Crossfostering in mice selectively bred for high and ...

Scandinavian Journal of Psychology, 2005, 46, 21– 29

Crossfostering in mice selectively bred for high and low levels of openfield thigmotaxis Blackwell Publishing, Ltd.

PIA K. LEPPÄNEN and S. BÉATRICE M. EWALDS-KVIST University of Turku, Finland

Leppänen, P. K. & Ewalds-Kvist, S. B. M. (2005). Crossfostering in mice selectively bred for high and low levels of open-field thigmotaxis. Scandinavian Journal of Psychology, 46, 21–29. The main purpose of this research was to investigate whether the difference in open-field (OF) thigmotaxis between mice selectively bred for high and low levels of wall-seeking behavior originated from genetic or acquired sources. Unfostered, infostered, and crossfostered mice were compared in two experiments in which the effects of strain, sex, and fostering on ambulation, defecation, exploration, grooming, latency to move, radial latency, rearing, thigmotaxis, and urination were studied. These experiments revealed that OF thigmotaxis was unaffected by the foster condition and thus genetically determined. The selected strains of mice also diverged repeatedly with regard to exploration and rearing. The findings are in line with the previously described existence of an inverse relationship between emotionality and exploration. Key words: Thigmotaxis, crossfostering, selective breeding, HOFT and LOFT mice, open field, emotionality, exploration. Pia Leppänen, Sauvontie 3 A 1, 00280 Helsinki, Finland. Tel: +358-505015454, +358-92419768 ; E-mail: [email protected]

In animal psychology, the open-field (OF) test is one of the most frequently used methods in the study of rodent behavior. The device was originally designed for measuring the natural or spontaneous direction of behavior in rats (Hall & Ballachey, 1932), with an emphasis on emotionality (Hall, 1934a,b). Hall (1934b) defined emotionality in the OF test as “a general upset or excited condition of the animal” elicited by the strangeness or novelty of the situation. In addition to emotionality, exploration and activity are often the focus of interest in OF research (Elias, Elias & Eleftheriou, 1975; Makino, Kato & Maes, 1991). As a general feature, the OF test comprises an enclosed open area in which the animal is placed and subjected to behavioral recordings (Choleris, Thomas, Kavaliers & Prato, 2001; Ossenkopp, Sorenson & Mazmanian, 1994). Over a period of 70 years, a broad variety of OF apparatuses and procedures have been employed and more than 30 OF parameters have been recorded (for reviews, see Choleris et al., 2001; Walsh & Cummins, 1976; Weiss & Greenberg, 1998). Behavioral registering is nowadays seldom performed manually (e.g., Ewalds-Kvist, Selander & Kvist, 1999), more often automatically (e.g., Gershenfeld & Paul, 1997) or by means of using a videotape recorder (e.g., Bronikowski, Carter, Swallow, Girard, Rhodes & Garland, 2001). The use of the OF test has spread from psychology to other disciplines, such as neuroscience and psychopharmacology, where rodents’ emotional behavior is commonly the focus of interest (Choleris et al., 2001). Emotionality in rodents is traditionally tested in the OF by recording defecation and ambulation (Archer, 1973). More specifically, high levels of defecation and low levels of ambulation have been interpreted as indicators of high levels of emotionality (DeFries, Wilson & McClearn, 1970; Flint, Corley, DeFries et al., 1995; Gervais, 1976; see also Whimbey

