Limbic Lesions and the Energizing, Aversive, and Inhibitory Effects of Non-Reward in Rats* PETER G. HENKE
St Francis Xavier University
ABSTRACT
Rats with bilateral lesions in the amygdala, septum, hippocampus, stria terminalis, and fornix were tested on a multiple reinforcement schedule in which barpressing in one component was associated with variable-interval reinforcement (S+) and the other component with extinction (S—). Responses on a second lever turned off S— for 5-sec periods during the extinction component. All groups, with the exception of animals with lesions in the amygdala, exhibited behavioral contrast. Rats with hippocampal or fornical lesions showed greater resistance to extinction. Response rates on the lever which turned off S— were higher after lesions in the septum and stria terminalis, whereas lower rates were obtained from rats with lesions in the amygdala. It was concluded that lesions in the amygdala attenuate the energizing and aversive effects of non-reward, lesions in the septal area and stria terminalis increase the aversive effects, and lesions in the hippocampus and fornix interfere with the inhibitory effects of non-reward.
Studies have shown that the limbic system determines certain responses to nonreward. The specific behavorial contributions of the various limbic structures, however, depend upon the ways in which the non-reward experience influences behavior. For example, we have found it helpful to categorize the behavorial effects of non-reward into: (1) energizing effects, i.e.,
the so-called frustration-effect (FE); (2) aversive effects, possibly analogous to punishment; and (3) response interference or inhibitory effects, i.e., extinction. Recent data suggest that these effects of nonreward are mediated by separate neural substrates in the limbic system. We have found that lesions in the amygdala eliminated the FE in a double-runway (Henke, 1977; Henke & Maxwell, 1973), but the lesion did not interfere with extinction performances after partial or continuous reinforcement (Henke, 1977; Henke & Bunnell, 1971). Similar studies of rats with septal lesions showed, however, that the FE was unimpaired (Henke, 1977; Mabry & Peeler, 1972) but extinction behavior was altered by the lesion (Gray, Quintao, & Araujo-Silva, 1972; Henke, 1974, 1977). In paradigms thought to measure the aversive effects of non-reward, rats with septal lesions pressed a lever that turned off a stimulus that had been associated with non-reward more frequently than did the controls. Similarly, the brain-damaged rats showed higher barpress performance when this response allowed the animal to escape from a compartment that was associated with non-reward. These results were interpreted to indicate that non-reward experiences were more aversive after the lesion (Dickinson, 1972; Henke, 1976). The hippocampus has also been implicated in controlling behavior following non-reward. In fact, it has been suggested that this limbic structure mediates Pavlovian inhibition (Kimble, 1968) or registers non-reward events (Douglas & Pribram, 1966). For example, animals with lesions in the hippocampus were found to be deficient in extinction behavior (Henke & Bunnell, 1971; Jarrard, Isaacson, & Wickelgren, 1964) but also showed a normal FE in a double runway apparatus (Swanson & Isaacson, 1969).
•Reprint requests should be sent to Peter G. Henke, St Francis Xavier University, Antigonish, Nova Scotia, Canada, B2G iCo. This research was supported by grants from the National Research Council and the Medical Research Council of Canada.
Canad. J. Psychol./Rev. canad. Psychol., 1979,33 (3)
133
The present study investigated the effects of lesions in the amygdala, septum, and hippocampus on the behavioral consequences of non-reward, i.e., the energizing, aversive, and inhibitory effects, simultaneously in the same animals. In addition, groups of rats with lesions in some of the fiber pathways connecting these structures, i.e., the dorsal fornix system and the stria terminalis, were also included. The behavioral task was a multiple reinforcement schedule that allowed withinsession tests of the presumed energizing effects (behavioral contrast), extinction performance, and escape responses from S—. It has been argued that contrast is due to the emotional or frustrative effects of non-reinforcement (Skull, Davies, & Amsel, 1970; Terrace, 1972). Behaviorally, it refers to opposing response changes after switching one component of a multiple reinforcement schedule to extinction conditions. Typically, response rates increase in the unaltered component (S+) as responding decreases in the extinction component (S—) (e.g., Henke, Allen, & Davison, 1972; Reynolds, 1961). During the extinction condition, presses on a second lever turned off S— for 5-sec periods. METHOD
Subjects
Thirty-six male albino rats of Wistar descent (Woodlyn Laboratories, Guelph, Ontario), 90 days old at the beginning of the experiment, were randomly assigned to six groups. Bilateral lesions were performed in the amygdala, septum, hippocampus, stria terminalis, and dorsal fornix. The control group consisted of one unoperated and five sham-operated rats. Surgery and Histology
Radio-frequency (100 kHz) lesions were produced while the rat was anesthetized with 50 mg/kg of sodium pentobarbital. The current was applied for 20 sec through the uninsulated tip of a stainless steel electrode. The circuit was closed with a rectal electrode. The coordinates for septal lesions were 1.5 mm anterior to bregma, .5 mm lateral to the midline, and 5.5 mm ventral to the surface of the skull (skull
