Kindling of the hippocampus induces spatial memory deficits in the rat

Neuroscienee Letters. 63 (1986) 115 120

II 5

Elsevier Scientific Publishers Ireland Ltd.

NSL 03688

KINDLING OF T H E H I P P O C A M P U S INDUCES SPATIAL M E M O R Y DEFICITS IN T H E RAT

F.H. LOPES DA SILVA, J.A. G O R T E R and W.J. W A D M A N

Neurophysioh~gy Group, Department of General Zoology, Biologisch Centrum, Bldg. 11, University ~/' Amsterdam, Kruislaan 320, 1098 SM Amsterdam (The Netherlands) (Received August 24th, 1985; Accepted September 15th, 1985)

Key wordsv hippocampus - working memory

reference memory - 8-arm maze - kindling

epilepto-

genesis - rat

Since kindling produces electrophysiological and morphological changes in the brain area stimulated, it may well affect behavioural functions dependent on the kindled area. Using an 8-arm maze, it was found that hippocampal kindling can induce specific memory deficits in spatial tasks. Reference (long-term) memory as well as working (short-term) memory were impaired. The largest impairment was observed during the period in which generalized convulsions occurred. Working memory but not reference memory impairment was reversible. Hippocampal kindling may be a useful experimental model for investigating behavioural deficits correlated with epileptogenesis.

Application of a tetanus to certain brain structures at regular intervals results first in local ictal and interictal phenomena, and ultimately in characteristic motor seizures that closely resemble those presented by epileptic patients. This is the so-called kindling model of epileptogenesis [3, 13]. Physiological [15] and minor morphological [6] changes have been demonstrated in and around the kindling area, but the cellular basis of the process is still unclear. It is also unknown to what extent the creation of such an epileptogenic focus may affect the functioning of the structure, although considerable abnormality can be expected. There is ample evidence, most of it resulting from studies employing bilateral lesions, that the hippocampus plays an important role in memory tasks [8], especially those involving spatial cues [4, 5, 7, 9]. We reasoned that if hippocampal kindling were to produce long-lasting deficits in hippocampal function, these ought to be revealed by testing rats in such a maze during the process of epileptogenesis. Until now, hippocampal kindling was only found to cause a decrease in locomotor activity, but this was limited to 2 h after a seizure [1]. An amnestic syndrome was reported in relation to hippocampal epileptic activity, but this was caused by tetanus toxin, and did not lead to lasting impairment of short-term memory [2]. Indeed, it has not yet been established whether hippocampal kindling can cause memory deficits. Fourteen male Wistar rats were successfully implanted with chronic electrodes in area CA 1, allowing stimulation and tetanization of the Schaffer collaterals of the left hippocampus as well as recording of the responses from the pyramidal cell area. 0304-3940/86/$ 03.50 © 1986 Elsevier Scientific Publishers Ireland Ltd.

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Under pentobarbitone anesthesia, two bundles o f 100-#m stainless-steel wires were placed in area C A I o f the left dorsal hippocampus; electrophysiological control was used to o p t i m i z e the position o f the t w o recording electrodes (around stratum pyramidale) and the three stimulation electrodes (at least one in stratum radiatum or lacunosum). Recordings o f field potentials were used to m o n i t o r electrode positions. Tetani (2 s, 50 Hz, 1 ms pulses of 2 0 0 - 3 0 0 #A), were applied twice daily by way of two o f the stimulation electrodes as described before [14]. Afterdischarges were recorded on paper for control. The behavioural tests were performed in an 8-arm radial m a z e [11]: the same 5 adjacent arms were baited with food at the beginning o f each trial and the other 3 were always e m p t y [10]. The correct response o f the rat was to enter each o f the 5 baited arms only once. Deviations from this response were scored as errors and could be divided into two different classes: (1) if the rat entered any arm a second time or m o r e times, this was scored as a working m e m o r y ( W M ) error; (2) when it entered an unbaited arm for the first time, this was scored as a reference m e m o r y ( R M ) error. It m a y be noted that if a rat entered a non-baited arm m o r e than once, this was P e r f o r m a n c e in a w o r k i n g m e m o r y t a s k control rots 0

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Fig. 1. Distribution along trials (days) of errors made in WM by kindled rats and controls. The period when the animals showed generalized seizures is indicated by the hatched bar. Controls were matched at random to kindled rats in order to obtain the same total time of observation in the two groups. Note (a) the increase in error scores of kindled rats as compared to controls and (b) the differences among periods I, II and III for the kindled rats; statistics indicated in Table I.

