BEHAVIORAL AND NEURAL BIOLOGY 58, 8--15 (1992)
Selective Lesions in the TemporaI-Hippocampal Region of the Rat" Effects on Acquisition and Retention of a Visual Discrimination Task TROND MYHRER 1
Division for Environmental Toxicology, Norwegian Defence Research Establishment, Kjeller, Norway
The first purpose of this study was to investigate whether lesions in the temporal region may affect acquisition or retention of a discrimination task. In Experiment 1, rats with lesions of the temporal cortex (TC), the lateral entorhinal cortex (LEC), or their interconnections were tested postoperatively in simultaneous brightness discrimination. The results show that neither TC lesions nor LEC lesions affected acquisition of the task, and only LEC lesions impaired retention. TC/LEC transections impaired both acquisition and retention. The second purpose was to investigate effects of hippocampal lesions and perforant path transections on the discrimination task (Experiment 2). Both hippocampal and perforant path lesions impaired acquisition of the task, whereas retention was unaffected. It is suggested that TC and LEC are primarily involved in information storing and that hippocampal function is primarily involved in information processing. © 1992 Academic Press, Inc. The parahippocampal cortex (perirhinal and entorhinal cortices) appears to play an important role in relaying reciprocal connections between neocortical association areas and the hippocampal formation in the rat, cat, and monkey (cf. Room & Groenewegen, 1986). In the rat, the temporal cortex (TC) projects predominantly to the lateral entorhinal cortex (LEC) via the perirhinal cortex (Deacon, Eichenbaum, Rosenberg, & Eckman, 1983; Vayssettes-Courchay & Sessler, 1983; Fig. 1). In turn, LEC projects heavily to TC, whereas the contribution of the medial entorhinal cortex (MEC) is very modest in this respect (Kosel, Van Hoesen, & Rosene, 1982). The fiber connections of TC and LEC in the rat are routed via the adjacent white matter (Vogt, 1974). These corticocortical connections seem to be relayed in the superficial layers in the above areas (Myhrer, Iversen, & Fonnum, 1989). It has been known for some time that the ento1 Please send r e p r i n t requests to Trond Myhrer, Norwegian Defence Centre of Psychology and Education, Oslo m i l / A k e r s h u s , N-0015, Oslo 1, Norway. 0163-1047/92 $5.00 Copyright © 1992 by Academic Press, Inc. All rights of reproduction in any form reserved.
rhinal area is involved in mnemonic processes in rodents (Collier & Routtenberg, 1978; Gauthier & Soumireu-Mourat, 1981; McLardy, 1970; Staubli, Ivy, & Lynch, 1984). Results from a recent study of rats show that damage to either LEC or TC both impaired performance of a preoperatively acquired brightness discrimination task, whereas damage to MEC was without effect. Furthermore, transections of the fiber connections betweent TC and LEC caused an even more pronounced impairment of the discrimination performance than separate damage to these structures (Myhrer, 1991). It may be argued, however, that the impaired performance was associated with a relearning deficit rather than a retention deficit. To clarify this issue it is important to investigate whether lesion effects influence the ability to acquire the task. One intent of the present study was to test effects of damage to TC and LEC and the connections between these areas on acquisition and retention of the discrimination task previously used (Experiment 1). A second intent was to ascertain whether large hippocampal lesions or hippocampal perforant path transections might affect acquisition of the discrimination task (Experiment 2). The projections from LEC and MEC to the hippocampal formation consist of the perforant path systems which originate in the superficial layers of the entorhinal cortices (Segal & Landis, 1974). Thus, a lesion within LEC is able to disconnect this structure from both TC and the hippocampal formation. EXPERIMENT 1
Methods Subjects. Thirty-six male Wistar rats from a commercial supplier (M¢llegaard Breeding Laboratories, Denmark), weighing 290-320 g at the time of surgery, served as subjects. They were randomly
TEMPORAL REGION AND MEMORY
TC
t
FIG. 1. Diagram of a horizontal section showing a simplified ,version of major neural connections in the temporal region (dashed lines). Abbreviations used: LEC, lateral entorhinal cortex; MEC, medial entorhinal cortex; PC, perirhinal cortex; TC, temporal cortex.
assigned to four groups: 8 rats received bilateral TC lesions, 9 received LEC lesions, 9 received transections between TC and LEC, and 10 served as controls. Of these control rats 5 received bilateral control lesions and 5 had only their scalp reflected. The rats were housed individually and had free access to commercial rat pellets and water. The rats were ]handled individually 3 days preoperatively and 1 day postoperatively, being allowed to explore a l~able top (80 x 60 cm) for 3 min per day. The climatized (21°C) vivarium was illuminated from 0700 t~o 1900 h.
Surgery. The rats were anesthetized ip with diazepam (10 mg/kg) and fenatyl fluanisone (2 mg/kg) and placed in a stereotaxic head holder with their skulls horizontal. The TC and LEC lesions were made by aspiration by means of a syringe with cannula (23G, diameter 0.6 mm) connected with a vacuum pump. A hole in the syringe (with piston removed) made it possible to control for suction power. The cannula was provided with a collar to control for insertion depth. The point of insertion for TC lesions was 4.0 mm behind bregma and 6.0 mm lateral to midline. The cannula was inserted 6.0 mm from top of skull in an angle of 15 ° with tip of cannula pointing laterally. Even high power suction only removes a small portion of tissue (about
9
0.6 mm ~) leaving a cavity around the tip of cannula. Additional suction was made 0.8 mm anterior and posterior to the original insertion site by tilting the ll-cm-high syringe parallel to the sagittal plane 4 cm in each direction as measured from top of syringe. The point of insertion for LEC lesions was 7.8 mm behind bregma and 6.0 mm lateral to midline. The cannula was inserted 8.0 mm (from top of skull) in an angle of 10 ° with tip of cannula pointing laterally. Additional suction was made 0.8 mm anterior to the original insertion site by tilting the syringe 4 cm (measured from top of syringe) posteriorly in an approximate occipitothalamic axis. For all types of lesion the opening of the cannula was directed toward the superficial layers of neocortex. The corticocortical connections of the entorhinal and temporal cortices seem to be relayed in layers I-III (cf. Myhrer et al., 1989). These connections would also be disrupted by damage to the deeper layers of neocortex, but to avoid destruction of the white matter, the center of the lesion site was aimed at layers I-III. The TC/LEC lesions were made mechanically by means of the sharp edges of cannulas (diameter 0.5 ram). The cannula to be used was mounted on a syringe. The point of insertion was 7.8 mm posterior to bregma and 6.7 mm lateral to midline. Each cannula was inserted into the brain in a position deviating 20 ° from the vertical in the sagittal plane (tip of cannula pointing rostrally). From this position the syringe was moved 10 times back and forth in an axis deviating about 45 ° from the frontal plane (opening of angle pointing medially). These maneuvers were carried out in two stages with insertion depths 6 and 8 mm from top of skull. In this way, the distal part of the angular bundle was transected at a site corresponding approximately to the level of the rhinal fissure. The control lesions were performed in the same way except that the insertion was made in one stage only with a cannula 3.5 mm long and with a round end.
Histology. Upon termination of testing the brains were removed and frozen. The brains were sectioned horizontally on a CO2-freezing microtome at 30 t~m, every 12th section being preserved. The sections were stained with methylene blue. The percentage of lesion extent was estimated by means of a planimeter for TC and LEC lesions. The extent of fibers transected was estimated from the degree to which the white matter between TC and LEC was damaged at the three dorsoventral levels presented in Fig. 2. The white matter (not the alveus) was divided in four equal columns, each column representing 25% of the fibers. The occurrence of damage
10
TROND MYHRER
was evaluated under relatively high magnification. The number of columns affected at each dorsoventral level were counted, and the mean percentage of damage was computed for each animal.