134
horizontal). Multiple lesions in the amygdala were produced at .9 and . 1 mm posterior to bregma, 4.75 and 4.25 mm lateral to the midline, and g.o and 8.8 mm ventral to the surface of the dura (incisor bar raised 5.00 mm above the interaural line). For lesions in the stria terminalis the electrode was placed .5 mm posterior to bregma, 1.00 mm lateral to the midline, and 6.00 mm ventral to the dura (skull horizontal); for lesions in the fornix it was positioned 1.00 mm posterior to bregma, .5 and .8 mm lateral, and 4.00 mm ventral from the skull surface (skull horizontal). Hippocampal lesions were done by aspiration. The cortex overlying the hippocampus was removed and the hippocampus was then aspirated under visual guidance through a microscope. Control operations were similar to that of their corresponding experimental groups, except that the electrode was not lowered into the brain. In one animal the cortex overlying the hippocampus was removed by aspiration. After testing, the brain-damaged rats were perfused intracardially with saline and 10% formalin. After embedding in paraffin, sections were cut at 18/u.. Every fifth section was stained with thionin. Additional sections were stained according to Woelcke's or Weil's methods for myelin sheath. Apparatus
Two-lever operant conditioning boxes (BRS/LVE) were housed in sound-attenuating enclosures. The right-hand lever was always extended into the apparatus, whereas the left-hand lever was only inserted during the discrimination phase of training. Right-hand lever presses provided reinforcements (45 mg Noyes pellets) during the prediscrimination phase and during the S + (cue light on) component in discrimination training. Presses on the left-hand lever turned off S— (white noise) for 5-sec periods. Procedure
Following a 2-wk recovery period for operated rats, the animals were gradually reduced to 80% of their free-feeding body weights. They were maintained at this weight level throughout the study. After shaping of the lever press response, the rats received 100 reinforcements on a continuous reinforcement schedule. The next day, a variable-interval (vi) of 30 sec schedule was in effect for a l-hr session. During the subsequent l-hr testing sessions (prediscrimination phase) reinforcements were provided on a vi-i min schedule. Throughout the 20 prediscrimination sessions a white cue light, immediately above the
P.G. Henke
FIGURE i Drawings of representative lesions in the amygdala (A), septum (S), stria terminalis (ST), fornix (F), and hippocampus (H).
reward lever, was on (S+). The left-hand lever was in the retracted position during prediscrimination training. During the 15-day discrimination phase (1-hr sessions), a probability generator randomly alternated 2-min S+ components with a-min S— components. S— was a white noise stimulus at 70 db, re .0002 dyne/cm2, measured in the vicinity of the right-hand lever. During the S— component no reinforcements were delivered, but responses on the left-hand lever turned off the white noise for 5-sec periods. RESULTS Anatomical Data
Drawings of representative lesions are shown in Figure 1. Lesions in the amygdaloid complex were relatively extensive. The greatest amount of destruction was Limbic lesions and non-reward
seen in the cortical, lateral basal, and medial basal nuclei. Moderate damage was found in the anterior lateral, posterior lateral, central, and medial nuclei. In four animals, damage was also present in the pyriform cortex, and one rat sustained slight unilateral damage in the optic tract. Septal lesions were also large, in all cases. The medial and lateral septal areas were almost completely destroyed. The diagonal band, its nucleus at the level of the anterior commissure, and the hippocampal rudiment also sustained damage. Incidental damage to surrounding tissue, including the corpus callosum and caudate-putamen, was generally slight and not related to behavioral performance. Examinations of the brains of hippocam-
135
Mean responses/minute during the prediscrimination baseline and three-day discrimination blocks Baseline
Block 1
Block 2
Block 3
Block 4
Block 5
15.4 10.9 12.1 11.3
20.3 7.6 11.8 8.2
19.4 5.2 12.2 5.9
20.0 2.8 15.4 3.5
21.2 2.1 14.2 1.9
Controls
VI EXT
15.1
Amygdala
VI EXT
17.2
Septum
VI EXT
29.6
31.4 18.1
45.7 15.2
37.5 18.4
40.3 6.9
48.2 6.6
Hippocampus
VI EXT
16.8
15.1 15.7
21.6 12.7
20.1 10.2
21.9 5.9
21.0 4.7
Stria Terminalis
VI EXT
17.1
17.7 11.9
20.2 9.6
21.9 7.2
19.9 5.1
20.8 4.5
Fornix
VI EXT
27.5
30.9 24.9
32.7 22.9
35.1 17.5
36.9 12.3
34.4 9.8
pal subjects indicated that the hippocampus was not completely destroyed. The lesions varied from approximately 50% to 80% destruction. The most rostral portion, as well as the ventral tip of the hippocampus, escaped major damage. In two rats incidental damage to lateral thalamic areas was also found. However, these animals did not differ in any systematic fashion from the other hippocampal rats and were included in the data analysis. The amount of neocortex removed was similar across all hippocampal animals. The damage found in the cortical control was within the range seen in the hippocampal rats. Lesions in the stria terminalis were found to be most extensive at the level of the anterior commissure. The precommissural and postcommissural components were cut in all cases. The bed nucleus of the stria terminalis was damaged in all animals, as well. Slight damage to the anterior commissure and the stria medullaris was seen in four rats. Lesions in the fornix bundle interrupted the dorsal fornix fibers and the fimbrial system. The dorsal and ventral fornical (hippocampal) commissures and the triangular septal nucleus were also damaged in the rats. The subfornical organ was damaged in three animals. There was no dam136
