Entorhinal Cortex Lesions Disrupt the Transition Between the Use of ...

Behavioral Neuroscience 2003, Vol. 117, No. 3, 588 –595

Copyright 2003 by the American Psychological Association, Inc. 0735-7044/03/$12.00 DOI: 10.1037/0735-7044.117.3.588

Entorhinal Cortex Lesions Disrupt the Transition Between the Use of Intra- and Extramaze Cues for Navigation in the Water Maze C. J. P. Oswald

D. M. Bannerman, B. K. Yee, and J. N. P. Rawlins

Cardiff University

Oxford University

R. C. Honey and M. Good Cardiff University

This study with rats examined the effects of excitotoxic lesions to the entorhinal cortex (EC) and hippocampus (HPC) on using extramaze and intramaze cues to navigate to a hidden platform in a water maze. HPC lesions resulted in a disruption to the use of extramaze cues, but not intramaze cues, whereas EC lesions had no effect on the use of these cues when they were encountered for the first time. However, prior navigation training in which 1 type of cue was relevant disrupted navigation with the other type in rats with EC lesions. Results show that the EC contributes to the processing of spatial information, but that this contribution is most apparent when there is a conflict between 2 sources of navigational cues in the water maze.

regarding the platform’s location. Rats with EC lesions were impaired relative to both control rats and those with HPC lesions in locating the platform by using the landmark. This effect of EC lesions on landmark learning was unexpected given recent evidence showing that rats with large parahippocampal cortex lesions can use an intramaze landmark to locate a platform in the water maze (Oswald & Good, 2000). What is more, this latter evidence undermines the obvious possibility that the deficit observed in rats with EC lesions in the second stage of Experiment 1 reflected a problem in learning about landmarks per se. Instead, it seemed to us more plausible to suppose that, in rats with EC lesions, prior learning about one class of cue (i.e., extramaze cues) was exerting some particularly adverse effect on subsequent learning about another class of cues (i.e., intramaze cues). In Experiment 2, we assessed the implications of this line of reasoning concerning the pattern of results observed following lesions to the EC in Experiment 1. In the first stage of Experiment 2 control rats and those with EC lesions were placed in the water maze in which the location of the hidden platform was indicated by a landmark. This stage allowed us to confirm the suggestion that lesions to the EC do not, in and of themselves, disrupt navigation using a landmark (cf. Oswald & Good, 2000). In the second stage, rats from the two groups, Control and EC, were subdivided into two further groups, Present and Absent. For the rats in Group Present, the extramaze cues indicated the platform’s location and the landmark remained in the pool, but it no longer provided accurate information about the platform’s whereabouts. The treatment received by Group Present was formally equivalent to that received by the rats during the second stage of Experiment 1, in which the cues that were relevant with respect to locating the platform in Stage 1 remained present in Stage 2, but were no longer relevant. For the rats in Group Absent, the platform’s location was again indicated by extramaze cues, but the landmark was removed from the water maze. The removal of the intramaze landmark in Group Absent in Experi-

The entorhinal cortex (EC) provides the hippocampus (HPC) with its principal source of polymodal cortical information and is considered, in most models of hippocampal function, to be of central importance to the information processing carried out by the hippocampus (e.g., Eichenbaum, Otto, & Cohen, 1994; O’Keefe & Nadel, 1978; Witter, 1993). It is, therefore, surprising to find that place learning in the water maze, an ability that is severely disrupted by hippocampal lesions (Morris, Garrud, Rawlins, & O’Keefe, 1982), is not affected by neurotoxic lesions of the EC (Bannerman, Yee, Iversen, Rawlins, & Good, 2001; Hagan, Verheijck, Spigt, & Ruigt, 1992; Pouzet et al., 1999; see also Bouffard & Jarrard, 1988; Galani, Obis, Coutureau, Jarrard, & Cassel, 2002). Intrigued by this observation, we conducted a study (reported here as Experiment 1) in which we compared the effects of neurotoxic lesions of the EC and HPC on a matching-to-position task in the water maze. As expected, rats with lesions of the HPC were severely impaired in locating the platform using extramaze cues; and consistent with previous reports, rats with lesions to the EC showed no disruption to place learning. All rats then received training in the same water maze, but with the position of the platform identified by an intramaze landmark rather than the extramaze cues; the extramaze cues from the first stage of training remained present but no longer provided accurate information