Apparatus. Testing of simultaneous brightness discrimination was carried out in a Plexiglas cage (56 x 34 x 20 cm) previously described (Myhrer & Nmvdal, 1989). In brief, a Plexiglas wall with an opening (10 x 10 cm) in the middle divided the apparatus in two equal compartments; start compartment and goal compartment. Three interchangeable aluminium cylinders (3 x 7 cm) with a round well (2 x 2 cm) in the top served as discriminanda. The cylinders were located in fixed positions (equal distance between each) along the wall opposite to the partition wall in the goal compartment. The cylinders were natural gray (aluminium) or painted black (except for the well). The well of the positive cylinder was filled with water. The only light was a 15-W bulb 60 cm above the apparatus. Procedure. During acquisition and retention testing the rats were deprived of water for 231/2 h a day. On the first day (Postoperative Day 8), each rat was allowed to explore the empty test apparatus for 15 min. On the second day, the subjects were trained to run from the start compartment into the goal comparment in which they were rewarded with some laps of water from the well in the positive cylinder. The rats were given 10 trials, and intertrial interval was 20 s during which they stayed in their home cage. On the third day, the animals were given trials until the occurrence of five correct responses in succession. Because the task is easily learned, learning criterion was set low to avoid overlearning. Thirteen days after learning criterion had been reached, the animals were tested for retention of the discrimination task. Testing was terminated when the previous criterion was reached. The following behaviors were recorded: number of trials to criterion, numbers and type of errors to criterion. In order to drink or investigate whether the well in a cylinder contained water the rats had to stand on their hind legs with at least one forepaw on top of the cylinder. Error response was scored when a negative cylinder was mounted and found empty of water (e.g., licking the empty well). Approaching or investigating negative cylinders (except the well) was not scored as an error. The positive cylinder was either black or gray and the two cylinders of opposite color were negative. The position of the positive cylinder (left, middle, right) was changed in a prearranged randomized order. One set of randomized positions was used on Day 2 of training
FIG. 2. Reconstructionof brain sectionsindicatinglocations of lesions in three levels of horizontal section.Distancebetween sections: 1.5 mm. ExamplesofTC lesion(A),LEC lesion(B), and TC/LEC lesion (C). and another one on Day 3 and on retention testing. A counterbalanced paradigm was followed in which half of the subjects were trained with black cylinder as positive and the other half with gray cylinder as positive. During the initial phase of learning this task rats frequently put their snout close to negative cylinders and then leave them. Because olfactory cues are of no guidance in this respect, they most likely respond to the color. An approach to positive cylinder is immediately followed by rearing and drinking from the well. As training proceeds, rats gradually cease approaching negative cylinders and head for the positive cylinder when entering the goal compartment. It is not likely that they change their learning strategy at this stage of training by addressing the positive cylinder because of its odd appearance (one positive versus two negative cylinders), since approaching negative cylinders is seen now and then. Statistical overall analyses were made with Kruskal-Wallis one-way ANOVA and group comparisons with two-tailed Mann-Whitney U test.
Results Histology. In the TC group, the lesions were found in areas 36 and 41 of TC (Fig. 2A). The mean
11
TEMPORAL REGION AND MEMORY
TABLE 1 Performance of Simultaneous Brightness Discrimination in Experiment 1 Acquisition Day 1
Retention
Day 2
errors
Errors
Errors
Trials (total)
Trials
Group
N
Median
Range
Median
Range
Median
Range
Median
Range
Median
Range
TC LEC TC/LEC Cont
8 9 9 10
2.0 1.0 2.0 1.5
1-3 0-3 0-3 0-3
0.5 1.0 2.0 1.0
0-1 0-1 1-5 0-2
15 17 18 15
15-19 15-18 16-28 15-19
0.0 2.0 2.0 0.5
0-1 0-3 1-4 0-1
5.0 8.0 12.0 5.0
5-7 5-11 7-17 5-7
Note. Abbreviations used: Cont, control; LEC, lateral entorhinal cortex; TC, temporal cortex.
]percentage of damage that mainly affected the superficial layers was 41% (range 29-52%). In the LEC group, the lesions were limited to LEC (Fig. 2B). The mean percentage of total damage to the superficial layers was 45% (range 29-62%). The TC/LEC lesions appeared as a section through the white matter at a site between TC and LEC (Fig. 2C). The transections, which often affected the alveus of the hippocampal formation, were 0.5-1.0 mm long in rostrocaudal extent and 3 - 4 mm long in dorsoventral extent. In four rats the ventral part of subiculum was damaged unilaterally. Because the cannula transections could not follow the exact curvature of the rhinal fissure, TC/LEC connections between a relatively small part in the caudal end of TC and in the rostral end of LEC were probably not accessible for denervation (in total about one-third of the fibers). The mean percentage of fiber lesion was 92% (range 84-96%) indicating that a total of about 60% of the fibers between TC and LEC were disconnected. Behavior. The sham-operated and lesion-operated control rats did not differ reliably in any measures and were treated as a single control group. Because of uneven distribution of data, nonparametric statistics were applied. K r u s k a l - W a l l i s ANOVA did not confirm significant differences in errors on Day 1 of acquisition (Table 1). On Day 2, however, a significant t r e a t m e n t effect emerged in terms of errors to criterion, H(3) -- 16.00, p < .01. The TC/LEC group made significantly more errors than the Control group (U = 13, p < .02), the LEC group (U = 9, p < .02), and the TC group (U = 4, p < .002). The TC, LEC, and Control groups did not differ reliably from one another. ANOVA did not reveal a significant overall effect in trials to criterion. For retention performance ANOVA confirmed a
reliable effect in errors to criterion, H(3) = 18.10, p < .001. The TC/LEC group made significantly more errors than the Control group (U = 3, p < .002) and the TC group (U = 2, p < .002), but not the LEC group. Also the LEC group made reliably more errors than the Control group (U = 16, p < .05) and the TC group (U = l l , p < .05). A significant treatment effect was seen for trials to criterion, H(3) = 16.91, p < .001. The TC/LEC group used reliably more trials than the Control group (U = 3, p < .002), the TC group (U = 1, p < .002), and the LEC group (U = 15, p < .05). The LEC group also used more trials than the Control group (U = 9, p < .02) and the TC group (U = 7, p < .O2).
EXPERIMENT 2 The results from Experiment i show that damage to neither TC nor LEC affected acquisition of the discrimination task. However, impaired relearning was seen to follow LEC lesions, but not TC lesions. Moreover, TC/LEC disruptions affected both acquisition and relearning. In a previous study, measuring retroactive effects of corresponding lesions, it was seen that both TC and LEC lesions impaired performance and TC/LEC transections produced even stronger effects than the former lesions (Myhrer, 1991). Such retrograde effects on retention of the discrimination task were not seen in rats bearing large hippocampal lesions or perforant path transections (Myhrer & Nmvdal, 1989). Inasmuch as hippocampal function is more involved in cognitive processes than preservation of memory engrams (e.g., Gray & McNaughton, 1983), damage to hippocampal systems might affect acquisition of the brightness discrimination. To examine this possibility rats with hippocampal, perforant path, or neo-
12
TROND MYHRER
Surgery. Procedures were the same as described under Experiment 1. Hippocampal and neocortical lesions were made by aspiration. Total perforant path lesions were made mechanically as described for TC/LEC lesions. The point of cannula insertion was 8.5 mm posterior to bregma and 5.0 mm lateral to midline. Each cannula was inserted into the brain in a position deviating 20 ° from the vertical in the sagittal plane (tip of cannula pointing rostrally). From this position the syringe was moved 10 times back and forth in the frontal plane making a cut of 1.0-1.5 mm through the angular bundle. These maneuvers were carried out in three stages with depth of insertions 4, 6, and 8 mm from surface of the skull to tip of cannula. The control rats received a corresponding incision in the scalp only. Histology. The procedure was as described under Experiment 1. Brains with hippocampal or neocortical lesions were sectioned frontally, every 15th section being preserved.
