Download Magnitude of the Object Recognition Deficit

Behavioral Neuroscience
2009, Vol. 123, No. 1, 115–124
© 2009 American Psychological Association
0735-7044/09/$12.00 DOI: 10.1037/a0013829
Magnitude of the Object Recognition Deficit Associated With Perirhinal
Cortex Damage in Rats: Effects of Varying the Lesion Extent and the
Duration of the Sample Period
M. M. Albasser, M. Davies, J. E. Futter, and J. P. Aggleton
Cardiff University
The present study examines 2 factors that might moderate the object-recognition deficit seen after
perirhinal cortex damage. Object recognition by normal rats was improved by extending (from 4 to 8
min) the sample period during which an object was first explored. Furthermore, there was a significant
positive correlation between time spent in close exploration of the sample object and degree of successful
novelty discrimination. In contrast, rats with perirhinal cortex lesions failed to benefit from increased
close exploration and did not discriminate the novel object after even the longest sample period.
Nevertheless, the lesions did not disrupt habituation across repeated exposure to the same object. The
second factor was extent of perirhinal cortex damage. A significant correlation was found between total
perirhinal cortex loss and degree of recognition impairment. Within the perirhinal cortex, only damage
to the caudal perirhinal cortex correlated significantly with recognition memory deficits. This study
highlights the critical importance of the perirhinal cortex within the temporal lobe for recognition
memory and shows that the lesion-induced deficit occurs despite seemingly normal levels of close object
exploration.
Keywords: temporal lobe, recognition, rat, perirhinal cortex, memory
more, studies have used different terminologies for the perirhinal
region (e.g., Burwell, 2001; Swanson, 1992), allied to quite major
differences in the placement of the perirhinal border with its
adjacent cortical regions (e.g., Burwell, 2001; Burwell & Amaral,
1998, compared with Shi & Cassell, 1997, 1999). Related concerns
include the possible contribution from damage to adjacent cortical
areas (Nemanic, Alvarado, & Bachevalier, 2004). For these reasons the present study sought to relate the degree of perirhinal
cortex damage with object recognition. This analysis included
additional comparisons for tissue loss in the rostral, mid, and
caudal perirhinal cortices, as these regions have different connectional properties (Furtak, Wei, Agster, & Burwell, 2007).
A second potentially important variable is the length of time the
rat has with the sample object, prior to discriminating that object
from a novel alternative. Apart from the first lesion study to show
the importance of the rat perirhinal cortex for object recognition
(Mumby & Pinel, 1994), almost all studies have used spontaneous
tests of object recognition (Aggleton et al., 1997; Barker et al.,
2001, 2006; Bussey, Muir, & Aggleton, 1999; Davies, Machin,
Sanderson, Pearce, & Aggleton, 2006; Ennaceur, Neave, & Aggleton, 1996; Mumby, Glenn, Nesbitt, & Kyriazis, 2002; Mumby,
Piterkin, Lecluse, & Lehman, 2007; Norman & Eacott, 2004,
2005; Winters & Bussey, 2005; Winters, Forwood, Cowell, Saksida, & Bussey, 2004). These studies all comprise two phases
(Ennaceur & Delacour, 1988). First, the rat is exposed to a novel
object in a test arena and permitted to explore that object freely.
This sample phase is followed by a test phase in which the rat is
put back in the arena after a specified delay, but the arena now
contains a novel object along with an identical copy of the sample
object. Normal rats spend more time exploring the novel object,
and this difference provides a measure of novelty discrimination.
The perirhinal cortex (areas 35 and 36) is thought to be necessary for normal recognition memory. While the first evidence
came from electrophysiological (Brown & Aggleton, 2001;
Brown, Wilson, & Riches, 1987) and lesion (Meunier, Bachevalier, Mishkin, & Murray, 1993; Murray & Mishkin, 1986; ZolaMorgan, Squire, & Amaral, 1989) studies of recognition by monkeys, subsequent studies of the rat have repeatedly shown that the
perirhinal cortex appears to fulfill a very similar role (Aggleton,
Keen, Warburton, & Bussey, 1997; Mumby & Pinel, 1994; Zhu,
Brown, & Aggleton, 1995, Zhu, Brown, McCabe, & Aggleton,
1995, Zhu, McCabe, Aggleton, & Brown, 1996). As a consequence, information about the ways in which the rat perirhinal
cortex supports recognition memory is likely to generalize across
species. The present study addresses two variables (lesion extent
and stimulus sample time) likely to alter the severity of the
object-recognition deficit typically seen after perirhinal cortex
loss. Understanding the impact of these variables will help to
explain inconsistencies across studies and inform theories concerning the precise role of the perirhinal cortex in recognition memory.
