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Behavioral Neuroscience
2011, Vol. 125, No. 1, 29 –36
© 2011 American Psychological Association
0735-7044/11/$12.00 DOI: 10.1037/a0021952
This document is copyrighted by the American Psychological Association or one of its allied publishers.
This article is intended solely for the personal use of the individual user and is not to be disseminated broadly.
Strain-Specific Cognitive Deficits in Adult Mice Exposed to Early Life Stress
Mukti Mehta
Claudia Schmauss
New York State Psychiatric Institute
New York State Psychiatric Institute and Columbia University
Early life stress is a prominent risk factor for the development of adult psychopathology. Numerous
studies have shown that early life stress leads to persistent changes in behavioral and endocrine responses
to stress. However, despite recent findings of gene expression changes and structural abnormalities in
neurons of the forebrain neocortex, little is known about specific cognitive deficits that can result from
early life stress. Here we examined five cognitive functions in two inbred strains of mice, the stressresilient strain C57Bl/6 and the stress-susceptible strain Balb/c, which were exposed to an infant maternal
separation paradigm and raised to adulthood. Between postnatal ages P60 to P90, mice underwent a series
of tests examining five cognitive functions: Recognition memory, spatial working memory, associative
learning, shifts of attentional sets, and reversal learning. None of these functions were impaired in IMS
C57Bl/6 mice. In contrast, IMS Balb/c mice exhibited deficits in spatial working memory and extradimensional shifts of attention, that is, functions governed primarily by the medial prefrontal cortex. Thus,
like recently discovered changes in frontocortical gene expression, the emergence of specific cognitive
deficits associated with the medial prefrontal cortex is also strain-specific. These findings illustrate that
early life stress can indeed affect specific cognitive functions in adulthood, and they support the
hypothesis that the genetic background and environmental factors are critical determinants in the
development of adult cognitive deficits in subjects with a history of early life stress.
Keywords: early life stress, inbred mouse strains, recognition memory, working memory, attention
set-shifting
ment than the stress-resilient strain C57Bl/6 (Millstein & Holmes,
2007).
ELS can provoke long-lasting effects on endocrine responses to
subsequent stress (Ladd et al., 2004) and the related gene expression changes (Navailles et al., 2010). Moreover, ELS induces
alterations in dendritic morphology of pyramidal neurons of the
medial prefrontal cortex (mPFC; Pascual & Zamora-León, 2007)
that resemble those elicited by chronic corticosteroid administration (Cerqueira et al., 2007). Moreover, age-dependent changes in
the densities of calbindin- and parvalbumin-immunoreactive interneurons in subregions of the mPFC (Helmeke et al., 2008) and
altered monoaminergic innervation of the mPFC (Braun et al.,
2000) have been reported to result from ELS. Recently, strainspecific changes in fronto-cortical gene expression have also been
found in adult mice exposed to ELS. For example, Balb/c, but not
C57Bl/6, mice exhibit increased expression of G␣q (Bhansali et
al., 2007; Schmauss et al., 2010), decreased expression of the
plasticity-inducible transcription factor egr-1 (Navailles et al.,
2010), and increased expression of extensively edited mRNA
isoforms encoding 5-HT2C receptors with decreased G-proteincoupling efficiency (Bhansali et al., 2007; Schmauss et al., 2010).
These genes can play a role in forebrain neocortex-governed
cognitive functions, especially in the mPFC (working memory,
set-shifting) and the orbital fontal cortex (reversal learning): signaling through Gq-coupled receptors is necessary for working
memory (Runyan et al., 2005); induced egr-1 expression can lead
to long-lasting synaptic changes that influence behaviors associated with learning and memory (Cole et al., 1989); and 5-HT2Creceptor signaling in the orbital frontal cortex plays a role in early
phases of spatial reversal learning (Boulougouris & Robbins,
2010).
For several heritable psychiatric disorders, the interaction between gene and environment is thought to be critical for modulating outcome or mutating genetic risk (Kendler, 2005; Kendler &
Baker, 2007). This is best documented for mood disorders in
subjects with distinct genetic variants in serotonin-related genes
(Caspi et al., 2010). Moreover, the impact of environmental factors
on the development of psychopathology depends upon the age of
exposure. The most persuasive example, early life stress (ELS),
exerts profound effects on adult emotional behavior and increases
risk for depression, anxiety disorders, and substance abuse (Holmes et al., 2005).
In studies on the impact of ELS on adult psychopathology,
inbred strains of mice provide a source of natural genetic variability and phenotypic differences. For example, the isogenic strains
C57Bl/6J and Balb/cJ differ in their sensitivity to ELS and adult
stress (Holmes et al., 2005; Millstein & Holmes, 2007), with
Balb/c being more susceptible to stress-induced behavioral impair-
Mukti Mehta, Department of Molecular Therapeutics, New York State
Psychiatric Institute; Claudia Schmauss, Department of Molecular Therapeutics, New York State Psychiatric Institute and Department of Psychiatry, Columbia University.