age to the stria terminalis or stria medullaris. Behavioral Data
In agreement with previous findings (Gaffan, 1973; Henke, 1976), rats with lesions in the septal area and fornix responded at higher rates than the controls. Table 1 shows that animals with septal and fornical lesions reached higher prediscrimination baseline rates. An analysis of variance of the baseline data produced a significant lesion effect, ^(5, 30) = 5.01,p < .01. In order to facilitate between-groups comparisons during the multiple schedule condition, taking into account the unequal response baselines, the following transformation was used. In Figure 2, Period o represents as unity the mean barpress rates over the last three prediscrimination sessions. The discrimination response change ratios during successive three-session blocks were then computed by dividing the mean response rate of each rat, during the separate S+ and S— components, by the rat's mean baseline rate. The group mean ratios were then computed and plotted over Periods 1-5. Figure 2 shows that during the S+ component all groups increased responding relative to baselines, with the exception of P.G. Henke
1.75 A
/
1.50 -A 1.25 -
A
Q 1.00 _
\
>—
—•
%
.75
6
a.
.50 s« sc o— o — o •— • - - - • .25 _ s a H k STO—D—O A
I
I
0
1
-A
1 2
I
1
1
3
4
5
PERIOD
FIGURE 2 Response change ratios during S+ and S— of control rats (C) and rats with lesions in the amygdala (A), septum (S), hippocampus (H), stria terminalis (ST), and fornix (F). See text for details.
animals with lesions in the amygdala. In fact, inspection of Figure 2 indicates that rats with brain-damage in the amygdala reduced responding in S+ (negative induction) but their responding recovered to baseline levels during Periods 4 and 5. During the S— component (no reinforcements) all groups reduced responding relative to baselines. Figure 2 indicates a slower decline in responding for the rats with lesions in the hippocampus and fornix. The response rate ratios were analysed using a three-factor mixed-design analysis of variance with lesions as the betweensubjects variable and reinforcement condition and session blocks as the within-subjects variables. It produced a significant effect of lesion, F( 5 , 3 o) = 3.57, p < .05, reinforcement, F(i,$o) = 46.71,/? < .01, and sessions, F(4, 120) = 6.75,/) < .01. The Lesion X Reinforcement, F(5,30) = 9.29,p < .01, Lesion X Session, F(2O, 120) = 2.59,
Limbic lesions and non-reward
8 10 SESSION
FIGURE 3 Left-hand lever responses per S— component for control rats (C) and rats with lesions in the amygdala (A), septum (S), hippocampus (H), stria terminalis (ST), and fornix (F).
p < .01, Reinforcement x Session, ^"(4,120) = 62.08, p < .01, and triple interaction F(20,120) = 4.59,/* < .01, were also statistically significant. Additional comparisons of the control group with the groups of brain-damaged rats, using Dunnet's test (Winer, 1962, p. 89), showed that the animals with amygdala damage responded at significantly lower rates than the controls during S+ (p < .05). During the extinction component, Dunnet's test showed that both the group with hippocampal lesions and the group with fornical lesions responded at higher rates than the controls (Periods 2-5, p .10. Dunnet's test showed that, relative
137
to controls, rats with lesions in the septum and stria terminalis responded at higher rates during Sessions 2—14. Animals with lesions in the amygdala responded at reduced levels during Sessions 6—14. DISCUSSION
The present results show that, with the exception of animals with lesions in the amygdala, the remaining groups of rats exhibited behavioral contrast. It has been suggested that this increase in response rate is due to the emotional or frustrating consequences of non-reward (Skull et al., 1970; Terrace, 1972). This interpretation is supported to the extent that lesions in the amygdala eliminate both contrast effects and frustration effects (Henke, 1973; Henke & Maxwell, 1973; Henke et al., 1972). Figure 2 indicates that lesions in the stria terminalis did not significandy alter behavioral contrast, suggesting that this fiber pathway is not critically involved in the energizing effects of non-reward. The data in Table 1 show that the absolute response levels of animals with septal and fornical lesions were higher than those of the controls. However, as indicated in Figure 2, the relative magnitude of contrast was not changed by the lesions. The proposal has been made that septal lesions produce greater response output during positive reward conditions as the result of abnormally high incentive-motivation, i.e., the conditioned motivation to obtain rewards (Henke, 1975, 1977), or stronger approach tendencies (Fried, 1972). Based on the present data, these interpretations may be extended to account for the increase in the rate of responding after lesions in the fornix. In other words, it may be argued that the observed effects of lesions in the fornix system might be due to the cutting of fibres which have a septal origin. One possible explanation might be that these fibers normally transmit information about reductions in incentive-motivation, e.g., during extinction (cf. Henke, 1975).