FIG. 3. Reconstruction of brain sections indicating locations of lesions in coronal sections (A and B) and horizontal sections (C). Examples of hippocampal (A), neocortical (B), and perforant path (C) lesions.
Apparatus and procedure were the same as described under Experiment 1.
Results
cortical lesions were tested in the discrimination task used in Experiment 1.
Histology. The hippocampal lesions comprised about two-thirds of this structure (Fig. 3A). The neocortical areas predominantly affected were 4, 7, 17, and 18, as also seen for the Neocortical group (Fig. 3B). In the PP group, the perforant path was damaged bilaterally in the entire dorsoventral extent in all rats (Fig. 3C). Additional damage was occasionally observed as a small cut in parts of LEC and parasubiculum. A line count method (Myhrer, 1988) revealed a mean percentage lesion efficiency of 94% (range 88-100%).
Me~ods Subjects. Thirty-six male Wistar rats, weighing 290-320 g at the time of surgery, served as subjects. They were randomly assigned to four groups: 9 rats received bilateral hippocampal lesions, 8 received neocortical control lesions, 9 received perforant path lesions, and 10 served as controls. The animals were treated as described under Experiment 1.
TABLE 2 Performance of Simultaneous Brightness Discrimination in Experiment 2 Acquisition Day 1
Day 2
errors
Errors
Retention Trials (total)
Errors
Trials
Group
N
Median
Range
Median
Range
Median
Range
Median
Range
Median
Range
Hipp Neocort PP Cont
9 8 9 10
4.0 1.5 3.0 1.5
2-4 0-3 2-5 0-2
2.0 0.0 1.0 0.5
1-3 0-1 0-2 0-1
20.0 15.0 18.0 15.5
18-25 15-17 15-21 15-17
0 0 0 0
0-1 0-1 0-1 0-1
5 5 5 5
5-7 5-7 5-7 5-7
Note. Abbreviations used: Cont, control; Hipp, hippocampal formation; Neocort, neocortical; PP, perforant path.
13
TEMPORAL REGION AND MEMORY
Behavior. ANOVA confirmed significant differences in terms of errors on Day 1 of acquisition, H(3) = 11.40, p < .01 (Table 2). The Hippocampal (Hipp) group made reliably more errors t h a n the Control group and the Neocortical (Neocort) group (U = 3, p < .002 and U -- 4, p < .002, respectively), but not the Perforant Path (PP) group. The PP group also made significantly more errors t h a n the Control group and the Neocort group (U = 6, p < .002 and U = 11, p < .05, respectively). A significant overall effect was seen for errors on Day 2, H(3) = 16.73, p < .001. The Hipp group made reliably more errors t h a n the Control group (U = 3, p < .002), the Neocort group (U = 2, p < .002), and the PP group (U = 9, p < .02). The PP group did not differ significantly from the Control and Neocort groups. ANOVA confirmed a significant t r e a t m e n t effect for trials to criterion, H(3) = 20.10, p < .001. The Hipp group used significantly more trials t h a n the Control group and the Neocort group (U = 0 and U = 0, respectively, p < .002), but not the PP group. The PP group also used more trials t h a n the Control group and the Neocort group (U = 16 and 13, respectively, p < .05). No significant differences were revealed in retention scores among the groups. DISCUSSION The results from the present study show that acquisition and relearning of a visual discrimination task are differentially affected by various t e m p o r a l hippocampal lesions. In Experiment 1, TC/LEC transections impaired both acquisition and relearning. LEC lesions only impaired relearning, whereas TC lesions were without effects. In Experiment 2, both hippocampal lesions and perforant path transections impaired acquisition, whereas relearning was unaffected. Unimpaired acquisition of the discrimination task in rats with TC or LEC lesions strongly suggests that these lesions affected mnemonic processes rather than the capability to relearn the task in a previous study investigating retroactive effects (Myhrer, 1991). Interestingly, LEC lesions, that yielded a somewhat stronger retroactive effect t h a n TC lesions (Myhrer, 1991), also caused impairment of proactive memory, whereas TC lesions did not. In view of the present findings the acquisition deficit previously seen in rats with TC/LEC transections (Myhrer, 1989; M y h r e r et al., 1989) m a y also be related to mnemonic dysfunction. On Day 1 of acquisition, when all rats were given 10 trials, TC/LEC animals did not m a k e more errors t h a n
TABLE
3
Results Relative to Control Group from Present Experiments with Proactive Memory and Results from Previous Studies of Retroactive Memory Proactive Lesion TC LEC TC/LEC Hipp PP
Acquisition
Retention
Retroactive retention
-$ $
-$ $ --
$ --
$
--
_
-
-
$
Note. Abbreviations used are as for Tables 1 and 2. $, impaired; - - , unchanged.