The first variable is the extent of perirhinal damage. Comparing
this variable across studies is, however, difficult given that quantitative pathological information is typically not provided. Further-
M. M. Albasser, M. Davies, J. E. Futter, and J. P. Aggleton, School of
Psychology, Cardiff University, Cardiff, Wales, United Kingdom.
We thank Seralynne Vann, Janice Muir, and David Bilkey for their
assistance. The research was supported by the Wellcome Trust.
Correspondence concerning this article should be addressed to J. P.
Aggleton, School of Psychology, Cardiff University, 70 Park Place,
Cardiff, Wales, CF10 3AT, United Kingdom. E-mail: aggleton@
cardiff.ac.uk
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ALBASSER, DAVIES, FUTTER, AND AGGLETON
116
It is likely that the duration of time spent initially exploring the
sample object will affect the discrimination of novelty, though this
has rarely been examined. Some studies use a fixed sample period
from 3 to 5 min (Aggleton et al., 1997; Ennaceur et al., 1996;
Mumby et al., 2002), while others require the rats to explore the
objects actively for a preset period that is typically from 25 to 30 s
(Davies et al., 2006; Norman & Eacott, 2004, 2005; Winters &
Bussey, 2005; Winters et al., 2004). These latter studies also use a
maximum sample session duration if the rat does not reach the
preset sample time (typically 4 or 5 min). The only study that
appears to have examined whether the perirhinal lesion recognition
deficit can be partially overcome by increasing the exploration of
the sample object (Mumby et al., 2007) found that when perirhinal
cortex lesioned rats are given five sample periods with the same
object, each of 5 min, they are able to show relatively good novelty
discrimination after 24 hr retention but not after 3 weeks. With a
single 5-min sample period, the same rats showed the expected
perirhinal lesion deficit for the same 24-hr retention period
(Mumby et al., 2007).
The present study sought to examine the robustness of this
perirhinal sparing associated with increased sample exploration
(Mumby et al., 2007). This effect is of interest as it is predicted by
theories of perirhinal function that assume that this region supports
object identification, a process likely to be aided by extended
experience (Buckley, 2005; Eacott & Gaffan, 2005). This effect
was examined by increasing the length of single sample periods,
rather than by increasing the number of sample periods. Consequently, rats were tested on spontaneous object recognition using
three different sample durations (4, 6, and 8 min). In all cases the
retention delay was 24 hr, chosen because it is consistently sensitive to perirhinal damage after a single sample session (Mumby et
al., 2007; Norman & Eacott, 2004; Winters et al., 2004). A related
issue is whether rats with perirhinal lesions show abnormal rates of
habituation of exploration to a repeatedly presented object. This
process was also examined as abnormalities in rates of habituation
could confound the validity of the spontaneous recognition test.
Materials and Method
Subjects
Forty male rats of the pigmented DA (Dark Agouti) strain
(Bantin and Kingman, Hull) were used in this study. All subjects
were housed in pairs under diurnal conditions (14 hr light and 10
hr dark), and food and water were provided ad lib during testing.
At the start of testing the animals were 4 months of age and
weighed 220 –250 g. All experiments were performed in accordance with the U.K. Animals (Scientific Procedures) Act (1986)
and associated guidelines, thereby complying with American Psychological Association ethical standards for the treatment and care
of animals.
Surgical Procedures
Animals were deeply anesthetized by intraperitoneal injection
(60 mg/kg) of sodium pentobarbital (Sagatal, Rhone Merieux).
The 20 rats receiving perirhinal cortex lesions (Perirhinal) were
then each placed in a stereotaxic headholder (David Kopf Instruments, Tujunga, CA) with the nose bar at ⫹5.0. A sagittal incision
was made along the scalp and the temporal muscles retracted. An
area of skull was then removed over the parietal cortex in each
hemisphere, approximately 4 –7 mm posterior to bregma. The
perirhinal cortex lesions were made by injecting a solution of 0.09
M N-methyl-D-aspartic acid (NMDA; Sigma Chemical Company,
Ltd., U.K.) dissolved in phosphate buffer (pH 7.2) in three sites per
hemisphere using a 1-␮l Hamilton syringe (Bonaduz, Switzerland). The stereotaxic coordinates of the lesion placements relative
to ear-bar zero were (AP) 4.0, lateral (L) ⫾ 5.7; AP 2.5, L ⫾ 6.1;
and AP 0.9, L ⫾ 6.2. The depth (in millimeters), from bregma at
the three sites was ⫺9.2 (most rostral), ⫺9.5, and ⫺8.9 (most
caudal). Bilateral injections of 0.20 ␮l were made for all three
sites. The 20 animals acting as surgical controls (sham) received
the same procedure and drugs as did the animals receiving lesions.