We thank Kristin Bornello for assistance in behavioral testing. This
work was supported by National Institutes of Health Grant MH078993 to
C.S. and, in part, by a National Institutes of Health Conte Center Grant P50
MH062185.
Correspondence concerning this article should be addressed to Claudia
Schmauss, Department of Psychiatry and Molecular Therapeutics, Columbia University and New York State Psychiatric Institute, 1051 Riverside
Drive, New York, NY 10032. E-mail: cs581@columbia.edu
29
MEHTA AND SCHMAUSS
30
Despite the findings of altered neuronal morphology and gene
expression changes, the effects of ELS on frontocortical-governed
cognitive functions remain to be investigated. In the present study,
we tested the hypothesis that, like the persistent frontocortical gene
expression changes elicited by ELS exposure, ELS-induced cognitive deficits also occur in a strain-specific manner. Therefore, we
examined several cognitive functions, including recognition and
working memory, associative learning, attention-set-shifting, and
reversal learning, in C57Bl/6 and Balb/c mice with and without
exposure to ELS.
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Method
Subjects
Male and female Balb/cJ and C57Bl/6J mice were housed in a
temperature-controlled (26 ⫾ 2 °C) barrier facility with a 12-hr
light– dark schedule (lights on at 6:00 a.m.) and free access to food
and water. All experiments involving animals were performed in
accordance with the National Institutes of Health Guide for the
Care and Use of Laboratory Animals and approved by the Institutional Animal Care and Use Committees at Columbia University
and the New York State Psychiatric Institute. All efforts were
made to minimize both the number of animals and the discomfort
of the animals used.
Infant Maternal Separation (IMS)
Pups of first-time mothers were separated from their dam daily
for 3 hr (from 1:00 to 4:00 p.m.) from postnatal age Day 2 (P2)
until P15. Control animals were standard-facility-reared (SFR)
offspring of first-time mothers and were subjected to the same
housing and husbandry conditions. Only litters of 6 to 8 pups were
selected for this study. For SFR and IMS mice, a total of 10 and 12
litters, respectively, were used. Because IMS mice exhibit a delayed postnatal development as indicated, for example, by a 4-day
delay of eye opening and because they are still being nursed by
their dams at P21, all pups were weaned at P28. This ensured a
near 100% survival rate of the IMS pups, which throughout postnatal development have about a 20% reduced body weight, even as
young adults (Navailles et al., 2008). Four to five animals randomly selected from four to five different litters were grouphoused by sex. At P60, one cohort of IMS mice and their SFR
controls were selected for behavioral tests of depression- and
anxiety-like behavior (10 mice per group derived from 10 different
litters) while the other cohort was selected for cognitive–
behavioral testing (8 mice per group selected from 8 of the 10
[SFR] or 12 [IMS] litters described above). Behavioral tests of
depression and anxiety-related behaviors were conducted with
male mice. The groups of mice tested in cognitive– behavioral tests
were composed of equal numbers of male and female mice.
Elevated Plus Maze (EPM)
Mice were exposed to an EPM apparatus purchased from Stoelting (Wood Dale, IL), which had two open and two closed arms that
joined to form the center of the maze (lane width: 5 cm, arm
length: 35 cm, wall height: 15 cm, elevation above ground: 55 cm).
Testing (under 100 lux/m2 lightening) was performed between
1:00 and 4:00 p.m. Mice were placed in the center of the maze and
their time spent in open arms during a 5 min test period was
recorded by the investigators during the time of testing.
Forced Swim Test (FST)
The same mice tested in the EPM were tested in the FST 1 week
later. A modified version of the forced swim test was used as
previously described (Bhansali et al., 2007). Briefly, between 1:00
and 4:00 p.m., mice were placed into plastic buckets (23 cm deep
and 19 cm in diameter) filled with 25–28 °C water for 6 min. In
this modified version of the Posolt FST, mice alternate between
active swimming and passive floating, but climbing is not observed. Hence, the number of passive episodes and their duration
(in seconds) was monitored by the investigators during the last 4
min of the test. On the following day, mice were reexposed to the
FST protocol, and their behavior was recorded as described above.
Social Recognition
This test taxes short-term recognition memory and relies on the
ability of adult mice to recognize a juvenile conspecific and to
remember its perceptual features (olfactory, somatosensory, visual) for a period of 2 hr. Adult mice were singly housed 24 hr
before exposure of a nonfamiliar juvenile mouse. During a 5-min
exposure, the time interacting with the juvenile (following, sniffing, licking, etc.) was recorded (T1). Then, the juvenile was
removed, but placed back 2 hr later for a second 5-min test (T2).