controls. More errors were only committed on Day 2 of training. Thus, the impaired acquisition may be associated with a deficit in retaining information from one day to another. The notion that hippocampal function may be more involved in information processing than information storing receives support from the results of Experiment 2. Both hippocampal and PP lesions impaired acquisition, but not retention of the discrimination task. Furthermore, the effects of hippocampal lesions were more pronounced than those of PP lesions. This finding of differential effects is tenable with the possibility that PP transections are to be combined with septal disconnections in order to mimic effects of hippocampectomy. In several previous studies, retroactive effects of the lesions used in this study have been investigated in the same apparatus and with the same procedure as in this study (Myhrer, 1991; Myhrer & Nmvdal, 1989). These results along with the present ones are presented in Table 3. As seen from this overview, damage to TC, LEC, or their interconnections affects both proactive and retroactive memory, whereas damage to the hippocampal formation or PP only affects acquisition. As previously discussed, the acquisition impairment in rats with TC/LEC lesions is considered to be of mnemonic nature. Lesion of TC, LEC, or their interconnections impairs retroactive memory more severely than proacrive memory. Moreover, within proactive memory the severity of impairment seems to be related to the length of time in retaining the discrimination task (Myhrer, 1989; Myhrer et al., 1989). The differential effects on retroactive and proactive memory appear compatible with the idea of consolidation of memory. In retroactive memory, the surgical procedure most likely interrupts already established memory traces. In proactive memory, the organism
14
TROND MYHRER
is establishing engrams with an already existing lesion handicap. In the latter case, the acquisition phase may be affected, but when acquired the memory traces may be less vulnerable, because later on they are not interfered with experimentally. In lesion studies of monkeys, effects on proactive memory are reported to be stronger than effects on retroactive memory (cf. Zola-Morgan & Squire, 1986). In such studies, however, hippocampal lesions also comprise enthorhinal cortex, thus probably resulting in a deficit in both relearning and retention. Damage to LEC has previously been seen to impair retention of a discrimination task in rats and mice (Gauthier & Soumireu-Mourat, 1981; Staubli et al., 1984). In a recent study employing a test situation corresponding to the one used in this study, rats with excitotoxic lesions encompassing LEC displayed unimpaired acquisition and impaired retention (Levisohn & Isacson, 1991). Moreover, electrical stimulation of LEC disrupts retention, but not acquisition of a passive avoidance task in rats (Collier & Routtenberg, 1978). These findings correspond well with the present results of LEC lesions. Studies employing selective TC lesions in rats, apart from the one previously cited, do not seem to exist. Hippocampal lesions in previous studies of rats have also been found to interfere more with acquisition than retention of various tasks (e.g., Gray & McNaughton, 1983). The present findings that the hippocampus contributes largely to acquisition and consolidation, whereas neighboring cortical structures contribute largely to storage processes seem to correspond in part with current views derived from studies of primates. Important mnemonic functions have been suggested for the entorhinal and perirhinal cortices (Zola-Morgan, Squire, & Amaral, 1989) and the inferotemporal cortex (Cirillo, George, Horel, & Martins-Elkins, 1989). Previous studies of simultaneous brightness discrimination report that hippocampectomy affected neither acquisition nor retention (Kimble, 1963; Truax & Thompson, 1969). The lack of acquisition deficit contrasts with present findings. However, in both studies cited above a simple two-choice test was applied. In the present test, two negative cylinders versus one positive were used in order to increase cognitive loading of the task. Recent findings from our laboratory show that hippocampal rats learn like normals when using a two-choice version of the present discrimination task (Myhrer, unpublished data). The three-choice paradigm did not make rats respond to oddity (cf. Methods) which probably would require an ability to perceive the
wholeness of the situation. Rats seem to have limited capability in paying attention to wholeness according to Lashley's findings that they respond to restricted parts of discriminanda containing geometrical figures (Lashley, 1938). The lesions of TC and LEC were comparatively small (41 and 45%, respectively), whereas TC/LEC transections affected about 60% of the interconnections between these structures. The more pronounced effects of TC/LEC lesions than separate damage to these areas may be attributable to the differences in lesion efficiency. Another possibility may be that transections of fiber tracts have more disruptive effects on behavior than damage to assemblies of neurons (cf. Markowitsch, 1988). The small additional damage (sections in alveus and subiculum) occasionally seen in a part of the hippocampal formation in TC/LEC animals is not likely to explain the mnemonic effects, because hippocampal lesions did not influence retention. Whether TC/LEC lesions affected fibers of unknown origin important for memory functions remains to be ascertained. Because the present lesions were relatively small, it cannot be ruled out that more complete lesions would have produced somewhat different results. For instance, complete LEC lesions might have impaired acquisition, and complete hippocampal lesions might have impaired retention. Although significant differences were obtained, the reported effects are very small and transient, and animals in all of the lesion conditions rapidly attained norma] asymptotes. The LEC lesions did not affect acquisition of the discrimination task even if these lesions disrupted perforant path fibers. However, present LEC damage comprised less than half of the superficial layers of this structure, thus affecting only about one-fifth to one-sixth of the total fiber systems damaged in the PP animals, because the lateral part of the entorhinal cortex is smaller than the medial one. Clear-cut behavioral changes as a result of extent of neural damage may also be seen in effects of separate TC and LEC lesions in relation to effects ofTC/LEC transections. The site of lesion, however, is crucial, since the relatively large lesions in the Neocortical group were without effects. Results from anatomical studies show that LEC has more extensive projections to the entire neocortical mantle and is more highly differentiated than MEC (Swanson & KShler, 1986). LEC is also provided with a rich system of interneurons in layers II and III which predominantly project to TC and the hippocampal formation (cf. KShler, 1986).
TEMPORAL REGION AND MEMORY
In this central position, LEC appears able to influence information processing by way of its connecLions with the hippocampal formation and information storing by way of its connections with TC (cf. Myhrer, 1989). REFERENCES Cirillo, R. A., George, P. J., Horel, J. A., & Martin-Elkins, C. (1989). An experimental test of the theory that visual information is stored in the inferotemporal cortex. Behavioral Brain Research, 34, 43-53. Collier, T. J., & Routtenberg, A. (1978). Entorhinal cortex electrical stimulation disrupts retention performance when applied after, but not during, learning. Brain Research, 152, 411-417. Deacon, T. W., Eichenbaum, H., Rosenberg, P., & Eckman, K. W. (1983). Afferent connections of the perirhinal cortex in the rat. Journal of Comparative Neurology, 220, 168-190. Gauthier, M., & Soumireu-Mourat, B. (1981). Behavioral effects of bilateral entorhinal cortex lesions in the Balb/c mouse. Behavioral and Neural Biology, 33, 419-436. Gray, J. A., & McNaughton, N. (1983). Comparison between the behavioural effects of septal and hippocampal lesions: A review. Neurosc~ence and Biobehavioral Reviews, 7, 119-188. Kimble, D. P. (1963). The effects of bilateral hippocampal lesions in rats. Journal of Comparative and Physiological Psychology, 56, 273-283. Kosel, K. C., Van Hoesen, G. W., & Rosene, D. L. (1982). Nonhippocampal cortical projections from the entorhinal cortex in the rat and rhesus monkey. Brain Research, 244, 201213. K6hler, C. (1986). Cytochemical architecture of the entorhinal area. In Schwarcz, R. and Ben-Ari, Y. (Eds.). Excitatory amino acids and epilepsy (pp. 83-98). New York/London: Plenum Press. Lashley, K. S. (1938). The mechanism of vision. XV. Preliminary studies of the rat's capacity for detail vision. Journal of General Psychology, 18, 123-193. Levisohn, L. F., & Isacson, O. (1991). Excitoxic lesions of the rat entorhinal cortex. Effects of selective neuronal damage on acquisition and retention of a non-spatial reference memory task. Brain Research, 564, 230-244. Markowitsch, H. J. (1988). Diencephalic amnesia: A reorientation towards tracts? Brain Research Reviews 13, 351-370. McLardy, T. (1970). Memory function in hippocampal gyri but not in hippocampi. Internatmnal Journal of Neuroscience, 1, 113-118.