138
The present results show that during the extinction component of the multiple schedule the rats with lesions in the septum, amygdala, and stria terminalis responded at similar relative rates. Lesions in the hippocampus and fornix, however, retarded the extinction behavior (Fig. 2). These findings are interpreted to indicate that the normal learning of not to respond, possibly the result of the anticipation of non-reward (cf. Bolles, 1972), requires an intact hippocampus. Furthermore, the data suggest that the fornix contains fibers which are involved in this process. When absolute response rates are considered (Table 1), the data show that rats with septal lesions emitted more responses in extinction than the controls. On the other hand, as pointed out previously, adjustments for a greater overall response output following the lesions 'removed' these response differences (cf. Henke, 1975; Ross, Grossman, & Grossman, 1975). In line with the 'incentive-model' of septal lesion effects, these data also fit the idea of an abnormal increase of incentivemotivation after septal damage. More specifically, the argument has been made that extinction conditions not only produce response interference due to an anticipation of non-reward (a cognitive process) but also lead to the extinction of incentivemotivation which had been acquired during acquisition (Henke, 1975). The analysis of the escape responses from S— shows that lesions in the septum and the stria terminalis produced an increase in this behavior. Conversely, lesions in the amygdala reduced the escape performance (Fig. 3). These findings indicate that an amygdala-septum axis, through fibers in the stria terminalis, influence responses to the aversive effects of nonreward. Anatomically, such a system might include fibers originating in the medial septal nuclei which then project through the stria terminalis to the basolateral amygdala (Raisman, 1966). Data by Gray and his co-workers have, in fact, implicated the
P.G. Henke
medial septal area in responses to nonreward (Gray, 1972; Gray et al., 1972). It is possible that the presumed reciprocal relationship for 'emotionality' between amygdala and septum (King & Meyer, 1958; Schwartzbaum & Gay, 1966) also exists in regard to aversive non-reward. The incentive-model of septal lesion effects predicts that non-reward experiences are more aversive because of greater expectations of reward. The possibility that septal lesions altered the responsiveness to the white noise stimulus itself was not supported, however, by the findings of a previous study which measured the unconditioned escape responses to the same stimulus. No significant differences between septal and control rats were found (Henke, 1976). Figure 3 shows that barpress responses which turned off S— were significantly reduced following lesions in the amygdala. Wagner (1966) has suggested that the withholding of expected reinforcements is functionally equivalent to other forms of punishment. Based on this interpretation it may be concluded that lesions in the amygdala greatly attenuate the punishing effect of non-reward. In fact, Wagner (ig66) has also asserted that the increase in running behavior in the second alley of a double runway following non-reward in the first goal box, i.e., the FE, may actually be due to an addition of escape responses from the aversive non-reward situation. This analysis also fits the finding that amygdala lesions virtually eliminate the FE in a double runway (Henke & Maxwell, 1973). In summary, it is concluded that the affective responses following non-reward experiences, i.e., the aversive and energizing effects, are related to processes in the amygdala. It is suggested that the inhibitory influences from the septal area, through fibers of the stria terminalis, normally mediate the aversive effects of non-reward relative to current levels of reward expectations. A cognitive mechanism, normally used to learn to anticipate non-reward, is Limbic lesions and non-reward
associated with the hippocampus and fornix system. In other words, the hippocampus is assumed to be important when nonreward inhibits approach responses on which it was made contingent. In addition, the fornix also contains fibers which transmit information about reductions in incentive-motivation. This is suggested to occur following various manipulations to reduce reward availability, e.g., during extinction. RESUME
Des rats ayant subi des lesions bilaterales de l'amygdale, du septum, de l'hyppocampe, de la stria terminalis et du fornix sont soumis a un regime multiple de renforcement dans lequel la pression d'un levier, dans l'une des composantes, est associee a un renforcement (S+) a intervalles variables et la seconde composante associee a l'extinction (S—). Les reponses liees a un second levier eliminent le S— pendant des periodes de 5s au cours de l'extinction. Tous les groupes, exception faite des animaux ayant subi des lesions dans l'amygdale, manifestent un contraste comportemental. Les rats ayant subi des lesions dans l'hyppocampe ou dans le fornix montrent plus de resistance a l'extinction. Le taux des reponses liees au levier qui elimine le S— est plus eleve quand les lesions affectent le septum et la stria terminalis que quand elles affectent l'amygdale. La conclusion affirme que les lesions de l'amygdale attenuent les effets dynamiques et aversifs du non-renforcement, les lesions de l'aire septale et de la stria terminalis augmentent les effets aversifs, et les lesions de l'hyppocampe et du fornix interferent avec les effets inhibiteurs du non-renforcement. REFERENCES BOLLES, R.c. Reinforcement, expectancy, and learning. Psychol. Rev., 1972, 79, 394—409 DICKINSON, A. Septal damage and response output under frustrative non-reward. In R.A. BOAKES and M.S. HALLIDAY (Eds.), Inhibition and learning. London: Academic Press, 1972 DOUGLAS, R.J., PRIBRAM, K.H. Learning and limbic le-
