Pii: S0304-3940(01)01773-6

Using a four-arm plus-shaped maze, rats with neurotoxic lesions in the dorsal hippocampus were trained in a spatial reference memory task in which both the extramaze constellation of stimuli and an intramaze landmark (a piece of sandpaper covering the ̄oor of the goal) indicated the location of the goal arm. After reaching criterion, animals were given two transfer tests in order to investigate how much they had learned about the intra and extramaze cues. Results showed that during the intramaze transfer, based exclusively on the intramaze cue, hippocampal rats produced a performance clearly superior to that of the controls. Importantly, the average percentage of correct responses recorded in the hippocampal group during the extra versus the intramaze transfer test was similar. Thus, no decremental processing of the intramaze cue was detected in the lesioned group. In contrast, control rats showed a decremental processing of the intramaze landmark performing worse during the intramaze transfer as compared with the extramaze transfer. These results support the view that the hippocampus regulates decremental changes in the processing of normally irrelevant stimuli, extending this claim into a spatial domain. q 2001 Elsevier Science Ireland Ltd. All rights reserved. Keywords: Learning; Spatial memory; Hippocampus; Place learning; Navigation; Attention Recent research using the Morris water maze has shown that the de®cit in cartographic or locale learning typically observed in hippocampally damaged rats [4,15] or in rats with ®mbria-fornix lesions [14] can be overcome when the animals are trained in the presence of a salient landmark which predicts the location of the submerged platform. In our laboratory, we have replicated this phenomenon using a four-arm plus-shaped maze as apparatus and a spatial reference memory task [12]. In this case, an intramaze landmark was placed in the goal arm. When the animals reached the learning criterion, a transfer test without the intramaze landmark revealed that both control and hippocampal rats manifested a perfect retention of the location of the reward based only on the extramaze con®guration of stimuli. The present study was designed to investigate this phenomenon further. The general aim consisted of studying whether the hippocampal and control animals process the intramaze cue used during the training in the same way. Using classical conditioning paradigms, a number of studies have shown that hippocampal damage disrupts decrements but not increments in conditioned stimulus processing [7,8,13]. When spatial tasks are used and extra and intramaze signals simultaneously predict the location of the goal arm, as in our original work [12], neurologically intact rats manifest decrements in the processing of intramaze cues but not of extramaze landmarks [3]. Thus, the main objective of this study was to investigate whether hippocampal damage disrupts decrements in the processing of intramaze stimulus when intra and extramaze landmarks simultaneously indicate the location of the goal during a navigational problem. The goal of Experiment 1 was to replicate, in the spatial context of our laboratory and using speci®c hippocampus lesions, the well accepted fact that hippocampal lesions impair the normal acquisition of a cartographic or place learning [9]. Fifteen naive male Wistar rats (290±320 g) were individually housed in a room with constant temperature and a 12:12 h light-dark cycle. The animals were fooddeprived to 85% of their normal body weight during the training period. Rats were randomly assigned to a hippocampus (HIP, n ˆ 9) or to a control group (CON, n ˆ 6). Under sodium pentobarbital anaesthesia (50 mg/kg, i.p.), Neuroscience Letters 304 (2001) 89±92 0304-3940/01/$ see front matter q 2001 Elsevier Science Ireland Ltd. All rights reserved. PII: S0304-3940(01)01773-6 www.elsevier.com/locate/neulet * Tel.: 134-958-243763; fax: 134-958-246239. E-mail address: [email protected] (J.M.J. Ramos). the rats were placed in a David Kopf stereotaxic apparatus. The HIP subjects received bilateral injections of N-methyld-aspartate (Sigma, USA. PBS pH 7.4, 0.077 M) in eight sites of the dorsal hippocampus in relation to the interaural zero point [10]: APˆ 5.9, Lˆ^1, Vˆ 6.7; APˆ 5.9, Lˆ^2, Vˆ 6.7; APˆ 4.8, Lˆ^1.5, Vˆ 6.7; APˆ 4.8, Lˆ^3, Vˆ 6.7. The neurotoxin was administered in a 0.4 ml volume at each site through a 30 gauge stainless steel cannula attached to a 5 ml Hamilton microsyringe. Delivery of the solution was carried out with a Harvard Apparatus pump set (model 22, Molliston, USA) at an infusion