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Myhrer, T. (1988). The role of medial and lateral hippocampal perforant path lesions and object distinctiveness in rats' reaction to novelty. Physiology and Behavior 42, 371-377. Myhrer, T. (1989). Exploratory behavior, reaction to novelty, and proactive memory in rats with temporo-entorhinal connections disrupted. Physiology and Behavior, 45, 431-436. Myhrer, T. (1991). Retroactive memory of a visual discrimination task in the rat: Role of temporal-entorhinal cortices and their connections. Experimental Brain Research, 84, 517524. Myhrer, T., Iversen, E. G., & Fonnum, F. (1989). Impaired reference memory and reduced glutamergic activity in rats with temporo-entorhinal connections disrupted. Experimental Brain Research, 77, 499-506. Myhrer, T., & Nsevdal, G. A. (1989). The temporal-hippocampal region and retention: The role of temporo-entorhinal connections in rats. Scandinavian Journal of Psychology, 30, 72-80. Room, P., & Groenewegen, H. J. (1986). Connections of the parahippocampal cortex. I. Cortical afferents. Journal of Comparative Neurology, 251, 415-450. Segal, M., & Landis, S. (1974). Afferents to the hippocampus of the rat studied with the method of retrograde transport of horseradish peroxidase. Brain Research, 78, 1-15. Staubli, U., Ivy, G., & Lynch, G. (1984). Hippocampal denervation causes rapid forgetting of olfactory information in rats. Proceedings of the National Academy of Sciences of the United States of America, 81, 5885-5887. Swanson, L. W., & K6hler, C. (1986). Anatomical evidence for direct projections from the entorhinal area to the entire cortical mantle in the rat. Journal of Neuroscience, 6, 30103023. Truax, T., & Thompson, R. (1969). Role of the hippocampus in performance of easy and difficult visual discrimination tasks. Journal of Comparative and Physiological Psychology, 67, 228-234. Vayssettes-Courchay, C., & Sessler, F. M. (1983). Neurophysiologie. Mise en ~vidence de convergences sensorielles dans le cortex entorhinal du Rat. Comptes Rendues de l'Academie des Siences Paris 296, 877-879. Vogt, B. A. (1974). A reduced silver stain for normal axons in the central nervous system. Physiology and Behavior, 13, 837-840. Zola-Morgan, S., & Squire, L. (1986). Memory impairment in monkeys following lesions limited to the hippocampus. Behavioral Neuroscience 100, 155-160. Zola-Morgan, S., Squire, L. R., & Amaral, D. G. (1989). Lesions of the amygdala that spare adjacent cortical regions do not impair memory or exacerbate the impairment following lesions of the hippocampal formation. Journal of Neurosc~ence, 9, 1922-1936.
Selective Lesions in the TemporaI-Hippocampal Region of the Rat" Effects on Acquisition and Retention of a Visual Discrimination Task TROND MYHRER 1
Division for Environmental Toxicology, Norwegian Defence Research Establishment, Kjeller, Norway
The first purpose of this study was to investigate whether lesions in the temporal region may affect acquisition or retention of a discrimination task. In Experiment 1, rats with lesions of the temporal cortex (TC), the lateral entorhinal cortex (LEC), or their interconnections were tested postoperatively in simultaneous brightness discrimination. The results show that neither TC lesions nor LEC lesions affected acquisition of the task, and only LEC lesions impaired retention. TC/LEC transections impaired both acquisition and retention. The second purpose was to investigate effects of hippocampal lesions and perforant path transections on the discrimination task (Experiment 2). Both hippocampal and perforant path lesions impaired acquisition of the task, whereas retention was unaffected. It is suggested that TC and LEC are primarily involved in information storing and that hippocampal function is primarily involved in information processing. © 1992 Academic Press, Inc. The parahippocampal cortex (perirhinal and entorhinal cortices) appears to play an important role in relaying reciprocal connections between neocortical association areas and the hippocampal formation in the rat, cat, and monkey (cf. Room & Groenewegen, 1986). In the rat, the temporal cortex (TC) projects predominantly to the lateral entorhinal cortex (LEC) via the perirhinal cortex (Deacon, Eichenbaum, Rosenberg, & Eckman, 1983; Vayssettes-Courchay & Sessler, 1983; Fig. 1). In turn, LEC projects heavily to TC, whereas the contribution of the medial entorhinal cortex (MEC) is very modest in this respect (Kosel, Van Hoesen, & Rosene, 1982). The fiber connections of TC and LEC in the rat are routed via the adjacent white matter (Vogt, 1974). These corticocortical connections seem to be relayed in the superficial layers in the above areas (Myhrer, Iversen, & Fonnum, 1989). It has been known for some time that the ento1 Please send r e p r i n t requests to Trond Myhrer, Norwegian Defence Centre of Psychology and Education, Oslo m i l / A k e r s h u s , N-0015, Oslo 1, Norway. 0163-1047/92 $5.00 Copyright © 1992 by Academic Press, Inc. All rights of reproduction in any form reserved.
rhinal area is involved in mnemonic processes in rodents (Collier & Routtenberg, 1978; Gauthier & Soumireu-Mourat, 1981; McLardy, 1970; Staubli, Ivy, & Lynch, 1984). Results from a recent study of rats show that damage to either LEC or TC both impaired performance of a preoperatively acquired brightness discrimination task, whereas damage to MEC was without effect. Furthermore, transections of the fiber connections betweent TC and LEC caused an even more pronounced impairment of the discrimination performance than separate damage to these structures (Myhrer, 1991). It may be argued, however, that the impaired performance was associated with a relearning deficit rather than a retention deficit. To clarify this issue it is important to investigate whether lesion effects influence the ability to acquire the task. One intent of the present study was to test effects of damage to TC and LEC and the connections between these areas on acquisition and retention of the discrimination task previously used (Experiment 1). A second intent was to ascertain whether large hippocampal lesions or hippocampal perforant path transections might affect acquisition of the discrimination task (Experiment 2). The projections from LEC and MEC to the hippocampal formation consist of the perforant path systems which originate in the superficial layers of the entorhinal cortices (Segal & Landis, 1974). Thus, a lesion within LEC is able to disconnect this structure from both TC and the hippocampal formation. EXPERIMENT 1
Methods Subjects. Thirty-six male Wistar rats from a commercial supplier (M¢llegaard Breeding Laboratories, Denmark), weighing 290-320 g at the time of surgery, served as subjects. They were randomly
TEMPORAL REGION AND MEMORY
TC
t
FIG. 1. Diagram of a horizontal section showing a simplified ,version of major neural connections in the temporal region (dashed lines). Abbreviations used: LEC, lateral entorhinal cortex; MEC, medial entorhinal cortex; PC, perirhinal cortex; TC, temporal cortex.
assigned to four groups: 8 rats received bilateral TC lesions, 9 received LEC lesions, 9 received transections between TC and LEC, and 10 served as controls. Of these control rats 5 received bilateral control lesions and 5 had only their scalp reflected. The rats were housed individually and had free access to commercial rat pellets and water. The rats were ]handled individually 3 days preoperatively and 1 day postoperatively, being allowed to explore a l~able top (80 x 60 cm) for 3 min per day. The climatized (21°C) vivarium was illuminated from 0700 t~o 1900 h.
Surgery. The rats were anesthetized ip with diazepam (10 mg/kg) and fenatyl fluanisone (2 mg/kg) and placed in a stereotaxic head holder with their skulls horizontal. The TC and LEC lesions were made by aspiration by means of a syringe with cannula (23G, diameter 0.6 mm) connected with a vacuum pump. A hole in the syringe (with piston removed) made it possible to control for suction power. The cannula was provided with a collar to control for insertion depth. The point of insertion for TC lesions was 4.0 mm behind bregma and 6.0 mm lateral to midline. The cannula was inserted 6.0 mm from top of skull in an angle of 15 ° with tip of cannula pointing laterally. Even high power suction only removes a small portion of tissue (about
9
0.6 mm ~) leaving a cavity around the tip of cannula. Additional suction was made 0.8 mm anterior and posterior to the original insertion site by tilting the ll-cm-high syringe parallel to the sagittal plane 4 cm in each direction as measured from top of syringe. The point of insertion for LEC lesions was 7.8 mm behind bregma and 6.0 mm lateral to midline. The cannula was inserted 8.0 mm (from top of skull) in an angle of 10 ° with tip of cannula pointing laterally. Additional suction was made 0.8 mm anterior to the original insertion site by tilting the syringe 4 cm (measured from top of syringe) posteriorly in an approximate occipitothalamic axis. For all types of lesion the opening of the cannula was directed toward the superficial layers of neocortex. The corticocortical connections of the entorhinal and temporal cortices seem to be relayed in layers I-III (cf. Myhrer et al., 1989). These connections would also be disrupted by damage to the deeper layers of neocortex, but to avoid destruction of the white matter, the center of the lesion site was aimed at layers I-III. The TC/LEC lesions were made mechanically by means of the sharp edges of cannulas (diameter 0.5 ram). The cannula to be used was mounted on a syringe. The point of insertion was 7.8 mm posterior to bregma and 6.7 mm lateral to midline. Each cannula was inserted into the brain in a position deviating 20 ° from the vertical in the sagittal plane (tip of cannula pointing rostrally). From this position the syringe was moved 10 times back and forth in an axis deviating about 45 ° from the frontal plane (opening of angle pointing medially). These maneuvers were carried out in two stages with insertion depths 6 and 8 mm from top of skull. In this way, the distal part of the angular bundle was transected at a site corresponding approximately to the level of the rhinal fissure. The control lesions were performed in the same way except that the insertion was made in one stage only with a cannula 3.5 mm long and with a round end.