C. J. P. Oswald, R. C. Honey, and M. Good, School of Psychology, Cardiff University, Cardiff, United Kingdom; D. M. Bannerman, B. K. Yee, and J. N. P. Rawlins, Department of Experimental Psychology, Oxford University, Oxford, United Kingdom. This research was supported by a grant from the Biotechnology and Biological Sciences Research Council (UK) and the Wellcome Trust (UK) and was conducted when R. C. Honey was a Royal Society University Research Fellow. Correspondence concerning this article should be addressed to M. Good, School of Psychology, Cardiff University, Park Place, Cardiff CF10 3YG, United Kingdom. E-mail: [email protected] 588

ENTORHINAL CORTEX AND SPATIAL NAVIGATION

ment 2 allowed us to determine whether or not any disruption to learning in Stage 2 was dependent on the presence of the previously relevant cue. In the Discussion section, we consider the possible mechanisms that might underlie the pattern of observations in Experiments 1 and 2.

Method Experiment 1 Subjects The subjects were 24 Lister hooded rats (supplied by Harlan Olac, UK) with weights ranging between 400 and 550 g. The rats were housed in pairs on a 12-hr light– dark cycle (lights on at 0700) and maintained on ad-lib food and water throughout the experiment. Twelve rats received bilateral entorhinal cortex lesions (Group EC), 12 rats received bilateral hippocampal lesions, and 12 served as sham-operated controls (Group Sham). The subjects had participated in appetitive Pavlovian conditioning experiments before water maze testing (see Oswald et al., 2002).

Surgery The rats were anesthetized by an intraperitoneal injection of Avertin (for details, see Yee, Feldon, & Rawlins, 1995) and placed in a stereotaxic frame. A bone flap overlying the EC or the HPC was then removed. N-methyl-D-aspartate (NMDA; Sigma Chemicals, Poole, UK) dissolved in 0.1 M phosphate-buffered saline (pH 7.4) was infused at a concentration of 10 mg/ml. In brief, rats in the EC-lesioned group received 16 injections of NMDA at the coordinates specified by Yee et al. (1995). Injections (0.025– 0.100 ␮l) were made over approximately 120 s with a 5-␮l microsyringe (SGE, Milton Keynes, UK) mounted onto a stereotaxic frame. On completion of each infusion, the needle was left in place for a further 120 s to allow for the diffusion of the toxin away from the injection site. Rats in the HPC group received a similar surgical procedure, except that 28 injections (0.05– 0.10 ␮l) of NMDA were made at coordinates specified by Good and Honey (1997). For rats in the sham-operated control group, the track of the needle was limited to the cortex and no neurotoxin was infused.

Apparatus The water maze was 2 m in diameter and 60 cm in depth. The base of the pool was elevated 80 cm above the floor and housed in a room (3.5 m ⫻ 3.0 m) illuminated by four 500-W floodlights. The pool was filled to a depth of 30 cm with water (heated to 24 °C) made opaque by the addition of 2 L of semiskimmed milk. A platform, 10 cm in diameter, was placed in the pool. The top of the platform was 2 cm beneath the surface of the water. During Stage 2, in addition to the platform, an intramaze landmark was placed in the pool. The intramaze landmark was a spherical object with a diameter of 13 cm that was mounted on a stand in such a way that the bottom of the sphere was 2 cm below the surface of the water. The top half of the sphere was painted white to prevent it interfering with the automated tracking system (supplied by HVS Ltd, Hampton, UK), and the bottom half of the sphere was painted black. The extramaze cues were composed of a variety of black and white posters of different shapes and sizes placed on the wall. The walls of the room were painted white, except for the wall that contained the door to the room, which was painted gray. The rats’ behavior in the pool was tracked automatically by means of the system described in Oswald and Good (2000).

