of seconds each animal spent feeding was also noted. Each animal was scored “1” for an attack, “0” if it did not attack, and “.5” if it and its opponent attacked simultaneously.
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Large-field free aggression. Scoring procedures for this test were the same as those used in lapge-fieldfood aggression noted above, except that no food was present. Small-field free aggression and foud aggression. These 2 tests were conducted similarly to those noted above. After all aggression tests had been run, open-field behavior was measured a 2nd time in 3 sessions. Subjects were at this time 100 days old. They were weighed a 3rd time and placed on a schedule of ad lib food and water. Group living. Ten days after the subjects were returned to an ad lib feeding schedule, they were randomly housed together in groups of 4 containing 2 LL's and 2 SL's. Litter mates were is no case assigned to the same living group. Animals remained in these groups for a week, in the same plastic shoe-box cages used previously. The dominance hierarchy that developed over the week was scored. Ranking was based on the animal's physical Condition. The animal in each cage with the greatest number of tail scars andmost extensively chewed flanks (or those animals that were killed) were given a score of 4. That animal in each group which had the fewest scars and was in the best physical condition was given a score of 1. Each subject was ranked within its group and when no difference was apparent subjects were given tied ranks. Animals earning a score of 1 or 2 were considered dominant within the group: animals with scores of 3 or 4 were considered submissive.
Results Body Weight The mean weights of the SL and LL groups are summarized in Table 1. The SL's weighed significantly more than the LL's at weaning, 70 days, and 100 days of age ( F = 21.4, df = 1/SO, p < .0l). This difference in weight was maintained throughout testing and did not decrease significantly with age (F < I).
Open-Field Latency Latency to initiate activity in the open field was longer in the LL group that in the SL group (F = 8.03, df= 1/36, p < .01). Latency of the LL's decreased in the 2nd test whereas the latency of the SL's did not (F = 10.5, d f = 1/36, p < 01 ;see Fig. 1).
Open-Field Activity Open-field activity of the 2 groups did not differ either before or after dominance testing. Activity of both groups of animals increased following the dominance testing (F = 12.2,df = 1 / 3 6 , p < .01).
TABLE 1. Mean Body Weight.(g) ~
LL's SL'S
~~~~~~~
Weaning
70 Days
100 Days
33.1 38.5
44.1 47.5
42.0 45.3
LITTER SIZE AND AGGRESSION
367
70 100 DAYS of AGE ______-. =. L L'S = SL's
-
Fig. 1. Latency to move in the open field.
Aggression The 2 groups did not differ in the number of tests won in either size of enclosure with or without food ( a l l F s < 1). The SL's won a total of 159 tests; LL's won 147. Although the type of test did not appear to be a significant variable, previous experience was of considerable importance in determining the outcome of encounters over time (see Fig. 2). The LL's were initially more aggressive than were SL's. In the first 100 tests LL's aggressed 56.5 times (nearly twice the number of times that animals from small litters aggressed). Prolonged exposure to dominance testing resulted in a constant decrease in aggression for the LL's. Thus, this group aggressed only 21 times in the last 100 tests. The decrease in aggression in the LL's as a function of experience was the reverse of the trend found for the SL's. In the first 100 tests SL's aggressed only 32 times. After this initial experience these animals increased their aggressions to an average of 42 per 100 encounters. This interaction between test blocks was significant (F = 5.8, df = 1/36, p < .02). The extent to which the 4 types of brief dominance tests are correlated can be seen in Table 2; all coefficients are significant beyond the .01 level.
m. of wins
--__ 30 numbrer
--..--. --. -X
20
1-100
101-200
201-300
301-400
TESTS = L L's
-___--_ -=
SL's
Fig. 2. Dominance tests won over time.
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KYAN AND WEHMER
TABLE 2. Mutrix ofliaizk Correlutioris 011 Dominance Tests.