rate of 30 ml/h. The cannula was left in situ for an additional 3 min before being withdrawn. The rats in the CON group underwent the same surgical procedure except that vehicle injections were administered at the same eight co-ordinates. After a 10-day recovery period, all rats were handled on seven successive days for 5 min each. On the following day the behavioral training began, using a four-arm plus-shaped maze. The apparatus and the characteristics of the testing room have been described elsewhere [11]. During the training procedure, rats received eight trials per session and one session per day. Brie ̄y, at the beginning of a trial, the rat was placed at the end of one of the arms used for starting (S, N or E). Two 45-mg food pellets (P.J. Noyes Company Inc., UK) were placed in the food cup at the end of the west arm. After a choice was made and the rat passed the mid-way point of the chosen arm with all four of its limbs, a wooden cube measuring 10 £ 10 £ 10 cm was placed by the experimenter just behind the rat. In this way the animal remained at the end of the chosen arm for 5±8 s. Then the rat was picked up and con®ned in a box for an intertrial interval of 30 s. The maze was rotated 908 in a clockwise direction from trial to trial. Rats were trained until they reached a learning criterion of at least 14 correct trials on two consecutive days (87%). After the behavioral procedure, the rats were deeply anaesthetized with sodium pentobarbital and perfused intracardially. Several days later the brains were frozen and coronal sections were stained with Cresyl Violet. The histological analysis of Experiment 1 and the following one revealed appropriately positioned bilateral lesions. Representative lesions displaying the maximum and minimum damage resulting from the lesions are represented in Fig. 1. Behavioral results of Experiment 1 indicated that the HIP group presented a profound deterioration in the acquisition of the task. Thus, a one-way ANOVA revealed that the number of incorrect trials to criterion was signi®cantly higher in the HIP than in the CON group (F…1; 13† ˆ 4:97, P , 0:04, Fig. 2A). These results suggest that with our particular spatial task, spatial context and type of lesion it is possible to reproduce previous ®ndings that suggest an important involvement of the hippocampus in the acquisition of allocentric navigation tasks [9]. In Experiment 2 (31 naive male Wistar rats 290±310 g, HIP n ˆ 16 and CON n ˆ 15) two types of landmarks, an intramaze stimulus and the extramaze constellation of stimuli, were present simultaneously during the training, and the two types of landmarks consistently indicated the location of the goal arm. The intramaze cue consisted of a piece of sandpaper (thickness 00) completely covering the ̄oor of the goal arm. The procedure of Experiment 2 was identical to that followed in Experiment 1 except that now the intramaze cue (the sandpaper) was always located in the goal arm (west). When the animals reached the learning criterion, two transfer tests were performed in the 2 days following the end of the training. Each transfer test consisted of eight trials. The order in which the different starting arms were used was randomized, and it was the same for all animals. The aim of each of these transfer tests was to investigate how much each animal had learned about the intra and extramaze cues. During the extramaze transfer, the sandpaper was removed and the animals had only the extramaze con®guration of stimuli to resolve the task. During the intramaze transfer, the animals had only the intramaze cue to solve the task. To eliminate the extramaze con®guration of landmarks during the intramaze transfer test, the maze was surrounded by a black curtain from ceiling to ̄oor and the sandpaper was changed from arm to arm from one trial to the next. The order of the transfer tests was counterbalanced, in such a way that half of the animals carried out the intramaze transfer ®rst. J.M.J. Ramos / Neuroscience Letters 304 (2001) 89±92 90 Fig. 1. Coronal sections showing the largest (gray) and smallest (central white area) hippocampal lesions. Anteroposterior coordinates according to the interaural zero point. Results showed no signi®cant differences between groups in the number of incorrect responses before criterion (F…1; 29† ˆ 0:73). The results of the transfer tests were analyzed using a two-way ANOVA with one betweengroups factor (lesion) and another within subjects factor (type of transfer). This analysis revealed signi®cant differences for the lesion factor (F…1; 29† ˆ 4:10, P , 0:05), type of transfer (F…1; 29† ˆ 27:35, P , 0:00001) and interaction (F…1; 29† ˆ 8:57, P , 0:006, Fig. 2B). The interaction was analyzed using Newman±Keuls post-hoc tests. Interestingly, during the extramaze transfer the performance of the HIP and CON animals was similar, no signi®cant differences being detected (P ˆ 0:37). These results con®rm previous data obtained in our laboratory but this time we used neurotoxic lesions and not electrolytic lesions [12]. Also, these data are in agreement with recent works that indicate that the hippocampus is not absolutely necessary for the learning of a place response [1,4,6]. During the intramaze transfer, the HIP rats recognized the intramaze signal more effectively than the CON animals (P , 0:003). Importantly, post-hoc Newman±Keuls tests also indicated that the average percentage of correct responses recorded in the HIP group during the extra versus the intramaze transfer test, did not differ signi®cantly (P ˆ 0:11, Fig. 2B). Thus, no decremental processing of the intramaze cue was detected in the lesioned group. However, the CON rats performed signi®cantly worse during the intramaze transfer as compared to the extramaze transfer (P , 0:0001, Fig. 2B). So, in agreement with previous ®ndings [3], a decremental processing of the intramaze landmark was detected in normal rats. The central ®nding is that hippocampal lesions interrupted the decremental processing of the intramaze cue in comparison with controls. Using conditioning paradigms, several studies have shown that hippocampal lesions substantially reduced latent inhibition, suggesting that the hippocampus plays a role in regulating decremental changes in attentional processing of the conditioned stimulus [7,8]. In accordance with such ®ndings, our study shows that in an allocentric learning situation, if animals are presented with an intramaze landmark, HIP rats learn more about it than the CON subjects, which suggests that the hippocampus is critically involved in decrements in the processing of initially irrelevant spatial stimuli. The way in which incremented processing of the intramaze cue by HIP rats in ̄uences the normal acquisition of the allocentric task seen in these rats is not known, but one hypothesis would be that the heightened process favours the development of ̄exible navigation strategies [2]. Finally, the data of the present study are congruent with the early notion that the hippocampus is necessary to learn to ignore initially irrelevant stimuli [5]. This research was supported by a grant from the Ministerio de EducacioÂn y Cultura, DireccioÂn General de EnsenÄanza Superior, Spain, PB96±1425. I thank Dr AndreÂs Catena for his helpful comments on this manuscript. [1] Alyan, S.H., Jander, R. and Best, P.J., Hippocampectomized rats can use a constellation of landmarks to recognize a place, Brain Res., 876 (2000) 225±237. [2] Bunsey, M. and Eichenbaum, H., Conservation of hippocampal memory function in rats and humans, Nature, 379 (1996) 255±257. [3] Chamizo, V.D., Sterio, D. and Mackintosh, N.J., Blocking and overshadowing between intra-maze and extra-maze cues: a test of the independence of locale and guidance learning, Q. J. Exp. Psychol., 39B, (1985) 107±125. [4] Day, L.B., Weisend, M., Sutherland, R.J. and Schallert, T., The hippocampus is not necessary for a place response but may be necessary for pliancy, Behav. Neurosci., 113 (1999) 914±924. [5] Douglas, R.J., The hippocampus and behavior, Psychol. Bull., 67 (1967) 416±442. [6] Gaffan, E.A., Bannerman, D.M. and Healey, A.N., Rats with J.M.J. Ramos / Neuroscience Letters 304 (2001) 89±92 91 Fig. 2. (A) Experiment 1: mean number of errors before criterion for the HIP and CON groups (only relevant the extramaze con®guration of stimuli). (B) Experiment 2: mean percentage of correct responses observed in the HIP and CON groups during the transfer tests 1 or 2 days after reaching criterion. Vertical bars in (A,B) represent SEM. hippocampal lesions learn about allocentric place cues in a non-navigational task, Behav. Neurosci., 114 (2000) 895± 906. [7] Han, J.S., Gallagher, M. and Holland, P., Hippocampal lesions disrupt decrements but not increments in conditioned stimulus processing, J. Neurosci., 15 (1995) 7323± 7329. [8] Kaye, H. and Pearce, J.M., Hippocampal lesions attenuate latent inhibition of a CS and of a neutral stimulus, Psychobiology, 15 (1987) 293±299. [9] Packard, M.G. and McGaugh, J.L., Double dissociation of fornix and caudate nucleus lesions on acquisition of two water maze tasks: Further evidence for multiple memory systems, Behav. Neurosci., 106 (1992) 439±446. 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