This involved the removal of a bone flap and the needle being
lowered but without the injection of NMDA. At the completion of
all surgeries the skin was sutured and an antibiotic powder (Acramide, Dales Pharmaceuticals, Skipton, U.K.) applied. All rats
also received a 5-ml subcutaneous injection of glucose saline.
Apparatus
All testing was conducted in a square arena that had a wooden
floor 1 m by 1 m. The floor was covered by sawdust that was
agitated between trials and regularly replaced. The solid walls
were 43 cm high and painted matte gray. An overhead camera was
used to record animals’ behavior for subsequent analysis.
The stimuli consisted of triplicate copies of objects made of
glass or plastic that varied in shape, color, and size (9 ⫻ 8 ⫻ 5 cm
to 17 ⫻ 13 ⫻ 5 cm), and were too heavy for the animal to displace.
Pairs of test objects were placed in the arena 23 cm from the
corner, and hence 54 cm apart. Objects used included cans, bottles,
tins, glasses, and pots. The objects differed markedly and did not
appear to share many common features (see Bartko, Winters,
Cowell, Saksida, & Bussey, 2007). Every test session was videorecorded. The particular pairs of objects used were counterbalanced across the three sample conditions. Time spent exploring
each object was defined as having the rat’s head within 1 cm of the
object. This measure was recorded automatically using Ethovision
(Noldus, The Netherlands) to ensure consistency.
Behavioral Testing
Pretraining. Prior to the start of testing, animals received two
habituation sessions. During the first session, 2 rats were placed in
the empty arena for a period of 5 min. During the second habituation period, rats were placed individually into the arena.
Object recognition. Testing for the three different sample
times (4, 6, and 8 min) was counterbalanced in sequence. Testing
was also counterbalanced with a parallel series of experiments that
used a Y-shaped maze (after Winters et al., 2004) to assess object
recognition (results not reported). Animals were taken into the
experimental room and tested individually. During these sample
periods the rats were allowed to explore two identical copies of the
sample object. The total time spent exploring the two identical
objects was recorded for all trials, and no criterion was set for a
minimum amount of exploration during this period. A different
pair of identical objects was used for each sample phase. The
actual objects used and whether any given object was used as the
sample (familiar) were counterbalanced.
PERIRHINAL CORTEX AND OBJECT RECOGNITION
After a delay of 24 hr, which was spent in the home cage with
a cage mate, each rat was returned to the arena, which now
contained a novel object and an identical copy of the object
previously used during the familiarization phase. Again, these
objects were placed equidistant from the sides of the arena wall
and the placement of both the novel and familiar objects (left or
right) was counterbalanced between animals. Each rat was tested
once at each of the three sample durations. The test period lasted
for 5 min, and times spent exploring both novel and familiar
objects were recorded. For every rat there was a gap of 3 days after
the test session before the next trial.
Object habituation. After completion of object recognition,
we tested all rats for their rates of habituation to the same, repeated
object (a golden globe 7 cm in diameter). For this test, four
identical globes were placed 7 inches from each corner of the
square arena. The arena was the same as that used for object
recognition, but it was now surrounded by a circular black curtain,
so limiting extramaze cues. The total amount of exploration of the
objects was measured over four successive 5-min sessions. Each
session was 3 days apart, with the same objects for all four
sessions. Exploration was measured in the same way as for object
recognition.
Histology. All animals were anesthetized with sodium pentobarbital (1 mg/kg) and perfused intracardially with a 0.9% saline
solution, followed by a 5% formal saline solution. Their brains
were removed and postfixed in a 5% formal saline solution; prior
to cutting, they were transferred to a 25% sucrose solution overnight. The brains were placed on a freezing microtome and 40-␮m
coronal sections were cut. Every third section was kept and
mounted onto gelatin-coated slides. The sections were then stained
with cresyl violet, a Nissl stain.
Volumetric analyses. Estimates were made of the extent of the
lesions in all 20 perirhinal animals. Coronal sections were viewed
on a Leica DMRB microscope, photographed using an Olympus
DP70 camera and the images transferred to a computer. Lesion
measurements were carried out using the program analySISD̂
(Soft-Imaging Systems). A set of three bilateral, standard coronal
sections (rostral, mid, and caudal) were first constructed according
to the cytoarchitectonic divisions of Burwell (2001). Lesion borders were then drawn in a frame area including the perirhinal
cortex (areas 35 and 36), area TE, and the piriform cortex or the
entorhinal cortex (depending on AP level).