SrnaU-13eId I:ood
Small-Field Food
Small-Field No Food
1,arge-Field Food
-
.88“
.89a
3 1
-
.75a
.81“
-
.IP
Small-Field No Food Large-Field I;ood Large- Field No Food
L.arge-I.’ield N o Food
-~
-
The LL‘s spent niore time eating than the SL’s (F’ = 21 5,df = 1/36, p < .Ol). Allhough both groups of subjects spent more time eating i n the large encounter area than the small (F = 8.4. df = 1!36, p < .01) the IL’s demonstrated a greater decrease o f consummatory behavior in the small encounter area than the SL‘s (F = 5.4, clj‘= 1/36, p < .03,5).
Home Cage Dominance When the animals were placed 4 in a cage, with each cage containing 2 LL‘s and 3, SL‘s: an SL animal ranked highest in 8 o f 9 cages (Binomial Test, p < .02). Subjects were given a score of “ I ” for each subject of the alternate treatment over which they were domillant and a score o f “S” for each tie. Analysis (if the dominaiice rankings achieved by all the animals revealed that SL‘s were doinillant over 28.5 LL’s whereas LL’s were dominant over 7 . 5 SL’s (xz = 5.4, dj’ = I , I.] < .07_).Body weight and dominance status in the home cage were not correlated ( r = .1Y).
Discussion Body Weight The nutritional stress involved i n nuising a large litter was reflected in lower weight of offspring and persistent growth ietardation in adulthood even after postweaning ad lib feeding. These data support other studies of permanent growth retardation of animals reared in large litteis (Frahhovi. 1968; Gutlrrie, 1968).
Emotionality Animals from large litters were more emotional than animals froin small litters both before and after dominance testing. These results are comparable to those found by
LITTER SIZE AND AGGRESSION
369
IaBarba and White (1971) and Manosevitz and McCanne (1973). The sources of these differences in emotionality are ciirrently unknown, but several possibilities exist. Accelerated growth induces the early onset of function of several physiological systems, including the hypothalamic-pituitary-adrenal axis (Milkovic & Milkovic. 1959). The function of this system and levels of plasma corticosteroids have repeatedly been related to levels of emotionality (Bronson & Eleftheriou, 1963; Howard, 1973; Levine & Mullins, 1968). Although the hightened emotionality found in animals from large litters may be the consequence of differential functioning of the adrenocortical system, the contributions of nutritional and maternal stress factors have not yet been determined (Plaut, 1970).
Aggression Contrary to early reports (e.g., Seitz, 1954) animals from large litters did not win more often in food competition tests. This finding supports the negative finding receiitly reported by Manosevitz and McCanne (1 973) on the effects o f postweaning deprivation of food on competition in A/J and DBA/25 mice. However, the initial aggressiveness o f animals from large litters was in accord with expectations based on Seitz’ data and Bronft;.,brenner’s (1968) hypothesis of the drive-inducing effect of early deprivation. The change i n behavior over time with respect to dominance testing was an unexpected result. Ginzberg and Allee (1942) suggested that a history of successful dominance encounters increases aggressive behavior in mice. Yet subjects from large litters, which were significantly niore aggressive at first, became more submissive over time. Thus the reported initial aggressiveness of early malnourished animals (Levitsky & Barnes, 1972) may be a transient correlate of their heightened emotionality, and not reflective of their ability to establish long term successful aggressive postures. The lack of correlation between body weight and home-cage dominance ranking indicates that dominance was not merely a reflection of the size of animals. The cori-elations between the 4 types of brief tests of aggression are in contrast to previous reports that rodent hierarchies vary with test parameters (e.g., Baenninger. 1970; Lindzey et al., 1966). In the present experiment mice froni sinall litters were dominant after experience in all types o f ecnounters. Levitsky and Barnes (1970) suggested that early malnutrition lowers the thresliold of the stress response system. Differences in emotionality and stress thresholds between the large- and small-litter subjects may have been sufficient t o determine the outcomes of dominance encounters, regardless of type o f encoii n te r .