Procedure Pretraining. All rats initially received 2 days of nonspatial pretraining in the water maze. The purpose of this pretraining was to familiarize the rats with some of the procedural aspects of the water maze task, for

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example, learning to climb onto an escape platform, and to swim away from the walls of the pool. During pretraining, a curtain was drawn around the pool to conceal the visual extramaze cues. Rats received four trials on each day. On each trial, the rat was released into the pool from one of eight start locations (NE, SE, NW, SW, N, S, E, W; these start locations were used in a pseudorandom order) and was required to locate the submerged platform. The position of the platform moved from trial to trial. Eight of a possible 12 platform locations (NE, SE, NW, SW, N, S, E, W, NE2, NW2, SE2, SW2) were used in a pseudorandom sequence across trials. On a given trial, the rat swam until the platform was mounted or until 120 s (Day 1) or 90 s (Day 2) had elapsed. Failure to locate the platform within this time resulted in the rat being guided to the platform. Within each day, the intertrial interval (ITI) was 30 s, during which the rat remained on the platform. Rats were transported to and from the water maze in an enclosed carrying box. The box was always placed in the same location within the water maze room. At the end of each session, rats were dried with a towel and then returned to their home cages. Stage 1: Matching-to-position. Training on the matching-to-position task commenced 24 hr following the completion of pretraining. The curtain was removed from the perimeter of the pool and secured against the east wall of the room. Rats received 12 days of training. On each day, the platform was placed in a new position in the pool, where it remained for the four trials of that day. The 12 platform positions were used in a pseudorandom sequence, with the constraint that no platform position could be used more than once. On each trial, the rat was placed in the pool facing toward the experimenter and released from the side of the pool. The rat swam until the platform was located or until 120 s had elapsed. If the platform was not located within this time, then the rat was guided to it. Within each day, the ITI between Trials 1 and 2 was 15 s. The ITI between the remaining trials was 30 s. Rats remained on the platform during the ITIs. Other details of the procedure were the same as those described for pretraining. Stage 2: Intramaze landmark training. Stage 2 began 24 hr after the completion of Stage 1. On each of the next 12 days of training, there were 4 trials. In addition to the platform, an intramaze landmark was placed in the pool. The center of the landmark was always situated 30 cm from the center of the platform. For half of the rats in each surgical group, the landmark was always located to the south of the platform, and for the remainder, it was always located to the north. For each rat, the landmark–platform unit moved to a new location on every trial. Thus, on a given trial, the platform occupied one of the 12 platform positions and the landmark was placed, depending upon the subgroups that the rats were assigned to, either 30 cm north or 30 cm south of the platform. The platform positions were used in a pseudorandom order with the constraint that in each block of 12 trials (3 days), each platform position was used on one occasion. Within each day, the ITI was 30 s, during which time the rat remained on the platform. Other details of the procedure that have not been specified were the same as in Stage 1.

Experiment 2 Subjects, Surgery, and Apparatus A naive cohort of 44 Lister hooded rats (supplied by Harlan Olac, UK), with ad lib weights ranging between 400 and 520 g, was used. Twenty rats received bilateral neurotoxic lesions to the entorhinal cortex, and 16 served as sham-operated controls. The care of the rats, the surgical procedures, and the water maze were identical to those of Experiment 1.

Procedure Stage 1: Intramaze landmark training. All rats received 2 days of nonspatial pretraining followed by 12 days of training on the landmark task, as described in Stage 2 of Experiment 1. Immediately after the final session of landmark training, the rats received a probe trial to assess the control exerted by the landmark over searching for the platform. The

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landmark was placed at the center of the pool. The platform was removed from the pool, and the swim pattern of the rats over a 120-s trial was analyzed. The areas used were (a) the area where the platform should have been located (training area), (b) the area opposite to the training area, (c) the area adjacent to the training area on the left-hand side, and (d) the area adjacent to the training area on the right-hand side. Each area was 0.13 m2 and was managed by the computer software. The percentage of time spent during the probe trials in each of the areas was used to determine the approach direction and the distance covered by the rats in searching for the platform from the landmark. Other details of the probe that have not been specified were the same as during Stage 1 training of Experiment 1. Stage 2: Matching-to-position training. Stage 2 commenced 24 hr following the completion of Stage 1. For rats in Group Absent (Absent Sham and Absent EC), the Stage 2 procedure was identical to Stage 1 of Experiment 1. The platform was placed in a new location in the water maze at the start of each day and remained in that location for the four trials of that day; there was no landmark in the pool. For the remaining rats in Group Present (Present Sham and Present EC), the procedure was the same, but in addition to the escape platform, the intramaze landmark was also present in the pool. In contrast to Stage 1, however, the intramaze landmark was a poor cue to the exact location of the platform, as the landmark could occupy any one of the 12 positions within the water maze. The landmark was placed in a different location on every trial according to a pseudorandom sequence, with the constraint that the landmark could not occupy the same location as the platform. All other details of the procedure that have not been specified were the same as those described in Stage 1 of Experiment 1.