The perirhinal cortex was subdivided into three subregions:
rostral (posterior to AP ⫺2.80 in relation to bregma; Paxinos &
Watson, 1997), mid (posterior to AP ⫺3.80), and caudal (posterior
to AP – 4.80). It should be noted that our rostral perirhinal measurements would include much of the caudal parietal insular cortex
as described by Shi and Cassell (1999), who proposed a much
more restricted perirhinal region than did Burwell (2001). For all
lesion areas analyzed, measurements were taken from four consecutive sections from each hemisphere immediately caudal to
each of these AP levels. Consequently, the extent of the lesion was
mapped out on 12 coronal sections along the anterior–posterior
(AP) extent of the perirhinal cortex. The tissue loss from all 24
hemispheres was summed to produce a total lesion size.
Analysis of behavior. Measurements were taken of the total
exploration time for all the identical objects in the sample phase,
as well as the time spent exploring the individual objects in the test
phase. Recognition is typically assessed from two measures. D1
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corresponds to the total time (in seconds) exploring the novel
object minus the total time exploring the familiar object (i.e., the
sample object). The discrimination ratio, D2 (Ennaceur & Delacour, 1988) is the difference in time spent exploring the novel and
familiar objects divided by the total time spent exploring objects in
the test phase (i.e., D1 divided by total exploration).
These two measures of discrimination were calculated across
the entire test phase (5 min) and after the first 2 min. The latter
interval was included in light of previous evidence (Dix & Aggleton, 1999) that it may provide the most sensitive measure of
discrimination. Only the results for the D1 scores are presented, as
it was found that some rats from both groups showed unusually
low levels of exploration. A consequence was the generation of
extreme D2 scores (both positive and negative). The occasional
presence of these very high or very low scores led to increased
variance and so increased the likelihood of null results in an
experimental design that required multiple individual comparisons
across different conditions. While the profile of results for D2
mirrored those for D1, they are not reported.
The behavioral data for D1 (2 min and 5 min) were analyzed in
separate analyses of variance. When appropriate, the simple effects
for each brain region were analyzed as recommended by Winer
(1971). The probability level of 0.05 was taken as being statistically significant. In order to see whether the animals were performing above chance, one-sample t tests (two-tailed) were conducted. Associations between sample time and performance were
examined using Pearson correlations. Correlating extent of perirhinal and extraperirhinal damage with recognition performance has
the challenge that the variables that one wishes to compare are not
themselves independent. Indeed, the cause of the perirhinal lesion
is the same cause of the extraperirhinal damage, and so it is to be
expected that the extent of perirhinal damage will correlate positively with extent of extraperirhinal damage (as was the case; see
the Results section). For this reason we used partial regressions to
examine the relationships between brain damage and performance.
Results
After first describing the location and extent of the perirhinal
cortex lesions, the Results section considers whether increased
sampling affected the extent of novelty discrimination by both
the perirhinal lesioned rats and their controls, and then considers the relationships between amount of cortex loss and recognition performance.
Histological Findings
The extent and borders of the perirhinal and postrhinal cortices
were taken from Burwell (2001). All 20 rats suffered bilateral loss
of the perirhinal cortex, though there was some variability. Five
animals were, however, excluded from the perirhinal lesion group
on the basis of the location, extent, and symmetry of their lesions
(“excluded cases”).
All rats in the perirhinal lesion group (n ⫽ 15; Figure 1A) had
lesions centered in the perirhinal cortex that produced substantial
bilateral damage to both areas 35 and 36. The measured damage
involved between 32.9% and 79.2% of the total perirhinal cortex.
It should be noted that these percentages are almost certainly
underestimates of functional damage, as in all cases some of the
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ALBASSER, DAVIES, FUTTER, AND AGGLETON
Did Perirhinal Cortex Lesions Affect the Total Amount of
Active Object Sampling?
The first analyses considered the total times spent exploring the
sample objects for the 4-, 6-, and 8-min sample conditions (see
Figure 2). As was expected, there was a highly significant increase
in object exploration as the sample period was made longer, F(2,
66) ⫽ 12.3, p ⬍ .0001. The perirhinal group did not, however,
differ from the controls in the total amount of active sampling, F(1,
33) ⫽ 2.17, p ⫽ .15, nor was there an interaction between surgery
and sampling (F ⬍ 1; see Figure 2).
Did the Duration of Active Sampling Predict the Extent of
Novelty Discrimination?
Figure 1. Diagrammatic reconstructions of the perirhinal lesions showing the
cases with the largest (gray) and smallest (black) lesions. The numbers refer to the
distance (in millimeters) from bregma according to the atlas of Paxinos and
Watson (1997). (A) Perirhinal lesion group (15 animals); (B) excluded cases (5
animals). Adapted from The Rat Brain in Stereotaxic Coordinates (3rd ed.),
G. Paxinos and C. Watson, 1997. Copyright 1997, with permission from Elsevier.
perirhinal damage was confined to either superficial or deep lamina, that is, the “spared” cortex was potentially disconnected. Some
extraperirhinal damage occurred in all cases. In most animals there
was some cortical thinning in area TE immediately above area 36,
while the lesions often extended ventrally to include immediately
adjacent parts of the piriform cortex and lateral entorhinal cortex
(depending on AP). Most lesions stopped caudally just before the
border with the postrhinal cortex. As a consequence, bilateral cell
loss in the postrhinal cortex was seen only in three cases. This cell
loss was confined to the very rostral limit of the postrhinal cortex.