sions. Neuropsycholog., 1966,4, 197-220 FRIED, P.A. The septum and behavior: A review. Psychol. Bull. 1972,78,292—310 GAFFAN, D. Inhibitory gradients and behavioral contrast in rats with lesions in the fornix. Physiol. Behav., 1973,ii,215—220 GRAY.J.A. Effects of septal driving of the hippocampal
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theta rhythm on resistance to extinction. Physiol. Behav., 1972, 8,481—490 GRAY.J.A., QUINTAO, L., & ARAUJO-SILVA, M.T. The partial reinforcement extinction effect in rats with medial septal lesions. Physiol. Behav., 1972, 8,491-496 HENKE, p.G. Effects of reinforcement omission on rats with lesions in the amygdala./, comp. physiol. Psychol., 1973. 8 4. 187-193 HENKE, p.G. Persistence of runway performance after septal lesions in rats.J. comp. physiol. Psychol., 1974, 86, 760-767 HENKE, p.G. Septal lesions and the extinction of incentive-motivation. Physiol. Behav., 1975, 15, 537-542 HENKE, p.G. Septal lesions and aversive nonreward. Physiol. Behav., 1976,17,483-488 HENKE, P.G. Dissociation of the frustration effect and the partial reinforcement extinction effect after limbic lesions in rats./, comp. physiol. Psychol., 1977, 91, 1032-1038 HENKE, P.G., ALLEN, J.D., & DAVISON, c. Effects of lesions
in the amygdala on behavioral contrast. Physiol. Behav., 1972,8, 173—176 HENKE, P.G., & BUNNELL, B.N. Reinforcement and ex-
tinction interactions after limbic lesions in rats. Communic. behav. Biol., Part A, 1971,6, 329—333 HENKE, P.G., & MAXWELL, D. Lesions in the amygdala
and the frustration effect. Physiol. Behav., 1973, 10, 647-650 JARRARD, L.E., ISAACSON, R.L., & WICKELGREN, W.O. Ef-
fects of hippocampal ablation and intertrial interval on running acquisition and extinction./, comp. physiol. Psychol., 1964, 57,442—444 KIMBLE, D.P. Hippocampus and internal inhibition. Psychol. Bull., 1968, 70, 285-295 KING, F.A., & MEYER, P.M. Effects of amygdaloid lesions
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upon septal hyperemotionality in the rat. Science, 1958, 1*8, 655-656 MABRY, P.D., & PEELER, D.F. Effect of septal lesions on response to frustrative non-reward. Physiol. Behav., 1972,8,909-913 RAISMAN, G. The connexions of the septum. Brain, 1966,89,317-348 REYNOLDS, G.s. Behavioral contrast./, exp. Anal. Behav., 1961,4,57-71 ROSS, J.F., GROSSMAN, L., 8c GROSSMAN, s.p. Some be-
havioral effects of transecting ventral or dorsal fiber connections of the septum in the rat./, comp. physiol. Psychol., 1975,89,5-19 SCHWARTZBAUM, j.s., & GAY, P.E. Interacting behavioral effects of septal and amygdaloid lesions in the rat./. comp. physiol. Psychol., 1966, 61, 59-65 SKULL, j . DA VIES, K., & AMSEL, A. Behavioral contrast and
frustration effect in multiple and mixed fixedinterval schedules in the rat./, comp. physiol. Psychol., 1970,71,478-483 SWANSON, A.M., & ISAACSON, R.L. Hippocampal lesions
and the frustration effect in rats./, comp. physiol. Psychol., 1969, 68, 562—567 TERRACE, H.S. Conditioned inhibition in successive discrimination learning. In R.A. BOAKES and M.S. HALLI-
DAY (Eds.), Inhibition and learning. London: Academic Press, 1972 WAGNER, A.R. Frustration and punishment. In R.N. HABER (Ed.), Current research in motivation. New York: Holt, Rinehait and Winston, 1966 WINER, B.J. Statistical principles in experimental design. New York: McGraw-Hill, 1962 (First received 10 November 1978) (Date accepted 8 March 1979)
P.G. Henke
St Francis Xavier University
ABSTRACT
Rats with bilateral lesions in the amygdala, septum, hippocampus, stria terminalis, and fornix were tested on a multiple reinforcement schedule in which barpressing in one component was associated with variable-interval reinforcement (S+) and the other component with extinction (S—). Responses on a second lever turned off S— for 5-sec periods during the extinction component. All groups, with the exception of animals with lesions in the amygdala, exhibited behavioral contrast. Rats with hippocampal or fornical lesions showed greater resistance to extinction. Response rates on the lever which turned off S— were higher after lesions in the septum and stria terminalis, whereas lower rates were obtained from rats with lesions in the amygdala. It was concluded that lesions in the amygdala attenuate the energizing and aversive effects of non-reward, lesions in the septal area and stria terminalis increase the aversive effects, and lesions in the hippocampus and fornix interfere with the inhibitory effects of non-reward.