Notes This report describes work done in partial fulfilliuciit of the M.A. degree awarded to the first author by thc Dcpartmcnt o f Psychology. Wayne S t a t c University. This was supported by G r a n t No. 1 1:03 MH54703-01 froin the Nationd Institute of Mental llealth. Request for rcpririts d ~ o u l dbc sent to: Dr. Francine Wchmer. Department of Psychology, Wayne State University, Detroit. Michigan 48202. U.S.A.
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References Baenningcr, L. P. (1970). Social dominance orders in the rat: ‘Spontaneous” food and water competition. J. Comp. Physiol. Psychol., 71: 202-209. Bronfenbrenner, U. (1968). Early deprivation in mammals: A cross-species analysis. In G . Newton and S. Levine (Eds.), Early Experience and Behavior. Springfield: Thomas. Pp. 627-765. Bronson, F. J., and Elcftheriou, B. E. (1963). Adrenal response to crowding in peromyscus and C57BL/10J mice. Physiol. Zoo[., 36: 161-166. Candland, D. K., and Bloomquist, D. W. (1965). Interspecies comparisons of the reliability of dominance orders. J. Comp. Physiol. Psychol., 59: 135-137. Chase, H. P., Dorsey, J., and McKhann, G. M. (1967). The effect of malnutrition on the synthesis of a myelin lipid. Pediatrics, 40: 55 1-560. Egan, O., and Royce, J. R. (1973). Litter size and emotionality in two strains of mice. J. Comp. Physiol. Psychol., 83: 55-59. b’rankova, S. (1968). Nutritional and psychological factors in the development of spontaneous behavior in the rat. In N. S. Scrimshaw and J. E. Gordon (Eds.), Malnutrition, Learning and Behavior. Cambridge: Massachusetts Institute of Technology Press. Pp. 316-322. Frankova, S. (1970). Behavioral responses of rats to early overnutrition. Nutr. Metabol., 12: 228-239. Frankova, S. (1972). Influence of nutrition and early experience on behavior of rats. Bibl. Nutr. Dieia, 17: 96-1 10. Ginsberg, B. C., and Allee, W. C:. (1942). Some effects of conditioning on social dominance and subordination in inbred strains of mice. Physiol. Zool., 15: 485-506. Guthrie, H. A. (1968). Severe undernutrition in early infancy and behavior in rehabitated albino rats. Physiol. Behav., 3: 619-623. Howard, E. (19’73). Increased reactivity and impaired adaptability in operant behavior of adult mice given corticostcrone in infancy. J. Comp. Physiol. Psychol., 85: 21 1-220. LaBarba, R. C., and White, J. L. (1971). Litter size variations and emotional reactivity in BALB/c mice. J. Comp, Physiol. PsychoL, 75: 254-251. Levine, S., and Mullins, R. F. (1968). Hormones in infancy. In G. Newton and S. Levine (Eds.), Early Experience and Behavior. Springfield: Thomas. Pp. 169-197. Levitsky, D. A., and Barnes, R. I-I. (1970). Effects of early malnutrition on the reaction of adult rats to aversive stimuli. Nature (Lond.),225: 468-469. Levitsky, D. A., and Barnes, K. H. (1972). Nutritional and environmental interactions in the bchavioral dcvelopment of the rat: Long-term effects. Science, 176: 68-71. Lindzey, G., Manosevitz, M., and Winston, H. (1966). Social dominance in the mouse. Psychon. Sci., 5: 68-71. Manosevitz, M., and McCanne, T. R. (1973), Effects of early food deprivation on mouse behavior. J. Comp. Physiol. Psychol., 83: 3 14-323. Milkovic, K., and Milkovic, S . (1’359). Reactiveness of the pituitary-adrenal system in some laboratory mammals. Endokrinologie, 37: 301-310. Plaut, S. M. (1970). Studies of undernutrition in the young rat: Methodological considerations. Dev. Psychobiol., 3: 15’7-167. Seitz, P. I:. D. (1954). The effects of infantile experiences upon adult behavior in animal subject: I. Effects of litter size during infancy upon adult behavior in the rat. Am. J. Psychiatry, 110: 9 16-927. Simonson, M., and Chow, B. W. (1969). Maze studies on the progeny of underfed mother rats. J. Nutr., 100: 685-690. Wilson, W. J. (1968). Adaptation to the Dominance Tube. Psychon. Sci., 10; 119-120. Winick, M., and Noble, A. (1967). Cellular response with increased feeding in neonatal rats. J. Nutr., 91: 179-182.