In mice with intact recognition memory, the time of social interaction during the second exposure is consistently shorter (i.e.,
T1–T2 equals a positive number). To confirm the validity of this
test as a test of short-term memory, we also exposed additional
groups of mice to a different juvenile during the second exposure.
Attention-Set-Shifting Task (ASST)
A day after completion of the Social Recognition test, mice were
food-restricted such that they gradually (over the period of 5 to 7
days) lost 10% of their free-feeding body weight. Once mice
reached this 10% reduction in body weight, they were tested in the
ASST. The rodent ASST, developed by Birrell and Brown (2000),
uses two stimulus dimensions, odor and texture, which are appropriate for this species. The test apparatus was made of Plexiglas
that resembled the housing cage (dimensions: 32 ⫻ 27 ⫻ 15 cm).
A sliding door separated one third of the apparatus (holding box)
from the remaining two thirds (test area).
On the first day, mice were habituated to the test apparatus.
During this time, they learned to retrieve a food reward deeply
buried in unscented terra cotta pots filled with familiar bedding
media. The habituation period ended when mice retrieved the food
reward in 6 consecutive trials. On the next day, mice performed the
entire ASST in a single session. Briefly, they were first trained on
two simple discriminations (SD) between odor (scented terra cotta
pots) or texture (different digging media). Only one stimulus
dimension indicated the location of the food pellet, and both
dimensions were used in a randomized order. The next task was a
compound discrimination (CD), in which another stimulus dimension was introduced that was not a reliable predictor of food
reward, and the same positive stimulus (a particular odor or tex-
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EARLY LIFE STRESS AND ADULT COGNITIVE DEFICITS
ture) used in the initial SD still guided correct response selection.
In the following intradimensional shift phase (IDS), both relevant
and irrelevant stimuli were changed, but the relevant stimulus
dimension used in the SD and CD (odor or medium texture)
remained the same. Then, the formerly irrelevant dimension became relevant and required an extradimensional shift of attention
(EDS). Finally, the previously positive stimulus of the EDS became a negative one, and the previously irrelevant stimulus indicated food reward (reversal learning; EDS-Rev). In all test phases,
animals had to reach a criterion of six consecutive correct trials.
The number of trials to criterion is the response accuracy, and the
time between stimulus onset and response selection is referred to
as response latency. Details about the stimulus properties used in
the test are described in Glickstein et al. (2005).
Spatial Working Memory (WM)
We used a widely used delayed alternation task performed in a
T-maze as previously described (Glickstein et al., 2002). The maze
was made of 0.6 cm-thick Plexiglas. Its main alley (58 cm long)
was connected to two side arms (30 cm long). Alley and side arms
were 11 cm wide. All walls were 18.5 cm high.
One day after completion of the ASST, mice began their training
for alternate arm entries in the T-maze. The training period (lasting
on average 10 to 14 days) ended when mice reached at least 70%
correct arm entries (in 10 trials per day) on two consecutive days
with no intertrial delay (referred to as 5 s delay, i.e., the minimum
handling time). Then, mice performed the test with three intertrial
delays (15, 20, and 30 sec). Only correct arm entries resulted in
food reward, and their percentage of the total number of trials was
taken as a measure of response accuracy. We have previously shown
that, in this test, mice can hold spatial information in working memory
for up to 20 s, but exhibit chance performance (i.e., only 50% correct
arm entries) at 30 s delay (Glickstein et al., 2002).
31
Figure 1A summarizes results of the FST. There was a significant difference in the total time of immobility between SFR mice
of both strains (ANOVA, F(3, 27) ⫽ 6.61, p ⫽ .0017). Post hoc
Bonferroni Multiple Comparisons tests revealed that SFR C57Bl/6
mice exhibited significantly greater immobility than Balb/c mice
(Figure 1A). Nevertheless, the FST behavior of IMS C57Bl/6 mice
did not significantly differ from their SFR controls. IMS Balb/c
mice, however, exhibited significantly more immobility than their
SFR controls (ANOVA, F(3, 25) ⫽ 7.04, p ⫽ .0014; Figure 1A).
For the EPM test, ANOVA revealed a significant difference
between SFR mice of both strains, F(3, 33) ⫽ 9.608, p ⫽ .0001.
Post hoc Bonferrori tests revealed that SFR Balb/c mice spent
significantly less time in the open arms compared to SFR C57Bl/6
mice (Figure 1B). Moreover, there were differences between SFR
and IMS mice of both strains, F(3, 29) ⫽ 12.96, p ⬍ .00001, with
IMS mice having spent significantly less time in the open arms
compared to their SFR controls (Figure 1B).
In summary, while IMS mice of both strains exhibited similar
anxiety-like behavior in the EPM, only IMS Balb/c mice also
showed increased depression-like behavior in the FST.