EC group. Figure 1B presents photomicrographs of sections from a rat with a representative EC lesion. Hippocampal lesions. All rats in the HPC group sustained damage to the HPC (CA1–3 and dentate gyrus). The lesion incorporated, on average, approximately 95% of the dorsal HPC. There was little or no damage to the subiculum or the EC at this level. More ventrally, the extent of damage to the HPC was more variable between rats. All rats sustained damage to the CA1–3 fields, although the degree of cell loss in the dentate gyrus at ventral levels of the HPC was variable (between 20% and 100%). Cell loss extended into the ventral aspect of the subiculum in 9 of the 12 rats. Inspection of the behavioral data failed to indicate any systematic correlation between the extent of damage to the ventral HPC and performance. Therefore, no rats were excluded from the hippocampal lesion group.

Experiment 2 All rats in the EC group (n ⫽ 20) sustained cell damage that was centered on the EC at all dorsal and ventral levels. However, 1 rat had considerable sparing of the EC and was consequently removed

Histology Following the completion of Experiments 1 and 2, the rats were overdosed with Euthatal (2 ml/kg sodium pentobarbital) and perfused through the heart. The brains were removed from the skull and postfixed in 10% (vol/vol) Formalin for at least 24 hr. The brains were transferred to 25% (wt/vol) sucrose 24 hr prior to being sectioned in the horizontal plane at 60 ␮m on a freezing microtome. The sections were mounted on slides and left to dry prior to staining with Cresyl violet. Other details of the procedure are described in Good and Honey (1997).

Results Histology Experiment 1 EC lesions. All rats in the EC group sustained damage that was centered on both the medial and the lateral EC. Two rats had considerable sparing of the EC at all dorsal and ventral levels and were removed from the statistical analyses. Of the remaining rats, the most substantial damage was to the ventral EC and encompassed both superficial and deep layers. There was some sparing (maximum sparing, approximately 50%) to the deep layers of the most medial and most lateral EC at the dorsal level. The majority of rats (n ⫽ 8) sustained only a limited amount of additional damage to the ventral subiculum and to the ventral pre- and parasubiculum. The damage to these areas did not, however, extend to more dorsal levels (i.e., no greater than 4 mm from the brain surface). Five rats sustained very minor, unilateral damage to the most ventral areas of the adjacent perirhinal cortex. As the extent and selectivity of the lesions in the remaining 10 rats were broadly consistent between rats and there was no apparent damage to the brains of sham-operated rats, no further rats were removed from the statistical analyses. Figure 1A presents a schematic reconstruction of the largest and smallest acceptable lesions in the

Figure 1. The maximum and minimum extent of the lesion in Experiments 1 and 2. A: Entorhinal cortex (EC) lesions from Experiment 1. B: Hippocampus lesions from Experiment 1. C: EC lesions from Experiment 2. Black area represents the smallest lesion, and stippled area the largest, in each group. Each reconstruction is based on panels described by Paxinos and Watson (1998) and are at approximately 3.1, 3.6, 4.6, 5.6, 6.6, 7.6, and 8.6 mm ventral from the brain surface. Reprinted from The Rat Brain in Stereotaxic Coordinates, 4th ed., G. Paxinos and C. Watson, Figures 94, 98, 102, 106, 110, 114, and 116, Copyright 1998, with permission from Elsevier Science.


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from the statistical analyses. Of the remaining rats, the damage was most extensive ventrally and included both deep and superficial layers. In addition, the damage encompassed both the medial and lateral EC. At a more dorsal level, there was some sparing (approximately 40%) of the deep layers of both the medial and lateral EC. In addition to cell loss in the EC, almost all rats sustained cell loss to the ventral pre- and parasubiculum. Ten rats also sustained additional damage to the ventral subiculum. The damage to the ventral subiculum, presubiculum, and parasubiculum did not extend to more dorsal levels. No further damage was detected above 4 mm from the brain surface according to plates from Paxinos and Watson (1998). Finally, 5 rats sustained very minor and unilateral damage to the most ventral aspect of the perirhinal cortex. However, there were no systematic differences between the behavioral data from the rats with minor perirhinal cortex damage and the remaining 14 rats with EC lesions. Figure 1 shows a reconstruction of the largest and smallest lesions in the EC group and the HPC group from Experiment 1 (far left and middle panel, respectively) and the EC group from Experiment 2 (far right panel). Figure 2 presents photomicrographs of sections taken from a sham-operated rat (upper panel) and rats with representative lesions of the EC (middle panel) and the HPC (lower panel).