Histology. Upon termination of testing the brains were removed and frozen. The brains were sectioned horizontally on a CO2-freezing microtome at 30 t~m, every 12th section being preserved. The sections were stained with methylene blue. The percentage of lesion extent was estimated by means of a planimeter for TC and LEC lesions. The extent of fibers transected was estimated from the degree to which the white matter between TC and LEC was damaged at the three dorsoventral levels presented in Fig. 2. The white matter (not the alveus) was divided in four equal columns, each column representing 25% of the fibers. The occurrence of damage
10
TROND MYHRER
was evaluated under relatively high magnification. The number of columns affected at each dorsoventral level were counted, and the mean percentage of damage was computed for each animal.
Apparatus. Testing of simultaneous brightness discrimination was carried out in a Plexiglas cage (56 x 34 x 20 cm) previously described (Myhrer & Nmvdal, 1989). In brief, a Plexiglas wall with an opening (10 x 10 cm) in the middle divided the apparatus in two equal compartments; start compartment and goal compartment. Three interchangeable aluminium cylinders (3 x 7 cm) with a round well (2 x 2 cm) in the top served as discriminanda. The cylinders were located in fixed positions (equal distance between each) along the wall opposite to the partition wall in the goal compartment. The cylinders were natural gray (aluminium) or painted black (except for the well). The well of the positive cylinder was filled with water. The only light was a 15-W bulb 60 cm above the apparatus. Procedure. During acquisition and retention testing the rats were deprived of water for 231/2 h a day. On the first day (Postoperative Day 8), each rat was allowed to explore the empty test apparatus for 15 min. On the second day, the subjects were trained to run from the start compartment into the goal comparment in which they were rewarded with some laps of water from the well in the positive cylinder. The rats were given 10 trials, and intertrial interval was 20 s during which they stayed in their home cage. On the third day, the animals were given trials until the occurrence of five correct responses in succession. Because the task is easily learned, learning criterion was set low to avoid overlearning. Thirteen days after learning criterion had been reached, the animals were tested for retention of the discrimination task. Testing was terminated when the previous criterion was reached. The following behaviors were recorded: number of trials to criterion, numbers and type of errors to criterion. In order to drink or investigate whether the well in a cylinder contained water the rats had to stand on their hind legs with at least one forepaw on top of the cylinder. Error response was scored when a negative cylinder was mounted and found empty of water (e.g., licking the empty well). Approaching or investigating negative cylinders (except the well) was not scored as an error. The positive cylinder was either black or gray and the two cylinders of opposite color were negative. The position of the positive cylinder (left, middle, right) was changed in a prearranged randomized order. One set of randomized positions was used on Day 2 of training
FIG. 2. Reconstructionof brain sectionsindicatinglocations of lesions in three levels of horizontal section.Distancebetween sections: 1.5 mm. ExamplesofTC lesion(A),LEC lesion(B), and TC/LEC lesion (C). and another one on Day 3 and on retention testing. A counterbalanced paradigm was followed in which half of the subjects were trained with black cylinder as positive and the other half with gray cylinder as positive. During the initial phase of learning this task rats frequently put their snout close to negative cylinders and then leave them. Because olfactory cues are of no guidance in this respect, they most likely respond to the color. An approach to positive cylinder is immediately followed by rearing and drinking from the well. As training proceeds, rats gradually cease approaching negative cylinders and head for the positive cylinder when entering the goal compartment. It is not likely that they change their learning strategy at this stage of training by addressing the positive cylinder because of its odd appearance (one positive versus two negative cylinders), since approaching negative cylinders is seen now and then. Statistical overall analyses were made with Kruskal-Wallis one-way ANOVA and group comparisons with two-tailed Mann-Whitney U test.
Results Histology. In the TC group, the lesions were found in areas 36 and 41 of TC (Fig. 2A). The mean
11
TEMPORAL REGION AND MEMORY
TABLE 1 Performance of Simultaneous Brightness Discrimination in Experiment 1 Acquisition Day 1
Retention
Day 2
errors
Errors
Errors
Trials (total)
Trials
Group
N
Median
Range
Median
Range
Median
Range
Median
Range
Median
Range
TC LEC TC/LEC Cont
8 9 9 10
2.0 1.0 2.0 1.5
1-3 0-3 0-3 0-3
0.5 1.0 2.0 1.0
0-1 0-1 1-5 0-2
15 17 18 15
15-19 15-18 16-28 15-19
0.0 2.0 2.0 0.5
0-1 0-3 1-4 0-1
5.0 8.0 12.0 5.0
5-7 5-11 7-17 5-7
Note. Abbreviations used: Cont, control; LEC, lateral entorhinal cortex; TC, temporal cortex.
]percentage of damage that mainly affected the superficial layers was 41% (range 29-52%). In the LEC group, the lesions were limited to LEC (Fig. 2B). The mean percentage of total damage to the superficial layers was 45% (range 29-62%). The TC/LEC lesions appeared as a section through the white matter at a site between TC and LEC (Fig. 2C). The transections, which often affected the alveus of the hippocampal formation, were 0.5-1.0 mm long in rostrocaudal extent and 3 - 4 mm long in dorsoventral extent. In four rats the ventral part of subiculum was damaged unilaterally. Because the cannula transections could not follow the exact curvature of the rhinal fissure, TC/LEC connections between a relatively small part in the caudal end of TC and in the rostral end of LEC were probably not accessible for denervation (in total about one-third of the fibers). The mean percentage of fiber lesion was 92% (range 84-96%) indicating that a total of about 60% of the fibers between TC and LEC were disconnected. Behavior. The sham-operated and lesion-operated control rats did not differ reliably in any measures and were treated as a single control group. Because of uneven distribution of data, nonparametric statistics were applied. K r u s k a l - W a l l i s ANOVA did not confirm significant differences in errors on Day 1 of acquisition (Table 1). On Day 2, however, a significant t r e a t m e n t effect emerged in terms of errors to criterion, H(3) -- 16.00, p < .01. The TC/LEC group made significantly more errors than the Control group (U = 13, p < .02), the LEC group (U = 9, p < .02), and the TC group (U = 4, p < .002). The TC, LEC, and Control groups did not differ reliably from one another. ANOVA did not reveal a significant overall effect in trials to criterion. For retention performance ANOVA confirmed a
reliable effect in errors to criterion, H(3) = 18.10, p < .001. The TC/LEC group made significantly more errors than the Control group (U = 3, p < .002) and the TC group (U = 2, p < .002), but not the LEC group. Also the LEC group made reliably more errors than the Control group (U = 16, p < .05) and the TC group (U = l l , p < .05). A significant treatment effect was seen for trials to criterion, H(3) = 16.91, p < .001. The TC/LEC group used reliably more trials than the Control group (U = 3, p < .002), the TC group (U = 1, p < .002), and the LEC group (U = 15, p < .05). The LEC group also used more trials than the Control group (U = 9, p < .02) and the TC group (U = 7, p < .O2).