Statistical Analysis
The FST, the EPM, and the Social Recognition test data were
analyzed by factorial analysis of variance (ANOVA; effect of
strain and effect of treatment [SFR or IMS]), and statistical differences were resolved post hoc using Bonferroni Multiple Comparisons tests. The data obtained from the ASST and the WM tests
were first analyzed by Repeated Measures ANOVA followed post
hoc by Tukey-Kramer Multiple Comparisons tests to assess the
performances of SFR and IMS mice of each strain in the individual
test phases (ASST) or delay periods (WM test). In addition,
factorial ANOVA was used to test for differences between strains
and treatment. For all cognitive test results, two-tailed Student’s t
tests were used to compare results between groups of males and
females. The statistical analyses were carried out using Graph Pad
InStat Version 3.0 (GraphPad Software, San Diego, CA).
Results
Emotive Behavior of Adult IMS Balb/c and
C57Bl/6 Mice
We first evaluated the impact of early life stress exposure on
adult emotive behavior by comparing the behavioral responses of
SFR and IMS Balb/c and C57Bl/6 mice to FST and EPM exposure.
Figure 1. Performance of SFR and IMS Balb/c and C57Bl/6 mice in the
FST and EPM. (A) immobility measured during the last 4 min of the
second day of FST exposure. Data are mean ⫾ SEM of measures taken
from 10 animals per group and they were compared by two-tailed Student’s
t test. (B) Total time spent in open arms during a single 5 min exposure to
the EPM. Data are mean ⫾ SEM of measures taken from the same 10
animals per group shown in A, and they were compared by two-tailed
Student’s t test. IMS ⫽ infant maternal separation. SFR ⫽ standardfacility-reared controls.
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32
MEHTA AND SCHMAUSS
Social Recognition of SFR and IMS Balb/c and
C57Bl/6 Mice
Performance of SFR and IMS Balb/c and C57Bl/6
Mice in the ASST
In the Social Recognition test, both male and female mice were
used. Two-tailed Student’s t tests revealed no differences between
males and females of either group, neither in T1 measures (Balb/
c-SFR: 131.4 ⫾ 9.5 s [males] and 118 ⫾ 14.5 s [females], p ⫽ .5;
Balb/c-IMS: 117.8 ⫾ 15.5 s [males] and 105.3 ⫾ 14.8 s [females],
p ⫽ .6; C57Bl/6-SFR: 131.9 ⫾ 7.0 s [males] and 155.1 ⫾ 11.5 s
[females], p ⫽ .1; C57Bl/6-IMS: 124.5 ⫾ 14.0 s [males] and 146.4 ⫾
20.2 s [females], p ⫽ .57), nor in the T1-T2 values for reexposure to
the same juvenile (Balb/c-SFR: 56.8 ⫾ 7.2 s [males] and 59.4 ⫾ 9.9 s
[females], p ⫽ .9; Balb/c-IMS: 52.8 ⫾ 8.4 s [males] and 59.4 ⫾ 12.5 s
[females], p ⫽ .70; C57Bl/6-SFR: 45.0 ⫾ 13.4 s [males] and 51.6 ⫾
13.5 s [females], p ⫽ .74; C57Bl/6-IMS: 56.4 ⫾ 16.9 s [males] and
46.0 ⫾ 12.8 s [females], p ⫽ .57). Thus, data from males and females
were combined in each group.
SFR and IMS mice of both strains exhibited no significant
differences in their total times of interaction with a conspecific
juvenile when they were exposed to the same juvenile during the
second exposure (ANOVA, F(7, 35) ⫽ 7.393, p ⬎ .05), that is, the
time of social interaction with the juvenile was on average 50 sec
less than during the first exposure (see Figure 2). Moreover, SFR
and IMS mice of both strains exhibited significant differences in
their interaction times when exposed to the same or a different
juvenile in the second exposure (ANOVA, F(7, 35) ⫽ 7.393; p ⬍
.0001). Post hoc Bonferroni Multiple Comparisons tests revealed
that SFR and IMS mice of both groups spent significantly more
time interacting with the novel juvenile during the second exposure. In fact, the time of interaction was indistinguishable between
first exposure (T1) and second exposure (T2) (see Figure 2).
Altogether, these results indicate unaltered short-term recognition memory of species-relevant sensory information (olfactory, somatosensory, etc.) in both strains of mice exposed to the
IMS.