Studies have shown that the limbic system determines certain responses to nonreward. The specific behavorial contributions of the various limbic structures, however, depend upon the ways in which the non-reward experience influences behavior. For example, we have found it helpful to categorize the behavorial effects of non-reward into: (1) energizing effects, i.e.,
the so-called frustration-effect (FE); (2) aversive effects, possibly analogous to punishment; and (3) response interference or inhibitory effects, i.e., extinction. Recent data suggest that these effects of nonreward are mediated by separate neural substrates in the limbic system. We have found that lesions in the amygdala eliminated the FE in a double-runway (Henke, 1977; Henke & Maxwell, 1973), but the lesion did not interfere with extinction performances after partial or continuous reinforcement (Henke, 1977; Henke & Bunnell, 1971). Similar studies of rats with septal lesions showed, however, that the FE was unimpaired (Henke, 1977; Mabry & Peeler, 1972) but extinction behavior was altered by the lesion (Gray, Quintao, & Araujo-Silva, 1972; Henke, 1974, 1977). In paradigms thought to measure the aversive effects of non-reward, rats with septal lesions pressed a lever that turned off a stimulus that had been associated with non-reward more frequently than did the controls. Similarly, the brain-damaged rats showed higher barpress performance when this response allowed the animal to escape from a compartment that was associated with non-reward. These results were interpreted to indicate that non-reward experiences were more aversive after the lesion (Dickinson, 1972; Henke, 1976). The hippocampus has also been implicated in controlling behavior following non-reward. In fact, it has been suggested that this limbic structure mediates Pavlovian inhibition (Kimble, 1968) or registers non-reward events (Douglas & Pribram, 1966). For example, animals with lesions in the hippocampus were found to be deficient in extinction behavior (Henke & Bunnell, 1971; Jarrard, Isaacson, & Wickelgren, 1964) but also showed a normal FE in a double runway apparatus (Swanson & Isaacson, 1969).
•Reprint requests should be sent to Peter G. Henke, St Francis Xavier University, Antigonish, Nova Scotia, Canada, B2G iCo. This research was supported by grants from the National Research Council and the Medical Research Council of Canada.
Canad. J. Psychol./Rev. canad. Psychol., 1979,33 (3)
133
The present study investigated the effects of lesions in the amygdala, septum, and hippocampus on the behavioral consequences of non-reward, i.e., the energizing, aversive, and inhibitory effects, simultaneously in the same animals. In addition, groups of rats with lesions in some of the fiber pathways connecting these structures, i.e., the dorsal fornix system and the stria terminalis, were also included. The behavioral task was a multiple reinforcement schedule that allowed withinsession tests of the presumed energizing effects (behavioral contrast), extinction performance, and escape responses from S—. It has been argued that contrast is due to the emotional or frustrative effects of non-reinforcement (Skull, Davies, & Amsel, 1970; Terrace, 1972). Behaviorally, it refers to opposing response changes after switching one component of a multiple reinforcement schedule to extinction conditions. Typically, response rates increase in the unaltered component (S+) as responding decreases in the extinction component (S—) (e.g., Henke, Allen, & Davison, 1972; Reynolds, 1961). During the extinction condition, presses on a second lever turned off S— for 5-sec periods. METHOD
Subjects
Thirty-six male albino rats of Wistar descent (Woodlyn Laboratories, Guelph, Ontario), 90 days old at the beginning of the experiment, were randomly assigned to six groups. Bilateral lesions were performed in the amygdala, septum, hippocampus, stria terminalis, and dorsal fornix. The control group consisted of one unoperated and five sham-operated rats. Surgery and Histology
Radio-frequency (100 kHz) lesions were produced while the rat was anesthetized with 50 mg/kg of sodium pentobarbital. The current was applied for 20 sec through the uninsulated tip of a stainless steel electrode. The circuit was closed with a rectal electrode. The coordinates for septal lesions were 1.5 mm anterior to bregma, .5 mm lateral to the midline, and 5.5 mm ventral to the surface of the skull (skull
134