& Denenberg, 1967). However, some researchers (e.g., Lister, 1990; Ramos & Mormède, 1998) have found the use of defecation and ambulation as indices of emotionality highly questionable. The nature of the concept emotionality is also much debated: Some researchers have considered emotionality (or fearfulness) as a unitary trait (e.g., Boissy, 1995; Broadhurst, 1975; Gray, 1979), whereas others have suggested that it is a multidimensional construct (e.g., Archer, 1973; Ohl, Toschi, Wigger, Henniger & Landgraf, 2001; Ramos & Mormède, 1998). Researchers often agree that emotionality is, at least to a certain degree, genetically determined (e.g., Boissy, 1995; Flint et al., 1995; Trullas & Skolnick, 1993), but it has also been proposed that in mice emotional reactivity may be a case of nongenetic heredity, that is, determined by a mothering style (Calatayud & Belzung, 2001; Clément, Calatayud & Belzung, 2002). OF thigmotactic or wall-seeking behavior is a commonly used index of emotionality. As early as 1932, Hall and Ballachey paid attention to the rats’ tendency to move around the walls of the OF. Thigmotaxis refers to the propensity of a rodent to stay in close contact with the OF wall because of the underlying tendency to avoid open, unknown, and potentially dangerous places (see Choleris et al., 2001). Thigmotaxis is considered to belong to the category of phylogenetically prepared fear reactions for the purpose of avoiding, for example, avian predators (Fanselow & De Oca, 1998; Grossen & Kelley, 1972; Treit & Fundytus, 1989). It is assumed that the more thigmotactic, the more emotional an animal is (Clément & Chapouthier, 1998; Valle, 1970; see also Holmes, 2001). This assumption is supported by observations showing that an aversive stimulus, such as a foot shock (Grossen & Kelley, 1972) or a bright light (Hirsijärvi & Junnila, 1986; Valle, 1970), increases thigmotaxis and a repeated

© 2005 The Scandinavian Psychological Associations/Blackwell Publishing Ltd. Published by Blackwell Publishing Ltd., 9600 Garsington Road, Oxford OX4 2DQ, UK and 350 Main Street, Malden, MA 02148, USA. ISSN 0036-5564.

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P. K. Leppänen and S. B. M. Ewalds-Kvist

(Ossenkopp et al., 1994; Williams & Russell, 1972) or prolonged (Choleris et al., 2001) exposure to the OF decreases it. Thigmotaxis has been found to be a valid index of anxiety (a commonly used synonym for emotionality in pharmacological studies; Ramos & Mormède, 1998) in mice because anxiolytic drugs have been proven to decrease (Choleris et al., 2001; Simon, Dupuis & Costentin, 1994) and anxiogenic drugs to increase (Simon et al., 1994) wall-seeking behavior. In addition, it has been shown that thigmotaxis in rats is suppressed by anxiolytic agents with a relative potency that is similar to their relative potency in the treatment of human anxiety (Treit & Fundytus, 1989). The time spent in the most protected areas of the OF arena has been validated as an index of emotionality on the basis of genetic correlations as well (van der Staay, Kerbusch & Raaijmakers, 1990). One of the most powerful tools in the study of the inheritance of behavior in animals is the selective-breeding method (Crabbe, 1986; Streng, 1974). In OF research, selection experiments have been performed on mice and rats in order to study activity or ambulation (DeFries, Gervais & Thomas, 1978; Ewalds-Kvist et al., 1999), rearing behavior (van Abeelen, van der Kroon & Bekkers, 1973), and defecation (Broadhurst, 1975). However, no selection experiments based on mouse OF thigmotactic behavior have previously been conducted. Therefore, the mice used in the present study were selectively bred for high (High Open-Field Thigmotaxis [HOFT]) and low (Low Open-Field Thigmotaxis [LOFT]) levels of OF thigmotactic or wall-seeking behavior, respectively. Thigmotaxis is known to exhibit large interstrain differences in mice (e.g., Fredericson, 1953; Gershenfeld & Paul, 1997); hence, it is plausible to assume that it is, at least to a certain degree, genetically determined. In species requiring maternal care, however, parental behavior often influences the behavior of the offspring and, consequently, the emotional behavior of fostered mice may be dependent on the strain of their host mother (Calatayud & Belzung, 2001; Le Pape & Lassalle, 1984; van Abeelen, 1980; see also Russell, 1971). Comparison of the behavior of pups infostered by mothers from their own strain and of pups crossfostered by mothers from another strain provides a tool for separating the effects of postnatal maternal care from the hypothesized genetic effects on behavior (Le Pape & Lassalle, 1984). A comparison between infostered and unfostered (i.e., pups raised by their own mothers) mice may reveal effects of fostering per se (Le Pape & Lassalle, 1984). The crossfostering paradigm naturally does not control for prenatal maternal influences on animal behavior (Gray, 1987, p. 43; Poley & Royce, 1970). The main purpose of the present study was to investigate by means of using the crossfostering paradigm whether the difference in OF thigmotaxis between the selectively bred HOFT and LOFT mice originated primarily from innate or acquired sources. The effects of strain and fostering on the following OF parameters were also examined: ambulation, defecation, exploration, grooming, latency to move, radial latency, rearing, and urination. The importance of taking

Scand J Psychol 46 (2005)

coexisting behavioral aspects into account has been shown by the OF selection experiments indicating that in rodents the selection trait is often accompanied by other OF behaviors (Broadhurst, 1975; DeFries et al., 1978; van Abeelen et al., 1973; see also Wimer & Wimer, 1985). Furthermore, it was asked whether sex exerts an influence on the aforementioned OF parameters.