Growth, emotionality, food competition, and aggression were examined in mice nursed in litters of 3 or 9 and rearcd in isolation until testing. Animals from large litters were lighter at weaning and in adulthood and were more emotional in the open field than subjects from small litters. They did not win more food competition tests than subjects from small litters although their consummatory behavior during food competition tests was greater. Subjects from large litters were more aggressive in initial encounters, but over repeated encounters became more submissive. In a 2nd open-field test, emotionality of large-litter subjects was reduced more than that of subjects from small litters. When later placed in group-living cages, subjects from small litters sustained less long term physical assault than subjects from large litters. High correlations were found between the 4 measures of brief aggression.
The effects of postpartum nutrition during the preweaning period have been studied by fostering rodent pups at birth so as to constitute litters ranging in size from 3 to 18, thus increasing or decreasing the supply of milk available to each of the neonates (Chase, Dorsey, & McKhann, 1967; LaBarba & White, 1971; Winick & Noble, 1967). An alternate procedure for providing differential nutrition to nursing pups is to maintain nursing mothers on diets which differ with respect t o percent of protein calories (Fra'ikovi, 1972; Levitsky & Barnes, 1972). Rat subjects from large litters are more active (Fraskovi, 1972) and more successful in food competition (Seitz, 1954) than subjects from small litters. Although variations in the size of mouse litters affects components of emotionality (Egan & Royce, 1973; LaBarba &White, 1971), the possibility that differential litter size affects adult aggressive behavior has not been investigated. The outcomes of aggression tests between mice, varied with respect to litter size, might be influenced by several variables. Bronfenbrenner (1968) has hypothesized that drive deprivation during infancy increases general drive level in adulthood and specifically increases drive related to the previous deprivation, particularly when the animal is again placed in the deprived state. Aggression is also related t o dominance over conspecifics. The dominance hierarchies of rodents established under one set of testing conditions are not necessarily representative
Received for publication 22 July 1974 Revised for publication 20 November 1974 DevelopmentalPsychobiology, 8(4): 363-370 (1975) @ 1975 by John Wiley & Sons, Inc.
36 3
364
RYAN AND WEHMEK
of the hierarchies established by the same subjects when parameters of size of encounter area and presence of food or water are manipulated (Baenninger, 1970; Lindzey, Manosevitz, & Winston, 1966). In addition, rodent hierarchies are initially unstable, but tend to increase in stability after repeated testing (Candland & Bloomquist, 1965; Wilson, 1968). In the present study we investigated the effects of differential litter size on the aggressive behavior of mice. We also assessed the possible influences of presence of food, size of testing area, and number of encounters on the outcome of dominance tests. Further, we examined the representativeness of hierarchies established on the basis of dyadic test encounters, and compared these hierarchies to those established in group living conditions.
Method S u biect s Fifteen primiparous pregnant female mice (Mus rnusculus) of the Swiss-Webster strain were procured through the Animal Resources Department of Wayne State University and individually caged in plastic “shoe box” type cages (31.5 x 15.3 x 20.4 cm). On the day following parturition, half of the litters were randomly culled to 9 or 3 pups. Fostering was not required in that all litters were originally larger than 9. Pups remained with their mothers and littennates until 33 days of age. Purina Mouse Chow and water were available to the pups ad lib w h l e they remained in the maternal cages. At 3 3 days of age, pups were individually housed in cages identical to those described above. (We extended the weaning age to insure similar maternal care for pup groups maturing at different rates due to differential nutritional conditions [Simonson & Chow, 19691 . However, overnutrition which might result from extended maternal contact in small litters has not been shown to produce animals which demonstrate superior adult behaviors [Frayikovli, 19701 .) Those animals maintained in large litters prior to weaning were designated Large Litter subjects (LL‘s) and those animals from small litters were designated Small Litter subjects (SL‘s). Ten male LL‘s an 10 male SL‘s, no more than 2 from any given litter, were randomly selected as experimental subjects. Sixteeen additional LL’s and SL‘s from an earlier pilot study were used in the home cage dominance phase of the study, with n o more than 2 males from a litter used in the study.