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scored as a W M error; however, since it seldom happened in the kindled rats (6 times out of a total of 196 trials) and never in controls, scoring these errors as RM instead of WM errors would not affect the conclusions. Rats were maintained at an average weight o f 300-350 g during the entire experimental period in a reversed light-dark cycle. Rats were initially trained until a stable performance was obtained, i.e. at least 4 baited arms had been visited in the first 5 choices. Each trial lasted either until they had chosen all 5 baited arms or 15 min had elapsed. After implantation, the rats were allowed two weeks for recovery, followed by 25 days o f retraining until a stable performance was regained. Rats were randomly assigned to a control or an experimental kindling group of equal size, which was treated identically except for the tetanization. Each rat was submitted daily to a behavioural test starting 1-2 h after the onset of the dark period; a tetanus was applied 1-2 h following this test. A second kindling stimulus was administered about 5 h later, in order to reduce the time needed for epileptogenesis. Each stimulus evoked an afterdischarge that lasted ca. 20 s at the beginning, but 100 s by the end of the kindling process. Kindling was continued until the first generalized seizure was observed (Racine's stage 5 [13]). This period varied between rats (range 6 to 17. days) and will be called period I. From then onwards, the tetanus was applied once a day until 9 generalized seizures had been observed

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(period II). Thereafter, the behavioural tests were continued for 7 consecutive days without tetanization, in order to study possible recovery (period III). The performances of the rats in WM and in RM are given as a function of time in Figs. 1 and 2, respectively, while statistical comparisons between the two groups and between different periods are summarized in Table I. 'Error score' was defined as the mean number of errors per daily trial according to the definitions given above. Comparisons between kindled rats and controls were carried out by using the scores of rats from identical periods of observation. Since no significant differences (P > 0.05) were found for controls in the various periods, the mean score for controls over the whole time-span could equally well have been used; doing so had no influence on the conclusions shown in Table I. No significant difference in WM or in RM scores was found between the two groups in the period before kindling started (Mann-Whitney U-test, two-tailed). During kindling, the comparisons between kindled animals and controls showed a significant difference for all periods in the RM score. This difference increased slightly from period I to II, and persisted even after kindling was discontinued (III). For the WM score, a significant difference between kindled animals and controls was also found during kindling (I, II), but performance in WM improved once stimulation was stopped, and was not different from controls during period III. Comparisons among the 3 experimental periods were made for the kindled rats by means of the Wilcoxon matched-pairs signed-rank test. RM did not change significantly between either periods I and II or periods II and III, implying that there was little or no recovery after discontinuation of stimulation. In contrast, WM performance deteriorated from period I to II but recuperated from period II to III, to such an

TABLE 1 MEAN E R R O R SCORES 1N WM A N D RM Mean number of errors per trial in working and reference memory for rats (n = 7) and controls (n = 7) for the 3 experimental periods (I, II, III). For controls, no significant differences among periods were found. Experimental period

Mean error scores .................................. Kindled Controls rats

Comparison: controls vs kindled*

Comparison among periods for kindled rats** Period

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RM

I

WM RM

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P<0.01 P<0.05

I

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P<0.05

n.s.

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0.62 1.60

0.06 0.08

P<0.001 P<0.001

II vs III

P<0.01

n.s.

III

WM RM

0.18 1.39

0.11 0.56

n.s. P<0.001

1 v s III

n.s.

n.s.

*Mann Whitney U-test (two-tailed). **Wilcoxon matched-pairs signed-rank test (two-tailed).