Six of the 15 rats showed complete sparing of the postrhinal
cortex, while a final 6 cases had unilateral damage confined to the
extreme rostral limit of the postrhinal cortex.
The lesions in the 5 excluded cases were often asymmetric and
spared appreciable parts of the rostral (n ⫽ 4) or caudal (n ⫽ 1)
perirhinal cortex (Figure 1B). The exclusion of these 5 cases was
made prior to the formal volumetric analyses, and no rats were
reassigned on the basis of this process. These 5 rats did, however,
represent part of a continuum of perirhinal damage with the remaining
15 cases, because the total perirhinal cortex damage in the excluded
group was between 31.1% and 45.2%. For this reason, the correlations of recognition performance against extent of tissue damage
used all 20 cases. In contrast, all of the group comparisons with the
sham group just used the 15 cases with acceptable lesions.
For the sham animals, a clear, positive relationship (see Figure 3)
emerged between the length of time spent exploring the sample
objects and the subsequent degree of novelty discrimination. Correlations (Pearson r) between the total amount of sample exploration (combined across all the three time conditions) and the extent
to which the novel object was discriminated (D1 from all 5 min
combined across the three time conditions) showed a highly significant positive correlation: Pearson r ⫽ .674, p ⫽ .001 (see
Figure 3). Unlike the sham animals, the perirhinal group (n ⫽ 15)
showed no relationship between sample time and recognition in
the square arena (r ⫽ ⫺0.188, p ⫽ .50). The same pattern of
results was found for the D1 measures taken after 2 min of each
test session as the correlation for the sham animals was again
significant (r ⫽ .443, p ⫽ .05), but this was not found for the
perirhinal group (r ⫽ .290, p ⫽ .29).
Did the Length of the Sample Period (4, 6, or 8 min)
Alter the Perirhinal Lesion Effect?
Figure 4A shows the degree of object-recognition performance
(D1, 5 min) for the three different sample conditions. As was
expected, the perirhinal group was impaired in relation to the sham
Figure 2. Bar charts showing the mean sample exploration times (plus or
minus the standard error of the mean) of the sham and perirhinal groups
during the three different sample-phase conditions (4, 6, and 8 min).
PERIRHINAL CORTEX AND OBJECT RECOGNITION
119
Did Lesion Placement and Extent Correlate With the
Ability to Discriminate Novel From Familiar Objects?
A second goal was to determine whether the extent of perirhinal
damage (n ⫽ 20) predicted the degree of any recognition deficit.
Partial regressions were used to examine these relationships in two
stages. First, the extent of damage in the entire perirhinal cortex
was considered, along with the total damage dorsal to perirhinal
cortex (area TE), and the total damage ventral to perirhinal cortex
(piriform cortex plus lateral entorhinal cortex). Next, a separate
partial regression considered the extent of damage in the rostral,
mid, and caudal perirhinal cortices. Analyses of postrhinal cortex
damage could not meaningfully be conducted, because so many
cases had no cell loss in this region, and only the small minority
suffered any bilateral damage.
Figure 3. Correlations (Pearson r) between total time spent in close
exploration of the sample object (4, 6, and 8 min conditions combined) and
the overall D1 (novel minus familiar) measure of discrimination. While the
sham animals (n ⫽ 20) benefit from more exploration, the perirhinal group
(n ⫽ 15) show no improvement.
group, F(1, 33) ⫽ 10.67, p ⫽ .003. While simple effects showed
that this D1 difference was significant only for the 6-min sample
period, F(1, 99) ⫽ 7.20, p ⫽ .009, the perirhinal group had lower
D1 scores for all three sample conditions. Furthermore, their D1
scores consistently failed to differ from chance (one-sample t tests,
two-tailed; 4 min, p ⫽ .381; 6 min, p ⫽ .901; 8 min, p ⫽ .168), that
is, the perirhinal group failed to select the novel object. In contrast,
the sham group had mean D1 scores significantly higher than
chance for the two longer sample conditions (4 min, p ⫽ .055; 6
min, p ⫽ .013; 8 min, p ⫽ .001), that is, they successfully
distinguished the novel object from the familiar object.