The various test phases of the ASST engage three specific
cognitive processes: associative learning (CD), maintaining sets/
set-shifting (IDS, EDS), and reversal learning (EDS-Rev). For
SFR mice of both strains, Repeated Measures ANOVA revealed
significant differences in the number of trials to criterion between
test phases (C57Bl/6: F(4, 36) ⫽ 7.01; p ⫽ .0003 and Balb/c: F(4,
20) ⫽ 13.6; p ⬍ .0001). Post hoc analysis showed that both strains
required significantly more trials to complete the EDS (i.e., the
most difficult test phase) than SD, CD, or IDS phases (C57Bl/6:
p ⬍ .01 compared to SD and CD and p ⬍ .001 compared to IDS
and Balb/c: p ⬍ .001 compared to SD, CD, and IDS). Moreover,
whereas ANOVA factorial analyses revealed no significant differences in the number of trials to criterion in each test phase between
SFR mice of both strains, F(9, 68) ⫽ 4.344, p ⬎ .05, there were
strain differences between IMS mice, F(9.71) ⫽ 11.89, p ⫽ .0001.
Post hoc Bonferroni Multiple Comparisons tests resolved these
differences exclusively for the EDS phase in which IMS Balb/c
mice required significantly more trials to criterion that IMS
C57Bl/6 mice ( p ⬍ .001) (see Figure 3). Moreover, while the
performance of SFR and IMS C57Bl/6 mice was indistinguishable
(F(9, 75) ⫽ 5.68, p ⬎ .05 for all test phases), ANOVA revealed
significant differences between SFR and IMS Balb/c mice (F(9,
64) ⫽ 17.889; p ⬍ .0001). Post hoc Bonferroni Multiple Comparisons revealed significantly impaired performance of IMS Balb/c
mice in the EDS phase of the ASST ( p ⬍ .001) (see Figure 3).
Thus, IMS Balb/c, but not IMS C57Bl/6, mice exhibit a deficit in
extradimensional set-shifting.
Like in the Social Recognition test, no differences between
males and females in either group were found for their performance in the ASST, neither in the CD phase (Balb/c-SFR:
6.4 ⫾ 0.2 [males] and 6.5 ⫾ 0.3 [females], p ⫽ .8; Balb/c-IMS:
7.8 ⫾ 0.9 [males] and 6.3 ⫾ 0.3 [females], p ⫽ .15; C57Bl/6SFR: 7.3 ⫾ 0.9 [males] and 9.4 ⫾ 1.0 [females], p ⫽ .2;
Figure 2. Behavior of SFR and IMS Balb/c and C57Bl/6 mice in the Social Recognition test. The difference
in the total time of interaction between the first and second exposure is shown. For C57Bl/6 and Balb/c mice,
data are mean ⫾ SEM of measures obtained from 8 mice per group. SFR(s) and IMS(s), reexposure to the same
juvenile. SFR(d) and IMS(d), reexposure to a different juvenile. Significant differences revealed by ANOVA
were resolved post hoc using Bonferroni Multiple Comparisons tests as indicated.
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EARLY LIFE STRESS AND ADULT COGNITIVE DEFICITS
33
Figure 3. Performance of SFR and IMS Balb/c and C57Bl/6 mice in the ASST. The individual test phases are
indicated in the order they were performed. The numbers for the trials to criterion are mean ⫾ SEM of 8 animals
(4 males and 4 females) per group. Results of Repeated Measures ANOVA are summarized in the text. In this
figure, results of ANOVA factorial analyses are shown that were resolved post hoc using Bonferroni Multiple
Comparisons tests as indicated.
C57Bl/6-IMS: 7.8 ⫾ 0.6 [males] and 8.6 ⫾ 1.5 [females], p ⫽
.55), the IDS phase (Balb/c-SFR: 6.2 ⫾ 0.2 [males] and 6.8 ⫾
0.8 [females], p ⫽ .5; Balb/c-IMS: 10.0 ⫾ 1.4 [males] and 10.4 ⫾
1.7 [females], p ⫽ .8; C57Bl/6-SFR: 7.8 ⫾ 1.0 [males] and 6.8 ⫾ 0.8
[females], p ⫽ .45; C57Bl/6-IMS: 8.3 ⫾ 0.5 [males] and 7.8 ⫾ 0.8
[females], p ⫽ .6), the EDS phase (Balb/c-SFR: 9.4 ⫾ 1.2 [males]
and 11.7 ⫾ 1.8 [females], p ⫽ .32; Balb/c-IMS: 21.5 ⫾ 3.2 [males]
and 22.3 ⫾ 1.9 [females], p ⫽ .85; C57Bl/6-SFR: 7.0 ⫾ 0.7
[males] and 8.8 ⫾ 1.7 [females], p ⫽ .41; C57Bl/6-IMS: 12.5 ⫾
1.3 [males] and 11.0 ⫾ 1.5 [females], p ⫽ .71), nor the EDS-Rev
phase (Balb/c-SFR: 8.8 ⫾ 0.9 [males] and 9.0 ⫾ 1.5 [females],
p ⫽ .9; Balb/c-IMS: 8.8 ⫾ 0.8 [males] and 10.3 ⫾ 0.8 [females], p ⫽ .21; C57Bl/6-SFR: 8.8 ⫾ 1.8 [males] and 12.3 ⫾
1.0 [females], p ⫽ .2; C57Bl/6-IMS: 11.8 ⫾ 1.4 [males] and
9.7 ⫾ 0.9 [females], p ⫽ .3).