EXPERIMENT 2 The results from Experiment i show that damage to neither TC nor LEC affected acquisition of the discrimination task. However, impaired relearning was seen to follow LEC lesions, but not TC lesions. Moreover, TC/LEC disruptions affected both acquisition and relearning. In a previous study, measuring retroactive effects of corresponding lesions, it was seen that both TC and LEC lesions impaired performance and TC/LEC transections produced even stronger effects than the former lesions (Myhrer, 1991). Such retrograde effects on retention of the discrimination task were not seen in rats bearing large hippocampal lesions or perforant path transections (Myhrer & Nmvdal, 1989). Inasmuch as hippocampal function is more involved in cognitive processes than preservation of memory engrams (e.g., Gray & McNaughton, 1983), damage to hippocampal systems might affect acquisition of the brightness discrimination. To examine this possibility rats with hippocampal, perforant path, or neo-
12
TROND MYHRER
Surgery. Procedures were the same as described under Experiment 1. Hippocampal and neocortical lesions were made by aspiration. Total perforant path lesions were made mechanically as described for TC/LEC lesions. The point of cannula insertion was 8.5 mm posterior to bregma and 5.0 mm lateral to midline. Each cannula was inserted into the brain in a position deviating 20 ° from the vertical in the sagittal plane (tip of cannula pointing rostrally). From this position the syringe was moved 10 times back and forth in the frontal plane making a cut of 1.0-1.5 mm through the angular bundle. These maneuvers were carried out in three stages with depth of insertions 4, 6, and 8 mm from surface of the skull to tip of cannula. The control rats received a corresponding incision in the scalp only. Histology. The procedure was as described under Experiment 1. Brains with hippocampal or neocortical lesions were sectioned frontally, every 15th section being preserved.
FIG. 3. Reconstruction of brain sections indicating locations of lesions in coronal sections (A and B) and horizontal sections (C). Examples of hippocampal (A), neocortical (B), and perforant path (C) lesions.
Apparatus and procedure were the same as described under Experiment 1.
Results
cortical lesions were tested in the discrimination task used in Experiment 1.
Histology. The hippocampal lesions comprised about two-thirds of this structure (Fig. 3A). The neocortical areas predominantly affected were 4, 7, 17, and 18, as also seen for the Neocortical group (Fig. 3B). In the PP group, the perforant path was damaged bilaterally in the entire dorsoventral extent in all rats (Fig. 3C). Additional damage was occasionally observed as a small cut in parts of LEC and parasubiculum. A line count method (Myhrer, 1988) revealed a mean percentage lesion efficiency of 94% (range 88-100%).
Me~ods Subjects. Thirty-six male Wistar rats, weighing 290-320 g at the time of surgery, served as subjects. They were randomly assigned to four groups: 9 rats received bilateral hippocampal lesions, 8 received neocortical control lesions, 9 received perforant path lesions, and 10 served as controls. The animals were treated as described under Experiment 1.
TABLE 2 Performance of Simultaneous Brightness Discrimination in Experiment 2 Acquisition Day 1
Day 2
errors
Errors
Retention Trials (total)
Errors
Trials
Group
N
Median
Range
Median
Range
Median
Range
Median
Range
Median
Range
Hipp Neocort PP Cont
9 8 9 10
4.0 1.5 3.0 1.5
2-4 0-3 2-5 0-2
2.0 0.0 1.0 0.5
1-3 0-1 0-2 0-1
20.0 15.0 18.0 15.5
18-25 15-17 15-21 15-17
0 0 0 0
0-1 0-1 0-1 0-1
5 5 5 5
5-7 5-7 5-7 5-7
Note. Abbreviations used: Cont, control; Hipp, hippocampal formation; Neocort, neocortical; PP, perforant path.
13
TEMPORAL REGION AND MEMORY
Behavior. ANOVA confirmed significant differences in terms of errors on Day 1 of acquisition, H(3) = 11.40, p < .01 (Table 2). The Hippocampal (Hipp) group made reliably more errors t h a n the Control group and the Neocortical (Neocort) group (U = 3, p < .002 and U -- 4, p < .002, respectively), but not the Perforant Path (PP) group. The PP group also made significantly more errors t h a n the Control group and the Neocort group (U = 6, p < .002 and U = 11, p < .05, respectively). A significant overall effect was seen for errors on Day 2, H(3) = 16.73, p < .001. The Hipp group made reliably more errors t h a n the Control group (U = 3, p < .002), the Neocort group (U = 2, p < .002), and the PP group (U = 9, p < .02). The PP group did not differ significantly from the Control and Neocort groups. ANOVA confirmed a significant t r e a t m e n t effect for trials to criterion, H(3) = 20.10, p < .001. The Hipp group used significantly more trials t h a n the Control group and the Neocort group (U = 0 and U = 0, respectively, p < .002), but not the PP group. The PP group also used more trials t h a n the Control group and the Neocort group (U = 16 and 13, respectively, p < .05). No significant differences were revealed in retention scores among the groups. DISCUSSION The results from the present study show that acquisition and relearning of a visual discrimination task are differentially affected by various t e m p o r a l hippocampal lesions. In Experiment 1, TC/LEC transections impaired both acquisition and relearning. LEC lesions only impaired relearning, whereas TC lesions were without effects. In Experiment 2, both hippocampal lesions and perforant path transections impaired acquisition, whereas relearning was unaffected. Unimpaired acquisition of the discrimination task in rats with TC or LEC lesions strongly suggests that these lesions affected mnemonic processes rather than the capability to relearn the task in a previous study investigating retroactive effects (Myhrer, 1991). Interestingly, LEC lesions, that yielded a somewhat stronger retroactive effect t h a n TC lesions (Myhrer, 1991), also caused impairment of proactive memory, whereas TC lesions did not. In view of the present findings the acquisition deficit previously seen in rats with TC/LEC transections (Myhrer, 1989; M y h r e r et al., 1989) m a y also be related to mnemonic dysfunction. On Day 1 of acquisition, when all rats were given 10 trials, TC/LEC animals did not m a k e more errors t h a n
TABLE
3
Results Relative to Control Group from Present Experiments with Proactive Memory and Results from Previous Studies of Retroactive Memory Proactive Lesion TC LEC TC/LEC Hipp PP
Acquisition
Retention
Retroactive retention
-$ $
-$ $ --
$ --
$
--
_
-
-
$
Note. Abbreviations used are as for Tables 1 and 2. $, impaired; - - , unchanged.