The same comparisons were made for the just the first 2 min of
each test session (Figure 4B). The pattern of results was similar to
that after 5 min. Although there was now no overall lesion effect
for D1, F(1, 33) ⫽ 2.74, p ⫽ .108, one-sample t tests again showed
that only the sham animals discriminated the novel object with the
longer sample periods (4 min, p ⫽ .129; 6 min, p ⫽ .007; 8 min,
p ⫽ .003). Once again, the perirhinal group failed to discriminate
the novel object in any of the three conditions (one-sample t tests;
4 min, p ⫽ .116; 6 min, p ⫽ .886; 8 min, p ⫽ .297).
Figure 4. Mean D1 scores (plus or minus the standard error of the mean)
for the three sample durations (4, 6, and 8 min). Upper graph (A) shows the
D1 data from all 5 min of the test phase, while the lower graph (B) shows
the D1 data from the first 2 min of each test session. A score significantly
above 0 (one-sample t test) indicates discrimination of the novel from the
familiar object. ⴱ p ⬍ .05. ⴱⴱ p ⬍ .01. ⴱⴱⴱ p ⬍ .001.
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ALBASSER, DAVIES, FUTTER, AND AGGLETON
The first partial regression used the cumulative D1 results (all
three sample conditions combined, 5 min) and revealed a significant negative correlation between recognition memory and overall
perirhinal damage (r ⫽ ⫺0.47, p ⫽ .047; Figure 5, left), that is, the
greater the degree of damage, the poorer the recognition performance. A similar result was found when the D1 scores from just
the first 2 min of each test phase were considered (Figure 5, right),
as again there was a significant negative correlation with total
perirhinal damage (r ⫽ ⫺0.54, p ⫽ .021). Correlations were also
calculated for the total amount of TE damage (5 min, r ⫽ ⫺0.10,
p ⫽ .68; 2 min, r ⫽ ⫺0.051, p ⫽ .84) and of piriform plus
entorhinal cortex (5 min, r ⫽ .33, p ⫽ .18; 2 min, r ⫽ .22, p ⫽
.37), but there was no evidence of a significant relationship with
D1 (see Figure 5).
The second analysis just considered the perirhinal subregions
(rostal, mid, and caudal). Partial regression with the D1 scores
showed that there was always a negative slope, that is, more
damage, worse performance (see Figure 6). The results from the
first 2 min provided the only significant correlation (Figure 6,
right), because the degree of caudal perirhinal damage was related
to D1 performance (r ⫽ ⫺0.47, p ⫽ .04).
Finally, we examined the correlations between the degrees of
damage in the various regions of interest. There was a positive
correlation between the extent of perirhinal damage and extent of
damage in area TE (r ⫽ ⫺0.89, p ⬍ .0001), as well as between
perirhinal cortex and piriform plus entorhinal cortices (r ⫽ .78,
p ⬍ .0001). Likewise, there was a positive correlation between the
tissue loss in the mid and caudal levels of the perirhinal cortex (r ⫽
.77, p ⫽ .0007). No other correlations were significant.
Did the Perirhinal Lesions Affect Rate of Habituation to a
Repeated Object?
There was no evidence that the perirhinal group displayed
unusual levels of exploration or abnormal rates of habituation (see
Figure 7). An analysis of variance using the total exploration times
for each of the four sessions found a highly significant effect of
session as exploration times fell, F(3, 99) ⫽ 92.5, p ⬍ .0001, but
no evidence of a lesion difference, F(1, 33) ⫽ 1.33, or a lesion by
session interaction (F ⬍ 1). The simple effects showed that both
groups displayed a marked reduction in exploration across repeated sessions (both ps ⬍ .001).
Discussion
In keeping with numerous previous studies, perirhinal cortex
lesions disrupted spontaneous object recognition. A significant
recognition deficit was found when the results from the three
different sample phase conditions (4, 6, and 8 min) were combined. Increasing the sample period (from 4 to 8 min) not only
increased the amount of object exploration but also improved
recognition 24 hr later by the sham controls. One consequence was
Figure 5. Correlations (n ⫽ 20) between extent of tissue loss in perirhinal cortex and in two adjacent cortical
regions with recognition performance (cumulative D1 scores). Data are presented for the D1 scores from the
entire session (5 min, on the left) and for the first 2 min (on the right) of each session. The best fit slopes
correspond to the Pearson correlations, while the partial regression results are given in the boxes.