Apparatus Open field. Measures of emotionality and activity were taken in an open box, 94.5 cm square with sides 20.4 cm, constructed of wood and painted gray. Black lines were painted on the floor to form a grid. The lines, placed every 7.6 cm, were sequentially numbered on the interior sides of the box. Large-field free aggression. These tests were run in the same apparatus described above. The box was slotted on 2 sides to allow for the insertion of a partition constructed of #16 gauge, 2 x 2 (each square, 1.3 x 1.3 cm) hardware cloth. When this partition was in place, the box was bisected into equal sections measuring 94.5 x 46.8 x 20.4 cm.
LITTER SIZE AND AGGRESSlON
365
Large-field food aggression. These tests occurred in a box similar to that of large-field free aggression but with a 1.9-cm hole in the middle of the field. A cup of Purina Mouse Chow was placed beneath the hole in the floor of the field. The food was accessible to only 1 animal at a time and was reached by the animal placing its head through the hole into the cup. A hardware cloth partition similar to that described above was also provided for this apparatus. When the partition was in place, bisecting the box, it also bisected the food hole and prevented all access to the food. Small-field ,free aggression. These tests took place in a box measuring 8.9 x 17.8 x 15.2 cm. A hardware cloth partition as described for the large-field free aggression apparatus was provided and, when placed in position, this partition bisected the box into equal 8.9-cm-square compartments. A #16 gauge, 2 x 2 hardware cloth cover was provided for the top. Smal1,ft’eld ,food aggression. The apparatus for these tests was similar to that described for small-field free aggression. However, like the apparatus described for larg-e-fieldfood aggression, it was provided with a hole in the center and a food cup from which the animals could feed.
Procedure When the subjects were 33 days old, they were weighed and placed in individual cages. Food and water were available ad lib. At 70 days of age subjects were again weighed, coded with indelible ink of various colors painted on different parts of the body, and given 5 2-min testing sessions in the open field. Animals were placed in one corner of the apparatus and their latencies to move were noted. These latencies constituted their emotionality scores. The numbers of lines the animals crossed as they moved around the apparatus were also noted and constituted their activity scores. After these initial tests, when the subjects were 75 days old, they were placed on a mild restricted feeding schedule of 4.5 g of Purina Mouse Chow per day until the termination of the brief period of dominance testing. Average weight loss on this schedule over the 30 days during which it was enforced was 5%. After this schedule was implemented, subjects were placed alone in each of the apparatuses for a 5-min period 3 days before formal testing began. In the apparatus containing food cups the 5-min period did not commence until the animal had approached the food. When 80 days old, every animal in the LL group was tested against every animal in the SL group in each type of encounter apparatus. Thus each animal met every animal that was not in his litter size group 4 times, In this way each animal had 40 dominance tests. Tests were arranged so that any given pair of animals encountered each other only after they had encountered 9 other animals since their last encounter. Each animal received a unique sequence of tests and no animal was given the same test twice in a row. The mice were tested over a 3-week period and were given an average of 4 tests per day at I-hr intervals, 3 days each week. Large-field food aggression. Two animals were placed simultaneously in this apparatus, separated from each other by the partition. The partition was removed after 30 sec. Animals remained in this apparatus for 2 min or until one animal attacked the other, whichever came first. A record of which animal attacked was made. The number of seconds each animal spent feeding was also noted. Each animal was scored “1” for an attack, “0” if it did not attack, and “.5” if it and its opponent attacked simultaneously.