Spatial Working Memory of SFR and IMS Balb/c
and C57Bl/6 Mice
Figure 4 summarizes the performance of SFR and IMS Balb/c
and C57Bl/6 mice in the T-maze test. For SFR C57Bl/6 mice,
Repeated Measures ANOVA revealed that the percentage of correct arm entries differed significantly between delay periods (F(3,
15) ⫽ 7.18; p ⫽ .0063). Post hoc Tukey-Kramer Multiple Comparisons tests showed that the percentage of correct arm entries
was significantly higher at the 5 and 15 s ( p ⬍ .01 and p ⬍ .05,
Figure 4. Performance of SFR and IMS Balb/c and C57Bl/6 mice in a test of spatial working memory. The
percentages of correct arm entries at the various delay periods are mean ⫾ SEM of the 8 animals examined in
Figure 3. The line across the bars indicates performance by chance (50%). Results of Repeated Measures
ANOVA are summarized in the text. In this figure, results of ANOVA factorial analyses are shown that were
resolved post hoc using Bonferroni Multiple Comparisons tests as indicated.
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34
MEHTA AND SCHMAUSS
respectively) delays when compared to the 30 s delay (at which
performance by chance was reached). A similar result was obtained for SFR Balb/c mice, F(3, 21) ⫽ 5.0, p ⫽ .0093. Their
percentage of correct arm entries at the 5 s delay was significantly
higher compared to the corresponding percentages of correct arm
entries measures at 20 and 30 s delays ( p ⬍ .05).
For IMS C57Bl/6 mice, Repeated Measures ANOVA also revealed significant differences in the percentage of correct arm
entries at different delay periods (F(3, 18) ⫽ 15.87; p ⫽ .0001). In
these mice, the percentage of correct arm entries at 5 s delay was
significantly higher than the corresponding percentages at the 20
and 30 s delays ( p ⬍ .01 and p ⬍ .001, respectively), and the
percentage of correct arm entries at 15 s was higher compared to
corresponding measures obtained at 30 s delay ( p ⬍ .001). Finally,
for IMS Balb/c mice, significant differences revealed by Repeated
Measures ANOVA, F(3, 21) ⫽ 104.6, p ⫽ .0001 were resolved
post hoc for the percentages of correct arm entries at 5 and 15 s
delays, which were significantly higher than the corresponding
percentages at 20 and 30 s delays ( p ⬍ .001).
A comparison of the percentages of correct arm entries at the 5,
15, 20, and 30 s delays between SFR mice of both strains revealed
no significant differences (ANOVA, F(7, 45) ⫽ 3.85, p ⬎ .05 for
all delay periods). However, strain differences were revealed for
IMS mice (ANOVA, F(7, 52) ⫽ 27.68, p ⬍ .0001), which were
resolved post hoc for the 20 s delay period, at which IMS Balb/c
mice achieved a significantly lower percentage of correct arm
entries when compared to the corresponding percentage of IMS
C57Bl/6 mice ( p ⬍ .01) (see Figure 4).
In addition, there were significant differences between SFR and
IMS Balb/c mice (ANOVA, F(7, 49) ⫽ 19.301, p ⬍ .0001). Post
hoc analysis revealed significantly lower percentages of correct
arm entries for IMS mice at 20 and 30 s delays ( p ⬍ .01) (see
Figure 4). In contrast, there were no significant differences between SFR and IMS C57Bl/6 mice (F(7, 48) ⫽ 5.211, p ⬎ .05 for
all delay periods).