Behavioral Findings Experiment 1 Stage 1: Extramaze cue training. The mean escape latency (in seconds) for Trials 1– 4 on the matching-to-position task are shown in Figure 3A, averaged across the 12 sessions of training. Inspection of this figure indicates that the EC and sham-operated rats showed rapid within-session learning across the four trials of each session. In contrast, HPC rats showed little or no improvement across training trials. An analysis of variance (ANOVA) with group (Sham, EC and HPC), block (4 ⫻ 3 session blocks), and trial (training Trials 1– 4; see Figure 3A) as variables supported this interpretation, revealing main effects of group, F(2, 31) ⫽ 6.78, p ⬍ .01; block, F(3, 93) ⫽ 26.71, p ⬍ .01; and trial, F(3, 93) ⫽ 30.86, p ⬍ .01. The only significant interaction was between group and trial, F(6, 93) ⫽ 5.09, p ⬍ .01. Analysis of simple main effects revealed that there was a main effect of group on Trials 2, 3, and 4 ( ps ⬍ .05), but not on Trial 1 (F ⬍ 1). In addition, there was a main effect of trial for both sham-operated and EC-lesioned rats ( p ⬍ .02), but no effect of trial in rats with hippocampal lesions (F ⬍ 1). The results clearly show that lesions of the EC do not disrupt spatial navigation in a water maze task that is sensitive to hippocampal damage. Stage 2: Intramaze cue training. The mean escape latencies grouped in 3-day blocks are shown in Figure 3B. Inspection of this figure suggests that all of the groups improved over the course of training. However, the EC group showed consistently slower latencies to find the platform relative to rats with hippocampal lesions and sham-operated rats. An ANOVA with group (Sham, EC, and HPC) and block as variables confirmed the accuracy of this description of the results. The ANOVA revealed main effects of group, F(2, 31) ⫽ 4.53, p ⬍ .05, and block, F(3, 93) ⫽ 40.51, p ⬍ .0001, but no interaction between these factors (F ⬍ 1). Post hoc comparisons (using Newman–Keuls) revealed significant differences between the sham- and EC-lesioned rats ( p ⬍ .05) and

Figure 2. Photomicrographs of representative horizontal brain sections taken at approximately 5 mm from brain surface, illustrating the typical extent of damage in sham-lesioned control (upper panel) and entorhinal cortex-lesioned (middle panel) and hippocampus-lesioned (lower panel) rats.

between the HPC- and EC-lesioned rats ( p ⬍ .05). The sham- and HPC-lesioned group did not differ significantly from each other ( p ⬎ .10). Swim speed. Table 1 presents the mean swim speed for Groups Sham, EC, and HPC in each stage of training. Inspection of this table indicates that there was a tendency for EC rats to swim more slowly, and that the three groups of rats swam more slowly in Stage 2 than they did in Stage 1. An ANOVA with group (Sham, EC, or HPC) and stage as variables confirmed these impressions, revealing a main effect of stage, F(1, 31) ⫽ 68.25, p ⬍ .0001; a main effect of group, F(1, 31) ⫽ 4.06, p ⬍ .05; and no interaction between these factors, F(1, 31) ⫽ 1.73, p ⬎ .19. The main effect of group was analyzed with post hoc comparisons (Newman–

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Figure 3. A: Latency to find the platform over the four trials of each session averaged over the course of training on the matching-to-position task in the water maze in Experiment 1. B: Latency to find the platform averaged over three-session blocks in the intramaze landmark task. Present ⫽ intramaze landmark present during Stage 2; Absent ⫽ intramaze landmark absent during Stage 2; Sham ⫽ sham-operated; EC ⫽ entorhinal cortex-lesioned; HPC ⫽ hippocampus-lesioned. Data are means (⫾ SEM).

Keuls) and revealed a significant difference between the EC and HPC groups ( p ⬍ .05); neither lesion group, however, differed significantly from the sham-operated control group.