horizontal). Multiple lesions in the amygdala were produced at .9 and . 1 mm posterior to bregma, 4.75 and 4.25 mm lateral to the midline, and g.o and 8.8 mm ventral to the surface of the dura (incisor bar raised 5.00 mm above the interaural line). For lesions in the stria terminalis the electrode was placed .5 mm posterior to bregma, 1.00 mm lateral to the midline, and 6.00 mm ventral to the dura (skull horizontal); for lesions in the fornix it was positioned 1.00 mm posterior to bregma, .5 and .8 mm lateral, and 4.00 mm ventral from the skull surface (skull horizontal). Hippocampal lesions were done by aspiration. The cortex overlying the hippocampus was removed and the hippocampus was then aspirated under visual guidance through a microscope. Control operations were similar to that of their corresponding experimental groups, except that the electrode was not lowered into the brain. In one animal the cortex overlying the hippocampus was removed by aspiration. After testing, the brain-damaged rats were perfused intracardially with saline and 10% formalin. After embedding in paraffin, sections were cut at 18/u.. Every fifth section was stained with thionin. Additional sections were stained according to Woelcke's or Weil's methods for myelin sheath. Apparatus
Two-lever operant conditioning boxes (BRS/LVE) were housed in sound-attenuating enclosures. The right-hand lever was always extended into the apparatus, whereas the left-hand lever was only inserted during the discrimination phase of training. Right-hand lever presses provided reinforcements (45 mg Noyes pellets) during the prediscrimination phase and during the S + (cue light on) component in discrimination training. Presses on the left-hand lever turned off S— (white noise) for 5-sec periods. Procedure
Following a 2-wk recovery period for operated rats, the animals were gradually reduced to 80% of their free-feeding body weights. They were maintained at this weight level throughout the study. After shaping of the lever press response, the rats received 100 reinforcements on a continuous reinforcement schedule. The next day, a variable-interval (vi) of 30 sec schedule was in effect for a l-hr session. During the subsequent l-hr testing sessions (prediscrimination phase) reinforcements were provided on a vi-i min schedule. Throughout the 20 prediscrimination sessions a white cue light, immediately above the
P.G. Henke
FIGURE i Drawings of representative lesions in the amygdala (A), septum (S), stria terminalis (ST), fornix (F), and hippocampus (H).
reward lever, was on (S+). The left-hand lever was in the retracted position during prediscrimination training. During the 15-day discrimination phase (1-hr sessions), a probability generator randomly alternated 2-min S+ components with a-min S— components. S— was a white noise stimulus at 70 db, re .0002 dyne/cm2, measured in the vicinity of the right-hand lever. During the S— component no reinforcements were delivered, but responses on the left-hand lever turned off the white noise for 5-sec periods. RESULTS Anatomical Data
Drawings of representative lesions are shown in Figure 1. Lesions in the amygdaloid complex were relatively extensive. The greatest amount of destruction was Limbic lesions and non-reward
seen in the cortical, lateral basal, and medial basal nuclei. Moderate damage was found in the anterior lateral, posterior lateral, central, and medial nuclei. In four animals, damage was also present in the pyriform cortex, and one rat sustained slight unilateral damage in the optic tract. Septal lesions were also large, in all cases. The medial and lateral septal areas were almost completely destroyed. The diagonal band, its nucleus at the level of the anterior commissure, and the hippocampal rudiment also sustained damage. Incidental damage to surrounding tissue, including the corpus callosum and caudate-putamen, was generally slight and not related to behavioral performance. Examinations of the brains of hippocam-
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Mean responses/minute during the prediscrimination baseline and three-day discrimination blocks Baseline
Block 1
Block 2
Block 3
Block 4
Block 5
15.4 10.9 12.1 11.3
20.3 7.6 11.8 8.2
19.4 5.2 12.2 5.9
20.0 2.8 15.4 3.5
21.2 2.1 14.2 1.9
Controls
VI EXT
15.1
Amygdala
VI EXT
17.2
Septum
VI EXT
29.6
31.4 18.1
45.7 15.2
37.5 18.4
40.3 6.9
48.2 6.6
Hippocampus
VI EXT
16.8
15.1 15.7
21.6 12.7
20.1 10.2
21.9 5.9
21.0 4.7
Stria Terminalis
VI EXT
17.1
17.7 11.9
20.2 9.6
21.9 7.2
19.9 5.1
20.8 4.5
Fornix
VI EXT
27.5
30.9 24.9
32.7 22.9
35.1 17.5
36.9 12.3
34.4 9.8
pal subjects indicated that the hippocampus was not completely destroyed. The lesions varied from approximately 50% to 80% destruction. The most rostral portion, as well as the ventral tip of the hippocampus, escaped major damage. In two rats incidental damage to lateral thalamic areas was also found. However, these animals did not differ in any systematic fashion from the other hippocampal rats and were included in the data analysis. The amount of neocortex removed was similar across all hippocampal animals. The damage found in the cortical control was within the range seen in the hippocampal rats. Lesions in the stria terminalis were found to be most extensive at the level of the anterior commissure. The precommissural and postcommissural components were cut in all cases. The bed nucleus of the stria terminalis was damaged in all animals, as well. Slight damage to the anterior commissure and the stria medullaris was seen in four rats. Lesions in the fornix bundle interrupted the dorsal fornix fibers and the fimbrial system. The dorsal and ventral fornical (hippocampal) commissures and the triangular septal nucleus were also damaged in the rats. The subfornical organ was damaged in three animals. There was no dam136