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RYAN AND WEHMER
Large-field free aggression. Scoring procedures for this test were the same as those used in lapge-fieldfood aggression noted above, except that no food was present. Small-field free aggression and foud aggression. These 2 tests were conducted similarly to those noted above. After all aggression tests had been run, open-field behavior was measured a 2nd time in 3 sessions. Subjects were at this time 100 days old. They were weighed a 3rd time and placed on a schedule of ad lib food and water. Group living. Ten days after the subjects were returned to an ad lib feeding schedule, they were randomly housed together in groups of 4 containing 2 LL's and 2 SL's. Litter mates were is no case assigned to the same living group. Animals remained in these groups for a week, in the same plastic shoe-box cages used previously. The dominance hierarchy that developed over the week was scored. Ranking was based on the animal's physical Condition. The animal in each cage with the greatest number of tail scars andmost extensively chewed flanks (or those animals that were killed) were given a score of 4. That animal in each group which had the fewest scars and was in the best physical condition was given a score of 1. Each subject was ranked within its group and when no difference was apparent subjects were given tied ranks. Animals earning a score of 1 or 2 were considered dominant within the group: animals with scores of 3 or 4 were considered submissive.
Results Body Weight The mean weights of the SL and LL groups are summarized in Table 1. The SL's weighed significantly more than the LL's at weaning, 70 days, and 100 days of age ( F = 21.4, df = 1/SO, p < .0l). This difference in weight was maintained throughout testing and did not decrease significantly with age (F < I).
Open-Field Latency Latency to initiate activity in the open field was longer in the LL group that in the SL group (F = 8.03, df= 1/36, p < .01). Latency of the LL's decreased in the 2nd test whereas the latency of the SL's did not (F = 10.5, d f = 1/36, p < 01 ;see Fig. 1).
Open-Field Activity Open-field activity of the 2 groups did not differ either before or after dominance testing. Activity of both groups of animals increased following the dominance testing (F = 12.2,df = 1 / 3 6 , p < .01).
TABLE 1. Mean Body Weight.(g) ~
LL's SL'S
~~~~~~~
Weaning
70 Days
100 Days
33.1 38.5
44.1 47.5
42.0 45.3
LITTER SIZE AND AGGRESSION
367
70 100 DAYS of AGE ______-. =. L L'S = SL's
-
Fig. 1. Latency to move in the open field.
Aggression The 2 groups did not differ in the number of tests won in either size of enclosure with or without food ( a l l F s < 1). The SL's won a total of 159 tests; LL's won 147. Although the type of test did not appear to be a significant variable, previous experience was of considerable importance in determining the outcome of encounters over time (see Fig. 2). The LL's were initially more aggressive than were SL's. In the first 100 tests LL's aggressed 56.5 times (nearly twice the number of times that animals from small litters aggressed). Prolonged exposure to dominance testing resulted in a constant decrease in aggression for the LL's. Thus, this group aggressed only 21 times in the last 100 tests. The decrease in aggression in the LL's as a function of experience was the reverse of the trend found for the SL's. In the first 100 tests SL's aggressed only 32 times. After this initial experience these animals increased their aggressions to an average of 42 per 100 encounters. This interaction between test blocks was significant (F = 5.8, df = 1/36, p < .02). The extent to which the 4 types of brief dominance tests are correlated can be seen in Table 2; all coefficients are significant beyond the .01 level.
m. of wins
--__ 30 numbrer
--..--. --. -X
20
1-100
101-200
201-300
301-400
TESTS = L L's
-___--_ -=
SL's
Fig. 2. Dominance tests won over time.
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TABLE 2. Mutrix ofliaizk Correlutioris 011 Dominance Tests.
SrnaU-13eId I:ood
Small-Field Food
Small-Field No Food
1,arge-Field Food
-
.88“
.89a
3 1
-
.75a
.81“
-
.IP
Small-Field No Food Large-Field I;ood Large- Field No Food
L.arge-I.’ield N o Food
-~
-
The LL‘s spent niore time eating than the SL’s (F’ = 21 5,df = 1/36, p < .Ol). Allhough both groups of subjects spent more time eating i n the large encounter area than the small (F = 8.4. df = 1!36, p < .01) the IL’s demonstrated a greater decrease o f consummatory behavior in the small encounter area than the SL‘s (F = 5.4, clj‘= 1/36, p < .03,5).