PERIRHINAL CORTEX AND OBJECT RECOGNITION
121
Figure 6. Correlations (n ⫽ 20) between extent of tissue loss in the rostral, mid, and caudal perirhinal cortex
with recognition performance (cumulative D1 scores). Data are presented for the D1 scores from the entire
session (5 min, on the left) and for the first 2 min (on the right) of each session. The best fit slopes correspond
to the Pearson correlations, while the partial regression results are given in the boxes.
that the perirhinal lesion deficit became more evident with the
longer sample periods as the use of a sample period that was too
short (4 min, 24-hr retention) produced floor effects that obscured
any lesion effect. Correlations based on the extent of tissue damage
showed that perirhinal cortex damage correlated significantly with
object recognition, with greater damage associated with poorer
recognition. Within the perirhinal cortex, significant correlations
were found between the loss of recognition memory and the extent
of damage to the caudal perirhinal cortex. Damage to other temporal lobe regions did not correlate with recognition performance.
The first goal was to look at the impact of increasing the sample
time on the effect of perirhinal cortex lesions. The rationale for this
goal derives from the repeated finding that perirhinal cortex lesions do not always impair object recognition. Performance at a
normal level has been reported in rats when there is essentially no
delay (Bartko et al., 2007; Winters et al., 2004) or a short retention
interval of less than 10 min (Ennaceur et al., 1996; Norman &
Eacott, 2004, 2005). The implication is that other brain regions are
better able to support this function if the retention interval is short
and the objects easy to discriminate (Bartko et al., 2007). In
contrast, studies with monkeys imply a delay-independent deficit
after perirhinal lesions (Baxter & Murray, 2001; Ringo, 1991). At
the same time, the effects of rhinal cortex lesions in monkeys
seemingly disappear if object recognition is repeatedly tested with
a small set size (Eacott, Gaffan, & Murray, 1994), that is, repeated
exposure to the same objects disproportionately aids discrimination by those animals lacking the perirhinal cortex. These findings
suggest an encoding deficit that can be compensated for if the
animal spends more time exploring the object in the sample period,
thus increasing its discriminability (Bartko et al., 2007; Eacott et
al., 1994). Support for this suggestion comes from the report that
rats with perirhinal lesions are able to discriminate the novel object
from a familiar object after a 24-hr retention interval when the
familiar object had been presented to the rat for a total of 25 min
over 5 days (Mumby et al., 2007). The present study used sample
times of 4 to 8 min, which includes times longer than those used
as standard but less than the 25 min used by Mumby et al. (2007).
The increase in sample periods in the present study (4 to 8 min) led
to an almost doubling of close-proximity exploration of the sample
object (see Figure 2), but did not aid subsequent discrimination of
novelty after 24 hr by rats with perirhinal cortex lesions.
The failure of increased sampling, within the limits used in the
present study, to ameliorate the perirhinal lesion deficit is reflected
in the correlations (see Figure 3). No relationship was found in the
perirhinal group between the degree of sampling and subsequent
discrimination, and the slightly negative slope of the line of best fit
indicates that the lack of an effect was not due to a failure to use
slightly longer sample periods. While it is possible that the lack of
122
ALBASSER, DAVIES, FUTTER, AND AGGLETON
Figure 7. Bar charts showing the mean sample exploration times (plus or
minus the standard error of the mean) of the sham and perirhinal groups across
the four repeated-sample phases using the same object. Both groups show a
clear and equivalent reduction in exploration, that is, they both habituate.
an improvement by the perirhinal lesion group reflects a floor
effect, this explanation seems less likely given the fourfold increase in sample time by individual rats (Figure 3B) that was not
reflected by any discrimination improvement. Similar sampling
increases by the control group led to marked improvements in
familiarity discrimination (Figure 3A). The lack of an improvement in the perirhinal lesion group might be seen as contradictory
to the predictions of perceptual models of perirhinal cortex function (e.g., Bussey, Saksida, & Murray, 2005) for which factors that
aid the discriminability of visual stimuli should ameliorate the
impact of the surgery. There is, however, a circular problem unless
you have independent measures of (a) what makes stimuli easier to
discriminate and (b) how easy the object is to discriminate initially. In
contrast, the sham control rats showed a significant positive
correlation between sampling time and novelty discrimination.
While this latter relationship is to be expected, the present study
is one of the few occasions for which it has been formally
demonstrated.
A further finding was that habituation across sessions to the
same object appeared normal. This finding is informative, because
any marked deficit in rates of habituation could confound the
spontaneous recognition test. Although habituation within a session was not assessed, the consistent finding that rates of overall
exploration for each sample session did not differ from that exhibited by the control animals would strongly suggest that this was
again normal. Normal levels of habituation after perirhinal damage
might seem surprising, but this would be predicted if the object
was sufficiently salient and could be readily discriminated (Bussey
& Saksida, 2002; Eacott & Gaffan, 2005). A possible shortcoming
is that object recognition for the particular object used in the
habituation study was not subsequently tested for recognition,
though recent findings (Mumby et al., 2007) indicate that the
perirhinal group should have been able to demonstrate clear recognition of the now-familiar object. That same study also found
that the amount of sample explorations did not differ between the
sham and perirhinal lesion groups across sessions (Mumby et al.,
2007), though surprisingly in that study neither group showed a
clear reduction in exploration across sessions that is, habituation.