GENERAL METHOD Subjects and housing conditions The mice were selectively bred for high (HOFT) and low (LOFT) levels of OF thigmotactic behavior. The HOFT and LOFT mice were derived from a Swiss albino outbred stock originating from Malmö, Sweden. They were bred, reared, and selected in the laboratory at the Department of Psychology at Åbo Akademi University, Finland. The selection was made from animals that had been exposed to the OF apparatus for at least 2-min periods on 4 consecutive days and tested on the fifth day. The selection experiment started in 1993. All of the animals were reared on a 12-hr light/ 12-hr dark cycle in a noiseless room at a normal room temperature (approximately 20–23 °C). They were fed standard laboratory pellets, R3 (1260 kJ/100 g), from Lactamin, Sweden, as well as fresh tap water ad lib. The cage had a bedding of wood shavings, and a new cage was introduced weekly. For the comfort of the animals, a thick paper roll (about 10.5 × 4.0 cm) was inserted in each cage. The present study was ethically approved.

Apparatus The open-field apparatus consisted of a circular, flat, white, wooden arena (40 cm in diameter) with a 20-cm high wall made of flat, white, metallic plate. The field was marked with thin black lines to delineate three concentric circles. The outer circle was further divided into 12 partitions and the middle circle (24 cm in diameter) into 6 partitions. The center circle (8 cm in diameter) was not divided. That is, the floor of the arena was subdivided into a total of 19 partitions to aid the recording of the ambulation of the animal, thereby providing raw scores for computation. The center circle and the 6 partitions of the middle circle were defined as 7 inner units and the 12 partitions of the peripheral circle as outer units. The floor of the arena was sponged with a washing fluid and dried with a paper towel between successive recordings.

Procedure Within each strain, the animals were randomly assigned to three different foster conditions: (a) unfostered (i.e., the pups were fostered by their own mother), (b) infostered (i.e., the pups were fostered by an alien mother of their own strain), and (c) crossfostered (i.e., the pups were fostered by a mother from the other strain). All the cages were coded to cover the animals’ identity, and two experimenters blind-tested the mice in a windowless, artificially lit (normal office fluorescent tubes) experimental room near the breeding room. One or 2 days before the experimental procedures took place, the mice were exposed to the OF apparatus for approximately 2 min in order to decrease their reactions to a novel environment. This was considered essential, since it has been proposed that scores from the first day in the OF may not provide very meaningful information (Whimbey & Denenberg, 1967; see also Tachibana, 1982; van der Staay et al., 1990).

© 2005 The Scandinavian Psychological Associations/Blackwell Publishing Ltd.