Finally, for the WM test, no differences were found between
males and females, neither at the 5 s delay (Balb/c-SFR: 77.5 ⫾
1.4 [males] and 78.8 ⫾ 5.5 [females], p ⫽ .83; Balb/c-IMS: 73.8 ⫾
2.4 [males] and 77.5 ⫾ 2.5 [females], p ⫽ .32; C57Bl/6-SFR:
77.1 ⫾ 2.6 [males] and 75.0 ⫾ 3.2 [females], p ⫽ .61; C57Bl/6IMS: 72.5 ⫾ 0.6 [males] and 73.3 ⫾ 3.3 [females], p ⫽ .85), the
15 s delay (Balb/c-SFR: 68.3 ⫾ 4.4 [males] and 65.0 ⫾ 3.1
[females], p ⫽ .88; Balb/c-IMS: 75.0 ⫾ 3.5 [males] and 72.5 ⫾ 2.5
[females], p ⫽ .80; C57Bl/6-SFR: 67.5 ⫾ 4.9 [males] and 71.0 ⫾
3.9 [females], p ⫽ .68; C57Bl/6-IMS: 65.0 ⫾ 2.0 [males] and
70.0 ⫾ 2.9 [females], p ⫽ .85), the 20 s delay (Balb/c-SFR: 55.0 ⫾
2.9 [males] and 63.8 ⫾ 7.4 [females], p ⫽ .32; Balb/c-IMS: 42.5 ⫾
4.7 [males] and 46.3 ⫾ 1.3 [females], p ⫽ .48; C57Bl/6-SFR:
56.7 ⫾ 4.6 [males] and 62.5 ⫾ 5.2 [females], p ⫽ .43; C57Bl/6IMS: 57.5 ⫾ 1.4 [males] and 58.3 ⫾ 1.7 [females], p ⫽ .72), nor
the 30 s delay (Balb/c-SFR: 61.7 ⫾ 8.8 [males] and 58.3 ⫾ 3.3
[females], p ⫽ .74; Balb/c-IMS: 48.8 ⫾ 1.3 [males] and 47.5 ⫾ 3.2
[females], p ⫽ .73; C57Bl/6-SFR: 52.5 ⫾ 7.5 [males] and 42.0 ⫾
7.8 [females], p ⫽ .43; C57Bl/6-IMS: 46.2 ⫾ 5.9 [males] and
45.0 ⫾ 8.7 [females], p ⫽ .91).
In summary, while there were no significant differences between SFR mice of both strains at either delay period, the performance of IMS Balb/c mice differed significantly from IMS
C57Bl/6 mice at the 20 s delay ( p ⬍ .01) and from SFR Balb/c
mice at 20 and 30 s delays. Hence, compared to SFR and IMS
C57Bl/6 mice and SFR Balb/c mice, IMS Balb/c mice exhibited
spatial working memory deficits.
Discussion
The present study uncovered two specific cognitive functions,
spatial WM and extradimensional shifts of attention, that were
impaired in adult mice exposed to early life stress. Other functions,
such as short-term memory, associative learning, and reversal
learning, were unaffected. Moreover, these ELS-induced deficits
were detected in the stress-susceptible strain Balb/c, but not in the
stress-resilient strain C57Bl/6. Thus, just as a number of persistent
forebrain neocortical gene expression changes occur in this strainspecific manner (Bhansali et al., 2007; Navailles et al., 2010;
Schmauss et al., 2010), cognitive deficits resulting from ELS are
also strain-specific.
The finding that both WM and extradimensional set-shifting
were affected in IMS Balb/c mice raises the question of a possibility of a common origin of their disruption. At the anatomic
levels, both functions are primarily associated with the mPFC.
Specifically, both functions are sensitive to lesions of the infralimbic and prelimbic subregions of the rodent mPFC (Aggleton et
al., 1995; Birrel & Brown, 2000). Moreover, after exposure to
maternal separation and later social isolation, the same anatomic
subregions experience changes in monaminergic innervation,
namely increased serotonergic innervation of the infralimbic and
decreased dopaminergic innervation of the prelimbic subregions
(Braun et al., 2000).
We also found that impaired performance in the EDS did not
lead to impaired performance in the EDS-Rev. A similar separation of impairment of these two functions has previously been
described for dopamine D2-receptor knockout mice that, despite
normal performance in the EDS phase of the ASST, exhibited
impaired performance in the EDS-Rev (DeSteno & Schmauss,
2009). Furthermore, it has been shown that these two cognitive
domains of attentional control functions also segregate at the
anatomic levels, that is, in rodents, the EDS-Rev performance is
sensitive to lesions of the orbital frontal cortex (McAlone &
Brown, 2004).
Finally, recognition memory, a function governed by the
perirhinal cortex and the hippocampus (Brown & Aggleton, 2001),
was also unaffected in IMS Balb/c mice. Hence, the present
finding suggests that cognitive functions governed primarily by the
infra- and prelimbic subregions of the mPFC are especially sensitive to early life stress exposure.
It is noteworthy that, despite unaffected cognitive functioning in
IMS C57Bl/6 mice, their adult emotive phenotype was affected by
IMS exposure. In fact, just like IMS Balb/c mice, IMS C57Bl/6
mice exhibited increased anxiety-like behavior in the EPM. However, in contrast to IMS Balb/c mice that also exhibited depressionlike behavior in the FST, the behavior of IMS C57Bl/6 mice in this
test did not significantly differ from their SFR controls. It is
possible that increased depression-like behavior further contributes
to the cognitive deficits detected in IMS Balb/c mice. However,
SFR C57Bl/6 mice exhibited significantly more immobility than
SFR Balb/c mice during FST exposure. This difference between
strains, and in particular the higher level of immobility of C57Bl/6
mice, could have obscured interpretation of the FST results de-
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This article is intended solely for the personal use of the individual user and is not to be disseminated broadly.