Experiment 2 Stage 1: Intramaze cue training. Inspection of Figure 4A suggests that the Groups Sham and EC both exhibited a comparable improvement in performance over the course of training. An ANOVA with group (Sham or EC), future condition (absent or present), and block as variables confirmed this description of the results. The ANOVA revealed a main effect of block, F(1, 31) ⫽ 47.59, p ⬍ .0001, but no effects of group, F(1, 31) ⫽ 1.55, p ⬎ .22, or future condition (F ⬍ 1), and no interactions between any of the factors (Fs ⬍ 1). Landmark probe test. To examine the swim patterns of the rats during the probe test, we analyzed the amount of time spent swimming in each of four areas around the landmark. In particular, the amount of time spent swimming in each of the areas was used to calculate an elevation ratio for each rat: The amount of time

spent swimming in the training area was divided by the combined total of the amount of time spent swimming in the training area and the average amount of time spent swimming in the other three areas. On the basis of this measure, a score above .50 indicates that the rats spent more time swimming in the training area than in the other three areas. The mean elevation ratios for Groups Sham and EC groups were .64 and .58, respectively. There was no significant difference in the average time spent swimming in the adjacent and opposite areas to the training location by the two groups: Sham ⫽ 17.6%, EC ⫽ 14.5%, F(1, 33) ⫽ 3.50, p ⬎ .07. Finally, one sample t tests comparing the elevation ratios from each group to a ratio of .50 (the ratio expected if the rats swim in all four areas at random) revealed that both rats with EC lesions and control rats showed a preference to swim in the training area compared to the other areas (both ps ⬍ .01). Stage 2: Extramaze cue training. Figure 4B shows the mean latencies to locate the escape platform on Trials 1– 4 averaged across all 6 days of training for the four groups of rats (Present Sham, Absent Sham, Present EC, Absent EC). Inspection of Figure 4B suggests that the presence of the intramaze landmark from Stage 1 training resulted in an initial disruption to acquisition of the matching-to-position task. However, both of the Sham groups and Group Absent EC showed an improvement in within-session performance. In contrast, the within-session improvement of Group Present EC was more modest. An ANOVA with group (Sham or EC), condition (Present or Absent), and trial as variables provided support for this description of the results and revealed main effects of group, F(1, 31) ⫽ 7.67, p ⬍ .01; condition, F(1, 31) ⫽ 9.05, p ⬍ .01; and trial, F(3, 93) ⫽ 57.58, p ⬍ .01. In addition, there was a significant interaction between group, condition, and trial, F(3, 93) ⫽ 3.92, p ⬍ .05. None of the other interactions were significant (Fs ⬍ 1). Additional ANOVAs (Present Sham vs. Present EC, Absent Sham vs. Absent EC) were conducted to examine further the interaction between group, condition, and trial. An ANOVA comparing the two Present groups (Present Sham and Present EC) revealed main effects of group, F(1, 15) ⫽ 4.72, p ⬍ .05, and trial, F(3, 45) ⫽ 32.78, p ⬍ .0001, and a significant interaction between these factors, F(3, 45) ⫽ 3.82, p ⬍ .02. Analysis of simple main effects revealed that Group Present Sham located the platform more rapidly than Group Present EC on Trial 2, F(1, 44) ⫽ 5.98, p ⬍ .05; Trial 3, F(1, 44) ⫽ 5.93, p ⬍ .05; and Trial 4, F(1, 44) ⫽ 4.59, p ⬍ .05, but there was no difference in the performance of the two Present groups on

Table 1 Average Swim Speeds During Each Stage of Training in the Water Maze Group

Stage 1

Stage 2

Sham EC HPC

Transfer from extramaze to intramaze cues 0.39 0.38 0.40

0.37 0.35 0.38

Sham EC

Transfer from intramaze to extramaze cues 0.37 0.38

0.37 0.37

Note. Swim speeds are in meters per second. Sham ⫽ sham-lesioned rats; EC ⫽ entorhinal cortex-lesioned rats; HPC ⫽ hippocampus-lesioned rats.


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Figure 4. A: Latency to find the platform averaged over three-session blocks in the intramaze landmark task in Stage 1 of training in Experiment 2. Each lesioned group is broken down into the condition to which rats were to proceed in Stage 2. B: Latency to find the platform averaged over the four trials of each session during training on the matching-toposition task in Stage 2. Sham ⫽ sham-operated; EC ⫽ entorhinal cortexlesioned. Data are means (⫾ SEM).