Crossfostering and thigmotaxis


Parameters The OF parameters used were ambulation, defecation, exploration, grooming, latency to move, radial latency, rearing, the thigmotactic ratio, and urination. Ambulation denotes unprovoked motor activity in the form of spontaneous whole-body movements from one OF unit to another during a 2-min period. The mouse always started from the center unit. Each visited unit (excluding the starting unit) was registered as a score on a similarly divided map when at least two feet of the animal moved into an adjacent field unit. Thus, each mouse received its individual sum of ambulatory scores. Defecation was recorded by counting the animal’s fecal boli after a 2-min OF test. Exploration was registered by counting the number of at-leastonce-entered OF units (Lhotellier, Perez-Diaz & Cohen-Salmon, 1993), that is, the area covered by the animal during a 2-min period. The number of exploratory scores ranged from 0 to 19. Grooming was recorded by counting the number of bouts of the animal’s cleaning or body scratching in the OF for 2 min. Latency to move signifies the time in seconds until the mouse left the OF center unit. The minimum latency to move was 1 s. Radial latency denotes the time in seconds until the mouse reached its first OF outer unit. The minimum radial latency was 2 s. If the animal did not reach an outer unit during the 120-s recording, it was given the maximum value. Rearing was registered by counting the number of times the animal rose onto its hind legs with the front limbs either against the wall or freely in the air (Streng, 1974) during a 2-min recording. The thigmotactic ratio signifies the animal’s orientation towards the OF peripheral units but does not necessarily imply bodily contact with the wall. Seven center parts delineated the two inner circles and 12 units the peripheral circle (see Apparatus). The thigmotactic ratio was calculated by dividing the number of inner parts entered by the total sum of units visited by the mouse (McIlwain, Merriweather, Yuva-Paylor & Paylor, 2001; Sanberg & Ossenkopp, 1977; Valle & Bols, 1976). Hence, the smaller the ratio, the more prone the mouse was to keeping close to the OF wall. The thigmotactic ratio ranged from 0.00 to 1.00. Urination was recorded by counting the number of urinary spots voided in the OF during a 2-min period.

Statistical Analysis The method used was a three-way (Strain × Sex × Fostering) analysis of variance (General Linear Model [GLM] Univariate procedure in SPSS 9.0 for Windows). Group differences were examined by using 95% confidence intervals (CIs) for estimated marginal means.

EXPERIMENT 1 The effects of strain, sex, and fostering on OF parameters were investigated in mice subjected to three different foster

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conditions. The adoption took place when the pups were at the age of approximately 1 week. One unfostered female HOFT mouse was considered an outlier and discarded. Method Subjects. A total of 219 mice (112 females and 107 males) were used in Experiment 1, originating from the 17th generation (S17) of the HOFT (N = 104) and LOFT (N = 115) strains. Procedure. Within each strain, the animals were assigned to three different foster conditions at the age of 5 to 8 days: 76 pups were fostered by their own mother, 65 were infostered by an alien mother of their own strain, and 78 were crossfostered by a mother from the other strain (Table 1). Some of the pups in the unfostered condition had participated in a previous experiment in which maternal behavior was assessed. The pups were weaned at the age of approximately 1 month. They were then separated according to sex and grouphoused in polycarbonate cages measuring 23 × 17 × 14 cm. In order to avoid the effects of social isolation interfering with later recordings, mice without siblings were discarded (Lassalle, BulmanFleming & Wahlsten, 1991). At the age of approximately 70 days, the animals were individually housed in aluminum cages measuring 14 × 14 × 12 cm. At the age of approximately 110 days, they were weighed and tested. The mean weight of the HOFT mice was 27.6 g (SD = 4.4) and of the LOFT mice 27.0 g (SD = 3.2), t = 1.21, n.s.

Results The means and standard deviations of the OF variables by strain, sex, and fostering are presented in Table 2. The table also shows statistically significant sources of variance, along with degrees of freedom and F-values. Main effects. The HOFT and LOFT mice were significantly differentiated by six parameters: exploration, grooming, radial latency, rearing, thigmotaxis, and urination. The LOFT mice explored a larger OF area and entered more inner OF units relative to outer units, or were less thigmotactic, than the HOFT mice. The LOFT mice also groomed and reared more but exhibited a shorter radial latency and voided fewer urinary spots than the HOFT mice. The females and males were significantly differentiated by two parameters: defecation and rearing. The females defecated more but reared less than the males. With regard to fostering, main effects were found on radial latency and rearing. That is, radial latency appeared to be longer in the crossfostered mice compared to the unfostered animals (although the CIs were slightly overlapping). In addition,

Table 1. Foster conditions for the HOFT and LOFT pups HOFT (S17)

Unfostered Infostered Crossfostered

LOFT (S17)

HOFT (S22)

LOFT (S22)

n

n

n

n

n

n

n

n

19 14 21

19 17 14

20 18 20

18 16 23

10 25 19

10 14 14

20 35 15

10 30 19


24 P. K. Leppänen and S. B. M. Ewalds-Kvist


Table 2. Significant main effects and interactions of strain, sex, and fostering on open-field parameters among the HOFT (S17) and LOFT (S17) mice HOFT