Home Cage Dominance When the animals were placed 4 in a cage, with each cage containing 2 LL‘s and 3, SL‘s: an SL animal ranked highest in 8 o f 9 cages (Binomial Test, p < .02). Subjects were given a score of “ I ” for each subject of the alternate treatment over which they were domillant and a score o f “S” for each tie. Analysis (if the dominaiice rankings achieved by all the animals revealed that SL‘s were doinillant over 28.5 LL’s whereas LL’s were dominant over 7 . 5 SL’s (xz = 5.4, dj’ = I , I.] < .07_).Body weight and dominance status in the home cage were not correlated ( r = .1Y).
Discussion Body Weight The nutritional stress involved i n nuising a large litter was reflected in lower weight of offspring and persistent growth ietardation in adulthood even after postweaning ad lib feeding. These data support other studies of permanent growth retardation of animals reared in large litteis (Frahhovi. 1968; Gutlrrie, 1968).
Emotionality Animals from large litters were more emotional than animals froin small litters both before and after dominance testing. These results are comparable to those found by
LITTER SIZE AND AGGRESSION
369
IaBarba and White (1971) and Manosevitz and McCanne (1973). The sources of these differences in emotionality are ciirrently unknown, but several possibilities exist. Accelerated growth induces the early onset of function of several physiological systems, including the hypothalamic-pituitary-adrenal axis (Milkovic & Milkovic. 1959). The function of this system and levels of plasma corticosteroids have repeatedly been related to levels of emotionality (Bronson & Eleftheriou, 1963; Howard, 1973; Levine & Mullins, 1968). Although the hightened emotionality found in animals from large litters may be the consequence of differential functioning of the adrenocortical system, the contributions of nutritional and maternal stress factors have not yet been determined (Plaut, 1970).
Aggression Contrary to early reports (e.g., Seitz, 1954) animals from large litters did not win more often in food competition tests. This finding supports the negative finding receiitly reported by Manosevitz and McCanne (1 973) on the effects o f postweaning deprivation of food on competition in A/J and DBA/25 mice. However, the initial aggressiveness o f animals from large litters was in accord with expectations based on Seitz’ data and Bronft;.,brenner’s (1968) hypothesis of the drive-inducing effect of early deprivation. The change i n behavior over time with respect to dominance testing was an unexpected result. Ginzberg and Allee (1942) suggested that a history of successful dominance encounters increases aggressive behavior in mice. Yet subjects from large litters, which were significantly niore aggressive at first, became more submissive over time. Thus the reported initial aggressiveness of early malnourished animals (Levitsky & Barnes, 1972) may be a transient correlate of their heightened emotionality, and not reflective of their ability to establish long term successful aggressive postures. The lack of correlation between body weight and home-cage dominance ranking indicates that dominance was not merely a reflection of the size of animals. The cori-elations between the 4 types of brief tests of aggression are in contrast to previous reports that rodent hierarchies vary with test parameters (e.g., Baenninger. 1970; Lindzey et al., 1966). In the present experiment mice froni sinall litters were dominant after experience in all types o f ecnounters. Levitsky and Barnes (1970) suggested that early malnutrition lowers the thresliold of the stress response system. Differences in emotionality and stress thresholds between the large- and small-litter subjects may have been sufficient t o determine the outcomes of dominance encounters, regardless of type o f encoii n te r .
Notes This report describes work done in partial fulfilliuciit of the M.A. degree awarded to the first author by thc Dcpartmcnt o f Psychology. Wayne S t a t c University. This was supported by G r a n t No. 1 1:03 MH54703-01 froin the Nationd Institute of Mental llealth. Request for rcpririts d ~ o u l dbc sent to: Dr. Francine Wchmer. Department of Psychology, Wayne State University, Detroit. Michigan 48202. U.S.A.