controls. More errors were only committed on Day 2 of training. Thus, the impaired acquisition may be associated with a deficit in retaining information from one day to another. The notion that hippocampal function may be more involved in information processing than information storing receives support from the results of Experiment 2. Both hippocampal and PP lesions impaired acquisition, but not retention of the discrimination task. Furthermore, the effects of hippocampal lesions were more pronounced than those of PP lesions. This finding of differential effects is tenable with the possibility that PP transections are to be combined with septal disconnections in order to mimic effects of hippocampectomy. In several previous studies, retroactive effects of the lesions used in this study have been investigated in the same apparatus and with the same procedure as in this study (Myhrer, 1991; Myhrer & Nmvdal, 1989). These results along with the present ones are presented in Table 3. As seen from this overview, damage to TC, LEC, or their interconnections affects both proactive and retroactive memory, whereas damage to the hippocampal formation or PP only affects acquisition. As previously discussed, the acquisition impairment in rats with TC/LEC lesions is considered to be of mnemonic nature. Lesion of TC, LEC, or their interconnections impairs retroactive memory more severely than proacrive memory. Moreover, within proactive memory the severity of impairment seems to be related to the length of time in retaining the discrimination task (Myhrer, 1989; Myhrer et al., 1989). The differential effects on retroactive and proactive memory appear compatible with the idea of consolidation of memory. In retroactive memory, the surgical procedure most likely interrupts already established memory traces. In proactive memory, the organism
14
TROND MYHRER
is establishing engrams with an already existing lesion handicap. In the latter case, the acquisition phase may be affected, but when acquired the memory traces may be less vulnerable, because later on they are not interfered with experimentally. In lesion studies of monkeys, effects on proactive memory are reported to be stronger than effects on retroactive memory (cf. Zola-Morgan & Squire, 1986). In such studies, however, hippocampal lesions also comprise enthorhinal cortex, thus probably resulting in a deficit in both relearning and retention. Damage to LEC has previously been seen to impair retention of a discrimination task in rats and mice (Gauthier & Soumireu-Mourat, 1981; Staubli et al., 1984). In a recent study employing a test situation corresponding to the one used in this study, rats with excitotoxic lesions encompassing LEC displayed unimpaired acquisition and impaired retention (Levisohn & Isacson, 1991). Moreover, electrical stimulation of LEC disrupts retention, but not acquisition of a passive avoidance task in rats (Collier & Routtenberg, 1978). These findings correspond well with the present results of LEC lesions. Studies employing selective TC lesions in rats, apart from the one previously cited, do not seem to exist. Hippocampal lesions in previous studies of rats have also been found to interfere more with acquisition than retention of various tasks (e.g., Gray & McNaughton, 1983). The present findings that the hippocampus contributes largely to acquisition and consolidation, whereas neighboring cortical structures contribute largely to storage processes seem to correspond in part with current views derived from studies of primates. Important mnemonic functions have been suggested for the entorhinal and perirhinal cortices (Zola-Morgan, Squire, & Amaral, 1989) and the inferotemporal cortex (Cirillo, George, Horel, & Martins-Elkins, 1989). Previous studies of simultaneous brightness discrimination report that hippocampectomy affected neither acquisition nor retention (Kimble, 1963; Truax & Thompson, 1969). The lack of acquisition deficit contrasts with present findings. However, in both studies cited above a simple two-choice test was applied. In the present test, two negative cylinders versus one positive were used in order to increase cognitive loading of the task. Recent findings from our laboratory show that hippocampal rats learn like normals when using a two-choice version of the present discrimination task (Myhrer, unpublished data). The three-choice paradigm did not make rats respond to oddity (cf. Methods) which probably would require an ability to perceive the
wholeness of the situation. Rats seem to have limited capability in paying attention to wholeness according to Lashley's findings that they respond to restricted parts of discriminanda containing geometrical figures (Lashley, 1938). The lesions of TC and LEC were comparatively small (41 and 45%, respectively), whereas TC/LEC transections affected about 60% of the interconnections between these structures. The more pronounced effects of TC/LEC lesions than separate damage to these areas may be attributable to the differences in lesion efficiency. Another possibility may be that transections of fiber tracts have more disruptive effects on behavior than damage to assemblies of neurons (cf. Markowitsch, 1988). The small additional damage (sections in alveus and subiculum) occasionally seen in a part of the hippocampal formation in TC/LEC animals is not likely to explain the mnemonic effects, because hippocampal lesions did not influence retention. Whether TC/LEC lesions affected fibers of unknown origin important for memory functions remains to be ascertained. Because the present lesions were relatively small, it cannot be ruled out that more complete lesions would have produced somewhat different results. For instance, complete LEC lesions might have impaired acquisition, and complete hippocampal lesions might have impaired retention. Although significant differences were obtained, the reported effects are very small and transient, and animals in all of the lesion conditions rapidly attained norma] asymptotes. The LEC lesions did not affect acquisition of the discrimination task even if these lesions disrupted perforant path fibers. However, present LEC damage comprised less than half of the superficial layers of this structure, thus affecting only about one-fifth to one-sixth of the total fiber systems damaged in the PP animals, because the lateral part of the entorhinal cortex is smaller than the medial one. Clear-cut behavioral changes as a result of extent of neural damage may also be seen in effects of separate TC and LEC lesions in relation to effects ofTC/LEC transections. The site of lesion, however, is crucial, since the relatively large lesions in the Neocortical group were without effects. Results from anatomical studies show that LEC has more extensive projections to the entire neocortical mantle and is more highly differentiated than MEC (Swanson & KShler, 1986). LEC is also provided with a rich system of interneurons in layers II and III which predominantly project to TC and the hippocampal formation (cf. KShler, 1986).
TEMPORAL REGION AND MEMORY
In this central position, LEC appears able to influence information processing by way of its connecLions with the hippocampal formation and information storing by way of its connections with TC (cf. Myhrer, 1989). REFERENCES Cirillo, R. A., George, P. J., Horel, J. A., & Martin-Elkins, C. (1989). An experimental test of the theory that visual information is stored in the inferotemporal cortex. Behavioral Brain Research, 34, 43-53. Collier, T. J., & Routtenberg, A. (1978). Entorhinal cortex electrical stimulation disrupts retention performance when applied after, but not during, learning. Brain Research, 152, 411-417. Deacon, T. W., Eichenbaum, H., Rosenberg, P., & Eckman, K. W. (1983). Afferent connections of the perirhinal cortex in the rat. Journal of Comparative Neurology, 220, 168-190. Gauthier, M., & Soumireu-Mourat, B. (1981). Behavioral effects of bilateral entorhinal cortex lesions in the Balb/c mouse. Behavioral and Neural Biology, 33, 419-436. Gray, J. A., & McNaughton, N. (1983). Comparison between the behavioural effects of septal and hippocampal lesions: A review. Neurosc~ence and Biobehavioral Reviews, 7, 119-188. Kimble, D. P. (1963). The effects of bilateral hippocampal lesions in rats. Journal of Comparative and Physiological Psychology, 56, 273-283. Kosel, K. C., Van Hoesen, G. W., & Rosene, D. L. (1982). Nonhippocampal cortical projections from the entorhinal cortex in the rat and rhesus monkey. Brain Research, 244, 201213. K6hler, C. (1986). Cytochemical architecture of the entorhinal area. In Schwarcz, R. and Ben-Ari, Y. (Eds.). Excitatory amino acids and epilepsy (pp. 83-98). New York/London: Plenum Press. Lashley, K. S. (1938). The mechanism of vision. XV. Preliminary studies of the rat's capacity for detail vision. Journal of General Psychology, 18, 123-193. Levisohn, L. F., & Isacson, O. (1991). Excitoxic lesions of the rat entorhinal cortex. Effects of selective neuronal damage on acquisition and retention of a non-spatial reference memory task. Brain Research, 564, 230-244. Markowitsch, H. J. (1988). Diencephalic amnesia: A reorientation towards tracts? Brain Research Reviews 13, 351-370. McLardy, T. (1970). Memory function in hippocampal gyri but not in hippocampi. Internatmnal Journal of Neuroscience, 1, 113-118.
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