LOFT

Females

Parameter Ambulation Defecation Exploration Grooming Latency to move Radial latency Rearing

Thigmotaxis Urination

Males

Females

Males

1

2

3

1

2

3

1

2

3

1

2

3

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

37.8 (18.0) 1.1 (1.5) 13.1 (3.8) 0.3 (0.6) 4.5* (4.2) 10.0* (10.4) 1.2 (1.5)

33.9 (17.0) 2.4 (1.9) 12.4 (3.9) 0.5 (1.0) 5.4 (9.0) 10.4 (11.4) 1.9 (2.2)

40.1 (24.0) 1.1 (1.7) 11.6 (4.4) 0.3 (0.8) 2.9 (4.2) 11.5 (25.3) 1.8 (3.5)

43.8 (18.3) 0.8 (1.2) 13.7 (3.6) 0.3 (0.9) 3.7 (3.3) 9.5 (8.6) 3.2 (3.4)

45.2 (19.3) 0.9 (1.5) 13.1 (3.4) 0.2 (0.4) 5.5 (13.0) 9.7 (13.9) 2.3 (2.8)

33.6 (13.2) 0.4 (0.9) 11.6 (2.9) 0.0 (0.0) 8.5† (9.4) 19.9† (14.6) 1.4 (1.5)

46.2 (22.0) 1.1 (1.5) 15.3 (3.6) 0.3 (0.7) 3.5 (3.3) 6.5 (4.2) 7.1 (5.3)

34.1 (13.4) 1.6 (1.4) 13.8 (3.4) 0.4 (1.0) 3.4 (2.6) 6.9 (4.5) 3.9 (4.8)

39.6 (15.7) 1.4 (1.3) 15.1 (2.4) 0.3 (0.7) 3.8 (4.2) 9.7 (9.2) 2.8 (3.6)

45.2 (17.5) 0.9 (1.4) 15.5 (3.7) 1.3 (2.0) 2.3 (1.5) 7.0 (6.4) 6.8 (5.5)

42.0 (15.5) 0.3 (0.6) 15.6 (4.3) 0.9 (1.2) 3.1 (2.0) 8.4 (6.1) 7.9 (6.7)

40.3 (15.8) 0.5 (1.0) 14.9 (3.3) 0.7 (1.4) 6.3‡ (5.4) 11.7‡ (7.2) 4.6 (3.5)

0.13 (0.11) 0.2 (0.4)

0.10 (0.06) 0.7 (1.6)

0.15 (0.21) 0.5 (1.2)

0.14 (0.11) 0.3 (1.2)

0.09 (0.07) 1.1 (1.5)

0.18 (0.12) 0.1 (0.3)

0.12 (0.09) 0.3 (0.7)

0.17 (0.10) 0.2 (0.5)

0.21 (0.11) 0.1 (0.2)

0.19 (0.08) 0.1 (0.3)

0.17 (0.11) 0.3 (1.5)

df

F

p

Sex Sex × Fostering Strain

1 2 1

19.2 3.5 24.1

< 0.001 0.033 < 0.001

Strain Strain × Sex Sex × Fostering

1 1 2

7.6 7.9 3.7

0.006 0.006 0.027

Strain Fostering Strain Sex Fostering Strain

1 2 1 1 2 1

4.7 3.8 42.0 5.0 4.5 14.4

0.032 0.023 < 0.001 0.026 0.013 < 0.001

Strain

1

6.0

0.016

_


Note: 1 = unfostered, 2 = infostered, 3 = crossfostered. * n = 17, † n = 13, ‡ n = 22.

0.09 (0.07) 0.8 (3.1)

Source

Crossfostering and thigmotaxis


the unfostered mice reared more than the crossfostered animals. Two-way interactions. Three two-way interactions were found: Strain × Sex for grooming and Sex × Fostering for both defecation and latency to move. With regard to grooming, an interstrain difference was found in the males. That is, the LOFT males groomed more frequently than the HOFT males, while there was no strain difference among the females. With respect to defecation, the infostered females defecated more than the unfostered females. Among the males, defecation was unaffected by fostering. With regard to latency to move, the crossfostered males exhibited greater values than the unfostered males. In contrast, the females were not influenced by their foster conditions with regard to this variable.