EARLY LIFE STRESS AND ADULT COGNITIVE DEFICITS
scribed—it is possible that the time SFR C57Bl/6 mice spent in
immobility had already reached a ceiling effect.
It is important to note that we found no evidence that the
cognitive deficits detected in IMS Balb/c mice differ between
males and females. This is in contrast to sex differences found for
rodents in certain tests of fear- and anxiety-related behaviors (see,
e.g., Wigger & Neumann, 1999) that may be linked (at least in
part) to different stress sensitivity of females at different stages of
the estrus cycle (Romeo et al., 2003; a finding that motivated us to
delimit our study of the FST and EPM behavior to male mice).
What causes the specific cognitive deficits in mice exposed to
early life stress? It is currently believed that there are two main
contributors to the strain-specific appearance of adult psychopathology after ELS exposure: (1) differences in maternal care and
(2) a genetic susceptibility. The differences in maternal care between C57Bl/6 and Balb/c mice at baseline are well documented:
C57Bl/6 dams exhibit significantly more arched-back nursing and
licking and grooming. They also spend significantly less time “off
the nest” compared to Balb/c dams (Millstein & Holmes, 2007;
Priebe et al., 2005). However, a daily 3-h IMS also alters the
maternal behavior, and dams of both strains have been shown to
spend an increased amount of time tending their pups immediately
upon being reunited. In fact, the strains with lowest levels of
maternal care at baseline, including Balb/c mice, exhibited the
greatest responses to IMS in terms of increased time spent “on the
nest” (Millstein & Holmes, 2007). Thus, it is not very likely that
the cognitive deficits detected in IMS Balb/c mice are solely or
predominantly routed in the maternal behavior they experience
during the IMS. Thus, it will now be important to investigate the
extent to which a genetic susceptibility to stress determines
whether subjects exposed to ELS will develop cognitive deficits.
In this regard, it is relevant to note that, in addition to anatomic
evidence for altered neuronal structure and monaminergic innervation within the mPFC, recent studies also found that IMS Balb/c
mice exhibit changes in the expression of specific genes in the
frontal cortex that are not found in IMS C57Bl/6 mice. Among
them is the G protein alpha q subunit, which is expressed at
increased levels in IMS Balb/c mice (Bhansali et al., 2007;
Schmauss et al., 2010). Interestingly, signaling through Gqcoupled receptors was found to be necessary for working memory
(Runyan et al., 2005), and stress-induced working memory impairments were shown to result from overactivation of PKC (Birnbaum et al., 2004), which can be activated by Gq-induced calcium
increase. Thus, future studies on mechanisms underlying changes
in the expression of this gene in IMS Balb/c mice could provide
important information about the role of genetic or epigenetic
variations that lead to altered G␣q expression following ELS in
one but not the other strain of mice. Moreover, it is important to
extend such studies to other genes implicated in the control of WM
and attention.
Finally, the two cognitive deficits found in the stress-susceptible
Balb/c mouse also serve as endophenotypes for a variety of psychiatric disorders (Erlenmeyer-Kimling et al., 2000; Marazziti et
al., 2010; Park & Holzman, 1992). This is of special interest as the
detection and treatment of these cognitive deficits could permit
early identification and possibly onset-prevention of the deficit’s
associated psychiatric disorders. As such, our present findings
underscore the need for a detailed assessment of the cognitive
deficits of subjects exposed to early life stress, regardless of
35
whether they already have developed clinically manifest psychopathology.
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Received August 9, 2010
Revision received September 24, 2010
Accepted October 11, 2010 䡲
Correction to Teufel et al. (2010)
In the article “On the Relationship Between Lateralized Brain Function and Orienting Asymmetries,” by Christoph Teufel, Asif A. Ghazanfar, and Julia Fischer (Behavioral Neuroscience, 2010,
Vol. 124, No. 4, pp. 437– 445), we wrote that “the likelihood of obtaining at least one significant
result at p ⬍ 0.05 is 3.125%*3 ⫽ 9.4%” (p. 443, “A Note on Statistics”). This is incorrect.
Considering the null hypothesis of equal left and right orienting responses, the (one-tailed)
probability of observing a left (or right) bias in five out of five subjects is 0.03125. With three
(independent) hypotheses, the probability of not obtaining any significant result is (1 ⫺ 0.03125)3 ⫽
0.968753 ⫽ 0.90915, and the probability of obtaining at least one significant result is 1 ⫺
0.968753 ⫽ 0.09085. More generally, the likelihood of obtaining no significant result in a single test
is 1 ⫺ ␣, and with n hypotheses, it is (1 ⫺ ␣)n. The probability of obtaining at least one significant
result is therefore 1 ⫺ (1⫺ ␣)n. We thank Mark Baxter and Jean-Paul Fischer for drawing our
attention to the error. —Christoph Teufel, Asif A. Ghazanfar, and Julia Fischer
DOI: 10.1037/a0022814
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