The ability of rats with perirhinal lesions to discriminate novel
objects after short intervals (Ennaceur et al., 1996; Norman &
Eacott, 2004, 2005) or to benefit from very extensive object
sampling (Mumby et al., 2007) raises questions about the involvement of this area for object-recognition memory. One potential
contributing factor to these examples of spared performance is that
the lesions were incomplete, that is, behavior was supported by
remaining perirhinal cortex. The correlations in the present study
show that sparing is an important element. Another likely factor is
the extent to which other temporal lobe areas can support object
recognition in the absence of the perirhinal cortex. The implication
is that these other areas require more time to compile an object for
recognition, but if so, it is beyond the time scale of the sample periods
used in the present study. One candidate region is the postrhinal
cortex (Nemanic et al., 2004), because it has many similar connections to the perirhinal cortex (Burwell & Amaral, 1998), although it
has been found that rats with postrhinal cortex damage appear to
perform normally on spontaneous object recognition with retention
delays of up to 10 min (Norman & Eacott, 2005). A second candidate
is area TE, because electrophysiological (Zhu, Brown, & Aggleton,
1995), immediate-early gene expression (Zhu, Brown, McCabe, et al.,
1995; Zhu et al., 1996), and lesion correlation (Nemanic et al., 2004)
studies have implicated this area in the visual recognition of novelty.
A third candidate region is the entorhinal cortex, because lesions here
can induce mild delayed nonmatching-to-sample deficits in monkeys
(Leonard, Amaral, Squire, & Zola-Morgan, 1995; Meunier et al.,
1993), though the few studies in rats suggest little or no effect on
recognition memory (Yee & Rawlins, 1998). A notable feature of all
three candidate regions is that they border the perirhinal cortex, and so
partial damage can occur when attempting to make complete perirhinal cortex lesions.
Arguably the most important finding from the structural correlations was the significant relationship between extent of total
perirhinal cortex damage and recognition memory performance.
This correlation is consistent with that found in monkeys (Baxter
& Murray, 2001) where, again, the larger the lesion, the poorer the
recognition performance. While this relationship may seem unsurprising, it helps to cement the notion that the perirhinal cortex is of
critical importance and links more closely findings from rodents
and monkeys (macaques). Furthermore, the slope of this relationship shows that with incomplete damage, you would expect to find
only partial effects, with the resultant need for larger group numbers to demonstrate any impairment. Consistent with this conclusion, an earlier study (Wiig & Bilkey, 1994) that examined rats
with subtotal perirhinal cortex lesions (mean of 25% damage)
found no deficit for the exploration of novel objects.
Within the perirhinal cortex the only significant correlation was
with damage in the caudal perirhinal cortex. This result is intriguing, because studies that have disrupted object recognition by
infusion into the perirhinal cortex have typically targeted the
caudal perirhinal cortex (e.g., Barker et al., 2006; Griffiths et al.,
2008; Warburton et al., 2005; Winters & Bussey, 2005). Furthermore, immediate-early gene expression studies have found that
c-Fos activity in the caudal perirhinal cortex is raised after exposure to novel stimuli (Zhu, Brown, McCabe, et al., 1995; Zhu et al.,
1996; Wan, Aggleton, & Brown, 1999). Caution is, however,
PERIRHINAL CORTEX AND OBJECT RECOGNITION
required in making a specific link with the caudal perirhinal cortex,
because none of the above studies included direct comparisons
with the rostral perirhinal cortex. Furthermore, in the present
study, damage in the caudal perirhinal cortex correlated with other
damage, for example, in the mid-perirhinal cortex. At the same
time, there was no evidence that extraperirhinal cortex damage
correlated with recognition. Although this null result does not
provide definitive evidence, because none of the candidate areas
was systematically targeted, the data clearly support the preeminence of the perirhinal cortex for object-recognition memory.
Finally, it is valuable to consider the implications of the positive
correlation found in the control rats between the amount of object
sampling and object recognition (D1). In both the sample and test
phases, both objects are potentially visible throughout the entire
test period. It is, however, assumed that only close active exploration (in the present study the criterion was ⬍1 cm distance)
provides the information that the animal requires to first learn
about the object and then discriminate the novel object (Ennaceur
& Delacour, 1988). This assumption is integral to the spontaneous
object recognition test, yet it does not appear to have been formally
examined. The present study provides clear support for the view
that close exploration is a valid measure and that it represents
activity that is qualitatively distinct from that which occurs during
the remainder of the session.
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Received May 13, 2008
Revision received July 10, 2008
Accepted July 28, 2008 䡲
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