EXPERIMENT 2 Experiment 1 investigated the effects of strain, sex, and fostering on OF parameters in mice adopted at the age of approximately 1 week. We thought that the pups might have been too old to be influenced by differing maternal care. Therefore, the mice used in Experiment 2 were adopted within 24 hours after birth. Because of the small procedural differences (the age of testing and housing) between Experiments 1 and 2, it is not justified, however, to assess the effects of the age of adoption by comparing the results of the two studies. Method Subjects. A total of 221 mice (124 females and 97 males) were used in Experiment 2, originating from the 22nd generation (S22) of the HOFT (N = 92) and LOFT (N = 129) strains. Procedure. Within each strain, the animals were assigned to three different foster conditions within 24 hr after birth: 50 pups were unfostered, 104 were infostered, and 67 were crossfostered (Table 1). The pups were weaned at the age of approximately 1 month. They were then separated according to sex and grouphoused 1 week with their siblings. After this period, they were housed in same-sex groups comprising 4 to 5 mice from the same strain and foster condition in polycarbonate cages measuring 23 × 17 × 14 cm. At the age of approximately 140 days, the animals were tested. At this time, it was discovered that some of the males of both strains had been fighting. For this reason, one crossfostered LOFT male mouse had to be removed and was not tested.

Results The means and standard deviations of the OF variables by strain, sex, and fostering are presented in Table 3. The table also shows statistically significant sources of variance, along with degrees of freedom and F-values. Main effects. The HOFT and LOFT mice were significantly differentiated by four parameters: exploration, latency to move, rearing, and thigmotaxis. The LOFT mice explored a

25

larger OF area, exhibited a longer latency to move, reared more, and were less thigmotactic than the HOFT mice. The females and males were significantly differentiated by five parameters: ambulation, defecation, exploration, rearing, and urination. The females defecated and urinated more but ambulated, explored, and reared less than the males. With regard to fostering, a main effect was found on ambulation: The crossfostered mice appeared to ambulate more than the infostered animals (although the CIs were slightly overlapping). Two-way interactions. Four two-way interactions were found: Strain × Fostering for both ambulation and rearing, Strain × Sex for rearing, and Sex × Fostering for defecation. With respect to ambulation, the crossfostered HOFT mice ambulated more than the unfostered and infostered HOFT mice. In contrast, fostering did not affect ambulation in the LOFT mice. The Strain × Fostering interaction for rearing indicates that the unfostered LOFT mice reared more than the infostered LOFT mice. Among the HOFT mice, rearing was unaffected by fostering. With regard to the interaction between strain and sex in predicting rearing, it was found that the LOFT males reared more than the females, whereas the HOFT mice did not exhibit sex differences in this respect. With regard to defecation, the sex difference was present in the unfostered and infostered mice but not in the crossfostered animals. Three-way interaction. A significant Strain × Sex × Fostering interaction was found in predicting ambulation. To facilitate the interpretation of the finding, a two-way analysis of variance was performed separately by sex. It was revealed that the crossfostered HOFT males ambulated more than the unfostered and infostered HOFT males. In contrast, fostering did not influence ambulation in the LOFT males. The Strain × Fostering interaction was nonsignificant among the females.

DISCUSSION In the present study, we used the crossfostering paradigm to investigate whether the differences in OF thigmotactic and possible subsidiary behaviors (ambulation, defecation, exploration, grooming, latency to move, radial latency, rearing, and urination) between the selectively bred HOFT and LOFT mice originated from genetic or acquired sources. Possible sex differences were also addressed. In spite of some differences in the procedures used in Experiments 1 and 2, the following findings emerged from the data of both experiments: The HOFT mice were more thigmotactic but explored and reared less than the LOFT mice. In addition, the males reared more but defecated less than the females. Only these replicated findings are discussed. It was clearly revealed that the difference in thigmotaxis between the HOFT and LOFT mice is primarily innate in


26

HOFT

LOFT

Females

Males

Females

Males

2

3

1

2

3

1

2

3

1

2

3

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

M (SD)

Ambulation

27.3 (9.8)

28.3 (15.6)

35.0 (17.3)

37.6 (16.5)

40.7 (20.1)

64.4 (28.2)