Trial 1 (F ⬍ 1). There was also an effect of trial in both Group Present Sham, F(3, 45) ⫽ 26.89, p ⬍ .01, and Group Present EC, F(3, 45) ⫽ 8.63, p ⬍ .01. An ANOVA comparing the performance of the two Absent groups (Absent Sham or Absent EC) revealed a main effect of trial, F(3, 48) ⫽ 25.22, p ⬍ .01, but no effect of group, F(1, 16) ⫽ 2.80, p ⬎ .11, and no interaction between these factors (F ⬍ 1). Swim speed. The mean swim speeds for each group in Stages 1 and 2 are shown in Table 1. Inspection of this table reveals that the swim speeds for the two groups were very similar in both stages of the experiment. An ANOVA performed on the swim speeds, with group (Sham or EC) and stage as variables, failed to find any effects of group or stage (Fs ⬍ 1), and no interaction between these factors, F(1, 33) ⫽ 2.65, p ⬎ .11.

The role of the EC in spatial navigation was investigated in tasks in which extramaze and intramaze cues provided information about the location of a hidden platform in the water maze. Rats with lesions of the EC had no difficulty learning the location of the platform with respect to either extramaze cues or an intramaze cue, provided that these cues were being encountered for the first time (see also Oswald & Good, 2000; Pouzet et al., 1999). This pattern of results is quite different from that observed in hippocampal rats, which exhibited a profound deficit in using extramaze cues to locate a hidden goal, but no deficit in using an intramaze cue to do so (see also Long & Kesner, 1996, 1998; Pearce, Roberts, & Good, 1998). The observation that rats with lesions of the EC do not show this pattern suggests that the navigational processes for which the HPC is critical can be supported by cortical–subcortical inputs other than those inputs provided by the EC (see Aggleton, Vann, Oswald, & Good, 2000). Indeed, taken in isolation, the finding that selective lesions to the EC had no marked effect or immediate influence on spatial learning in the water maze involving extramaze cues alone, or in combination with an intramaze cue, might suggest that the EC plays no role in processing spatial information (Pouzet et al., 1999). However, subsequent stages of Experiment 1 and Experiment 2 revealed that the EC contributes to spatial learning when there is a transition between a period of training in which one type of cue has been relevant to locating the platform (e.g., extramaze cues) and a second period of training in which these cues become irrelevant and a second set of cues (e.g., intramaze cues) become relevant. This pattern of results indicates that the EC does contribute to the processing of spatial information, and furthermore, they suggest what form this contribution might take. Lesions of the EC appear to compromise a process that is particularly likely to be engaged or recruited when there is a transition between the use of one type of cue (e.g., an intramaze cue) during the first stage of training and another type of cue during the second stage of training (e.g., extramaze). Furthermore, the disruption caused by the EC lesion was evident only under conditions in which a formerly relevant cue was present during the second stage of training (see Experiments 1 and 2). This suggests that the critical process involved learning not to respond on the basis of these cues, which were present but rendered irrelevant. There are a number of processes that might be involved in this form of learning. That is, there are a number of processes that might contribute to extinction during spatial navigation. For example, inhibitory learning processes are often thought to underlie extinction, and such learning might contribute to changes in spatial behavior in the water maze. To the best of our knowledge, however, there is no independent evidence to suggest that the EC contributes to inhibitory learning. Indeed, Pouzet et al. (1999) reported that rats with EC lesions learned a spatial reversal task in the water maze as efficiently as controls. Furthermore, on tasks such as nonspatial discrimination reversal learning, in which inhibitory learning might be expected to play an important role, lesions of the EC do not impair performance (e.g., Oswald et al., 2001; Otto, Schottler, Staubli, Eichenbaum, & Lynch, 1991; Yee & Rawlins, 1998). Another possibility is that lesions of the EC may disrupt the transition between the use of different navigation strategies that are engaged by working memory and reference memory procedures.