119 extent t h a t it was n o t significantly different in p e r i o d II1 from p e r i o d I. It m a y be c o n c l u d e d that k i n d l i n g s t i m u l a t i o n a p p l i e d to the d o r s a l h i p p o c a m p u s is sufficient to d i s r u p t the n o r m a l f u n c t i o n i n g o f b o t h h i p p o c a m p i in such a w a y that a deficit in W M a n d R M b e c o m e s a p p a r e n t . It s h o u l d be n o t e d that electrically induced h i p p o c a m p a l a f t e r d i s c h a r g e s result in the a c t i v a t i o n n o t o n l y o f the h i p p o c a m p a l f o r m a t i o n a n d septal nuclei b u t also o f a s s o c i a t e d areas such as the entorhinal cortex, a m y g d a l a , stria terminalis a n d its bed nucleus [16]. Therefore, it is likely that h i p p o c a m p a l kindling m a y p r o d u c e d i s r u p t i o n o f several limbic structures bey o n d the h i p p o c a m p u s . Each kindling stimulus m a y p r o d u c e a certain degree o f r e t r o g r a d e a m n e s i a [12]. The time between the b e h a v i o u r a l test a n d the tetanus was, however, never s h o r t e r than 45 min a n d usually I - 2 h, with the next b e h a v i o u r a l test c o m i n g 18-23 h later. Therefore, the findings presented here reflect a long-lasting d i s r u p t i o n o f processes involved in a spatial task that uses b o t h reference a n d w o r k i n g m e m o r y . W e should stress, however, that the task relied on spatial cues [7]. This implies that we c a n n o t , without explicit evidence, generalize o u r tindings to o t h e r m e m o r y tasks, which d o not involve such cues. Nevertheless, it will be o f p a r t i c u l a r interest to relate these findings to the clinical o b s e r v a t i o n that epileptic patients with limbic foci often have verbal m e m o r y deficits, a n d that i m p a i r m e n t s in a n u m b e r o f cognitive functions are c o r r e l a t e d with focal limbic e p i l e p t i f o r m activity [17]. In s u m m a r y , h i p p o c a m p u s kindling m a y be a g o o d m o d e l not only o f focal m o t o r seizures but also o f b e h a v i o u r a l deficits associated with epileptogenesis. The a u t h o r s w o u l d like to a c k n o w l e d g e the fruitful discussions with W. K a m p h u i s , B.P.C. Melchers a n d J.P.M. Pijn; technical assistance o f G . A . A d v o c a a t ; the statistical advice o f N. N a g e l k e r k e ; the stylistic revision o f M . A . Corner; a n d the secretarial assistance o f J.C. Cabi. I Ehlers, C.L. and Koob, G.F., Locomotor behaviour following kindling in three different brain sites, Brain Res., 326 (1985) 71 79. 2 George. G. and Mellanby, J., Memory deficits in an experimental hippocampal epileptiform syndrome in rats, Exp. Neurol., 75 (1982) 678-689. 3 Goddard. G.V., Development of epileptic seizures through brain stimulation at low intensity, Nature (London), 214 (1967) 102(~1021. 4 Jarrard, L.E., Selective hippocampal lesions and behaviour: effects of kainic acid lesions on perfl~rmance of place and cue tasks, Behav. Neurosci., 97 (1983) 873 889. 5 Jarrard, L.E., Okaichi, H., Steward, O. and Goldsmith, R.B., On the role of hippocampal connections in the performance of place and cue tasks: comparisons with damage to hippocampus, Behav. Neurosci., 98 (1984) 946-954. 6 Kamphuis, W.. Wadman, W.J. and Lopes da SilwL,F.H., Loss of inhibitory neurotransmitter 7-aminobutyric acid (GABA) in a kindling induced focus of epileptic activity, Neurosci. Lett., Suppl., 22 (1985) $383. 7 ()'Kcet'c. J. and NadeI, L., The Hippocampus as a Cognitive Map, Clarendon Press, Oxford, U.K.. 1978. 80lton, D.S., Becker, J.T. and Handelman, G.E., Hippocampus, space and memory, Behav. Brain Sci.. 2(1979) 313 365. 90lton, D.S., Walker, J.A. and Wolf, W.A., A disconnection analysis of hippocampal function, Brain Res., 233 (1982) 241 253.

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