40.3 (18.6)

35.3 (14.8)

39.1 (15.0)

48.3 (12.7)

51.2 (16.5)

43.9 (15.4)

Defecation

3.1 (1.5) 14.0 (2.6) 1.3 (1.6) 1.3 (0.7) 6.0 (6.6) 1.6 (2.1)

1.9 (1.4) 12.5 (4.9) 1.6 (1.9) 1.8 (1.4) 4.9 (3.4) 2.2 (3.3)

1.8 (1.5) 13.7 (4.1) 1.4 (1.6) 1.6 (1.6) 5.5 (9.5) 2.3 (3.2)

0.5 (0.7) 14.4 (3.4) 1.2 (2.1) 2.1 (1.4) 10.0 (13.1) 1.7 (2.9)

0.3 (0.6) 14.7 (3.8) 1.4 (1.8) 2.4 (3.3) 4.1 (3.3) 3.1 (3.3)

1.4 (2.2) 17.0 (2.6) 1.1 (1.7) 2.4 (2.0) 5.9 (6.0) 3.9 (4.5)

1.6 (1.3) 15.7 (3.2) 1.7 (2.1) 2.3 (2.2) 6.4 (6.6) 8.5 (6.4)

2.1 (1.6) 14.3 (3.5) 1.6 (2.2) 3.2 (4.8) 5.5 (5.2) 4.3 (4.9)

1.9 (1.6) 15.9 (3.3) 2.3 (1.4) 4.5 (4.8) 9.0 (8.8) 5.0 (4.2)

0.6 (1.1) 16.8 (1.6) 2.0 (2.5) 2.1 (1.4) 4.8 (5.5) 11.3 (3.9)

0.4 (0.9) 17.0 (1.9) 1.3 (1.8) 2.2 (1.6) 4.7 (2.7) 9.0 (5.5)

1.1 (1.9) 16.4 (2.4) 0.3 (0.8) 3.0 (2.0) 7.1 (4.3) 8.2 (6.7)

0.22 (0.12) 0.1 (0.3)

0.17 (0.09) 0.0 (0.2)

0.18 (0.13) 0.2 (0.4)

0.17 (0.10) 0.1 (0.3)

0.26 (0.09) 0.1 (0.3)

0.30 (0.15) 0.1 (0.3)

0.23 (0.12) 0.2 (0.4)

0.28 (0.16) 0.1 (0.4)

0.21 (0.11) 0.0 (0.0)

0.25 (0.12) 0.1 (0.3)

0.23 (0.14) 0.0 (0.0)

Parameter

Exploration Grooming Latency to move Radial latency Rearing

Thigmotaxis Urination

0.18 (0.11) 0.0 (0.0)

Note: 1 = unfostered, 2 = infostered, 3 = crossfostered.

Source

df

F

p

Sex Fostering Strain × Fostering Strain × Sex × Fostering Sex Sex × Fostering Strain Sex _

1 2 2 2 1 2 1 1

29.2 3.6 6.2 3.3 41.1 3.1 11.2 12.5

< 0.001 0.028 0.002 0.038 < 0.001 0.049 0.001 0.001

Strain

1

5.2

0.024

Strain Sex Strain × Sex Strain × Fostering Strain

1 1 1 2 1

58.3 10.5 3.9 4.0 9.4

< 0.001 0.001 0.049 0.020 0.002

Sex

1

4.3

0.040

_


1



Table 3. Significant main effects and interactions of strain, sex, and fostering on open-field parameters among the HOFT (S22) and LOFT (S22) mice