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References Baenningcr, L. P. (1970). Social dominance orders in the rat: ‘Spontaneous” food and water competition. J. Comp. Physiol. Psychol., 71: 202-209. Bronfenbrenner, U. (1968). Early deprivation in mammals: A cross-species analysis. In G . Newton and S. Levine (Eds.), Early Experience and Behavior. Springfield: Thomas. Pp. 627-765. Bronson, F. J., and Elcftheriou, B. E. (1963). Adrenal response to crowding in peromyscus and C57BL/10J mice. Physiol. Zoo[., 36: 161-166. Candland, D. K., and Bloomquist, D. W. (1965). Interspecies comparisons of the reliability of dominance orders. J. Comp. Physiol. Psychol., 59: 135-137. Chase, H. P., Dorsey, J., and McKhann, G. M. (1967). The effect of malnutrition on the synthesis of a myelin lipid. Pediatrics, 40: 55 1-560. Egan, O., and Royce, J. R. (1973). Litter size and emotionality in two strains of mice. J. Comp. Physiol. Psychol., 83: 55-59. b’rankova, S. (1968). Nutritional and psychological factors in the development of spontaneous behavior in the rat. In N. S. Scrimshaw and J. E. Gordon (Eds.), Malnutrition, Learning and Behavior. Cambridge: Massachusetts Institute of Technology Press. Pp. 316-322. Frankova, S. (1970). Behavioral responses of rats to early overnutrition. Nutr. Metabol., 12: 228-239. Frankova, S. (1972). Influence of nutrition and early experience on behavior of rats. Bibl. Nutr. Dieia, 17: 96-1 10. Ginsberg, B. C., and Allee, W. C:. (1942). Some effects of conditioning on social dominance and subordination in inbred strains of mice. Physiol. Zool., 15: 485-506. Guthrie, H. A. (1968). Severe undernutrition in early infancy and behavior in rehabitated albino rats. Physiol. Behav., 3: 619-623. Howard, E. (19’73). Increased reactivity and impaired adaptability in operant behavior of adult mice given corticostcrone in infancy. J. Comp. Physiol. Psychol., 85: 21 1-220. LaBarba, R. C., and White, J. L. (1971). Litter size variations and emotional reactivity in BALB/c mice. J. Comp, Physiol. PsychoL, 75: 254-251. Levine, S., and Mullins, R. F. (1968). Hormones in infancy. In G. Newton and S. Levine (Eds.), Early Experience and Behavior. Springfield: Thomas. Pp. 169-197. Levitsky, D. A., and Barnes, R. I-I. (1970). Effects of early malnutrition on the reaction of adult rats to aversive stimuli. Nature (Lond.),225: 468-469. Levitsky, D. A., and Barnes, K. H. (1972). Nutritional and environmental interactions in the bchavioral dcvelopment of the rat: Long-term effects. Science, 176: 68-71. Lindzey, G., Manosevitz, M., and Winston, H. (1966). Social dominance in the mouse. Psychon. Sci., 5: 68-71. Manosevitz, M., and McCanne, T. R. (1973), Effects of early food deprivation on mouse behavior. J. Comp. Physiol. Psychol., 83: 3 14-323. Milkovic, K., and Milkovic, S . (1’359). Reactiveness of the pituitary-adrenal system in some laboratory mammals. Endokrinologie, 37: 301-310. Plaut, S. M. (1970). Studies of undernutrition in the young rat: Methodological considerations. Dev. Psychobiol., 3: 15’7-167. Seitz, P. I:. D. (1954). The effects of infantile experiences upon adult behavior in animal subject: I. Effects of litter size during infancy upon adult behavior in the rat. Am. J. Psychiatry, 110: 9 16-927. Simonson, M., and Chow, B. W. (1969). Maze studies on the progeny of underfed mother rats. J. Nutr., 100: 685-690. Wilson, W. J. (1968). Adaptation to the Dominance Tube. Psychon. Sci., 10; 119-120. Winick, M., and Noble, A. (1967). Cellular response with increased feeding in neonatal rats. J. Nutr., 91: 179-182.