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However, this seems an unlikely explanation for the present pattern of results. In a recent study, Galani et al. (2002) examined the effects of NMDA-induced lesions of the EC on performance of a spatial radial maze task. Sham-operated and EC-lesioned rats were trained first on a working memory task and then on a reference memory version of the task in the same apparatus. The transfer from the working memory task resulted in a temporary increase in the number of errors in both sham-operated and EC-lesioned rats on the reference memory procedure. However, the performance of rats with EC lesions was indistinguishable from that of controls. Thus, under conditions in which the stimulus dimension that contributes to spatial navigation remains consistent, damage to the EC does not disrupt the transition between the use of strategies engaged by spatial working memory and reference memory tasks. The results of the present study suggest that the EC contributes to processes that are engaged when there is a transition between the type of cue that is used for navigation. One process that may be engaged when there is a transition between the use of one type of cue (e.g., intramaze cue) and another type of cue (e.g., extramaze) is attention. There is evidence that relevant and irrelevant cues gain and lose, respectively, the capacity to command attention and control spatial navigation (see Biegler & Morris, 1999; March, Chamizo, Mackintosh, 1992; Prados, Chamizo, & Mackintosh, 1998; Roberts & Pearce, 1999; Rodrigo, Chamizo, McLaren, & Mackintosh, 1997). Furthermore, there is a growing body of evidence showing that the EC contributes to the modulation of attention to cues. For example, rats with lesions of the EC do not show two effects that are considered to reflect the operation of attentional processes in animals: Rats with lesions of the EC show neither latent inhibition (Lubow & Moore, 1959; e.g., Coutureau, Galani, Gosselin, Majchrzak, & Di Scala, 1999; Coutureau, Gosselin, & Di Scala, 2000; Coutureau, Lena, Dauge, & Di Scala, 2002; Oswald et al., 2002; Yee et al., 1995, 1997), nor do they show an IDS/EDS (intra-dimensional shift/ extra-dimensional shift) effect (Oswald et al., 2001). In the present tasks, all rats enter the second stage of training with a tendency to respond to those cues that guided navigation in the first stage. This tendency directly interferes with performance during Stage 2, and any process that facilitates the rats’ ability to relinquish the reliance on the irrelevant cue (or cues) from the first stage will promote performance. Clearly if rats with EC lesions fail to, for example, “tune out” this irrelevant cue, there will be greater interference between what they learned in Stage 1 and what they are required to learn in Stage 2. Burwell (2000) has recently suggested an anatomical model illustrating how the EC may contribute to visuospatial attentional processes. According to Burwell, the connection between the lateral EC and the perirhinal cortex may provide access to prefrontal areas involved in attention. Together with reward information received from subcortical structures (see Pitkanen, Pikkarainen, Nurminen, & Ylinen, 2000), this system may support “focused attentional processing of behaviorally relevant stimuli” (p. 40). In addition, Burwell suggests that connections between the medial EC and the postrhinal cortex may provide this region with links to systems involved in visuospatial orienting, for example, the posterior parietal cortex and the lateral posterior nucleus of the thalamus. These parallel cortical systems may, therefore, provide the EC with appropriate sensory and motor information to modulate attention to environmental stimuli.

The results of the present study indicate that neurotoxic lesions of the EC do not disrupt spatial navigation per se. This finding contrasts with earlier evidence that mechanical lesions of the EC were sufficient to disrupt spatial navigation in the water maze (e.g., Schenk & Morris, 1985). An obvious explanation for this discrepancy is the use of different lesion techniques, for example, neurotoxic versus radiofrequency, and the concomitant differential disruption to fibers of passage and damage to related brain regions, such as the subiculum, that play a critical role in spatial learning (see Bannerman et al., 2001 for discussion). The absence of an effect of EC lesion on acquisition of spatial information in the water maze, but presence of such an effect on the transition between the use of different navigation cues, underlines the suggestion made by Aggleton et al. (2000) that multiple systems contribute to spatial learning. The EC may therefore play an important role in modulating the access of cortical (or subcortical) navigation systems to hippocampal processing as a result of changes in environmental conditions or contingencies during navigation (see Aggleton et al., 2000 for further discussion). In general terms, the results of Experiments 1 and 2 confirmed the finding that excitotoxic lesions of the EC have no immediate impact on spatial learning in the water maze. These findings are in agreement with anatomical and electrophysiological evidence in suggesting that the EC makes a distinctive contribution to learning (e.g., Burwell, 2000; Sharp, 1999; Witter et al., 2000; see also Gluck & Myers, 1993; Myers et al., 1995; Schmajuk, Cox, & Gray, 2001). This view received further support from our finding that the EC contributes to spatial navigation under circumstances in which there is a transition between different types of navigational cue that are relevant to locating the platform in the water maze. These findings are in general agreement with the proposition that lesions of EC disrupt the modulation of attention to cues both within and without the spatial domain (see Oswald et al., 2001, 2002).

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Received June 19, 2002 Revision received September 9, 2002 Accepted October 25, 2002 䡲