Crossfostering and thigmotaxis 27


origin and not influenced by differing maternal behavior, although it has to be remembered that the crossfostering paradigm does not control for prenatal maternal influences on animal behavior (Gray, 1987, p. 43; Poley & Royce, 1970). This finding agrees with the assumption that emotionality in rodents is genetically determined (Boissy, 1995; Flint et al., 1995; Trullas & Skolnick, 1993). In mice, support for a genetic explanation for individual differences in thigmotaxis also comes from studies showing interstrain differences (Fredericson, 1953; Gershenfeld & Paul, 1997). In addition, the positive selection effect in the HOFT and LOFT mice provides similar evidence (Gray, 1987, p. 43; Hurnik, Bailey & Jerome, 1973; Lagerspetz & Lagerspetz, 1983). Our results question the idea that differing mothering styles would provide the best explanation for individual differences in emotional reactivity of adult mice offsprings (Calatayud & Belzung, 2001; Clément et al., 2002). In the present study, thigmotaxis was not found to be affected by sex, as in a previous study of Kvist and Selander (1992) as well as of Gershenfeld and Paul (1997). On the other hand, Archer (1977) found female mice to be more thigmotactic than males, a finding which was confirmed in rats by Valle and Gorzalka (1980). In addition to thigmotaxis, the HOFT and LOFT mice also diverged with regard to exploration and rearing: The high-thigmotaxis HOFT mice explored and reared less than the low-thigmotaxis LOFT mice. This is in line with many OF studies suggesting that emotionality and exploration are inversely related (Archer, 1973; Holmes, 2001). That is, a high level of emotionality inhibits exploration and a low level facilitates it (Williams & Russell, 1972). Rearing represents one aspect of exploration (Archer, 1973; Hoover-Plow, Skomorovska-Prokvolit & Welsh, 2001; van Abeelen, 1970), given that it can assist an animal in obtaining information about its environment (Crusio, 2001). Consequently, rearing has also previously been found to be inversely related to thigmotaxis (Carrey, McFadyen & Brown, 2000; Hirsijärvi & Junnila, 1986; Rowan & Flaherty, 1991; Valle, 1970). Strain differences in OF parameters other than the one on which the selection is based have commonly been observed. DeFries et al. (1978) selected mice for high and low levels of ambulation and discovered that the strains also differed with regard to defecation. Likewise, van Abeelen et al. (1973) selected mice for high and low levels of rearing and found that the strains also differed with respect to ambulation. In addition, Broadhurst (1975) selected rats for high and low levels of defecation and discovered that his strains differed with regard to ambulation as well. On the basis of the previous findings (Choleris et al., 2001; Simon et al., 1994; Treit & Fundytus, 1989), we posit that thigmotaxis is a powerful index of emotionality or anxiety in mice and rats. However, the most traditional OF indices of emotionality are ambulation and defecation (Walsh & Cummins, 1976). Thus, it could have been expected that the HOFT and LOFT mice would also have differed in these two parameters, which turned out not to be the case. It has,

however, also previously been observed that strains differing in thigmotaxis do not always differ with regard to ambulation and defecation (Ramos, Berton, Mormède & Chaouloff, 1997; see also Ramos & Mormède, 1998). In addition, it has been found that drugs affecting the level of thigmotaxis do not alter locomotor activity or alter it in an inconsistent manner (Choleris et al., 2001; Simon et al., 1994; see also Gross, Santarelli, Brunner, Zhuang & Hen, 2000). This discrepancy may be due to the lack of validity of ambulation and defecation as indices of emotionality (Lister, 1990; Ramos & Mormède, 1998). On the other hand, it is also possible that these OF parameters may measure only different aspects of emotionality. With regard to sex differences, the males of both strains reared more than the females, which is in agreement with the findings of Mathis, Paul, and Crawley (1994). In addition, the HOFT and LOFT males defecated less than the females. In several studies, male mice have been shown to defecate more than females (e.g., Bronikowski et al., 2001; Halcomb, Hegmann & DeFries, 1975; LaBarba, Hibbs & White, 1973). However, Archer (1977) questioned typical sex differences in defecation and suggested that female mice may defecate more than males, especially during short testing periods. Furthermore, it has been proposed that the use of selectively bred strains often reverses sex differences in defecation (Gray, 1979), which agrees with the present finding. In summary, two separate experiments showed that when tested in adulthood, the thigmotactic behavior of the selectively bred HOFT and LOFT mice resembled that of their genetic strain, not that of their foster mothers’ strain. That is, the difference in OF thigmotaxis between the HOFT and LOFT mice originated from innate or genetic sources. The two types of mice also differed with regard to exploration and rearing. These findings support the existence of an inverse relationship between emotionality and exploration. The experiments were conducted in the laboratory at the Department of Psychology at Åbo Akademi University, Finland. The authors are also obliged to Niklas Ravaja, PhD, for statistical advice.

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