age to the stria terminalis or stria medullaris. Behavioral Data
In agreement with previous findings (Gaffan, 1973; Henke, 1976), rats with lesions in the septal area and fornix responded at higher rates than the controls. Table 1 shows that animals with septal and fornical lesions reached higher prediscrimination baseline rates. An analysis of variance of the baseline data produced a significant lesion effect, ^(5, 30) = 5.01,p < .01. In order to facilitate between-groups comparisons during the multiple schedule condition, taking into account the unequal response baselines, the following transformation was used. In Figure 2, Period o represents as unity the mean barpress rates over the last three prediscrimination sessions. The discrimination response change ratios during successive three-session blocks were then computed by dividing the mean response rate of each rat, during the separate S+ and S— components, by the rat's mean baseline rate. The group mean ratios were then computed and plotted over Periods 1-5. Figure 2 shows that during the S+ component all groups increased responding relative to baselines, with the exception of P.G. Henke
1.75 A
/
1.50 -A 1.25 -
A
Q 1.00 _
\
>—
—•
%
.75
6
a.
.50 s« sc o— o — o •— • - - - • .25 _ s a H k STO—D—O A
I
I
0
1
-A
1 2
I
1
1
3
4
5
PERIOD
FIGURE 2 Response change ratios during S+ and S— of control rats (C) and rats with lesions in the amygdala (A), septum (S), hippocampus (H), stria terminalis (ST), and fornix (F). See text for details.
animals with lesions in the amygdala. In fact, inspection of Figure 2 indicates that rats with brain-damage in the amygdala reduced responding in S+ (negative induction) but their responding recovered to baseline levels during Periods 4 and 5. During the S— component (no reinforcements) all groups reduced responding relative to baselines. Figure 2 indicates a slower decline in responding for the rats with lesions in the hippocampus and fornix. The response rate ratios were analysed using a three-factor mixed-design analysis of variance with lesions as the betweensubjects variable and reinforcement condition and session blocks as the within-subjects variables. It produced a significant effect of lesion, F( 5 , 3 o) = 3.57, p < .05, reinforcement, F(i,$o) = 46.71,/? < .01, and sessions, F(4, 120) = 6.75,/) < .01. The Lesion X Reinforcement, F(5,30) = 9.29,p < .01, Lesion X Session, F(2O, 120) = 2.59,
Limbic lesions and non-reward
8 10 SESSION
FIGURE 3 Left-hand lever responses per S— component for control rats (C) and rats with lesions in the amygdala (A), septum (S), hippocampus (H), stria terminalis (ST), and fornix (F).
p < .01, Reinforcement x Session, ^"(4,120) = 62.08, p < .01, and triple interaction F(20,120) = 4.59,/* < .01, were also statistically significant. Additional comparisons of the control group with the groups of brain-damaged rats, using Dunnet's test (Winer, 1962, p. 89), showed that the animals with amygdala damage responded at significantly lower rates than the controls during S+ (p < .05). During the extinction component, Dunnet's test showed that both the group with hippocampal lesions and the group with fornical lesions responded at higher rates than the controls (Periods 2-5, p .10. Dunnet's test showed that, relative
137
to controls, rats with lesions in the septum and stria terminalis responded at higher rates during Sessions 2—14. Animals with lesions in the amygdala responded at reduced levels during Sessions 6—14. DISCUSSION
The present results show that, with the exception of animals with lesions in the amygdala, the remaining groups of rats exhibited behavioral contrast. It has been suggested that this increase in response rate is due to the emotional or frustrating consequences of non-reward (Skull et al., 1970; Terrace, 1972). This interpretation is supported to the extent that lesions in the amygdala eliminate both contrast effects and frustration effects (Henke, 1973; Henke & Maxwell, 1973; Henke et al., 1972). Figure 2 indicates that lesions in the stria terminalis did not significandy alter behavioral contrast, suggesting that this fiber pathway is not critically involved in the energizing effects of non-reward. The data in Table 1 show that the absolute response levels of animals with septal and fornical lesions were higher than those of the controls. However, as indicated in Figure 2, the relative magnitude of contrast was not changed by the lesions. The proposal has been made that septal lesions produce greater response output during positive reward conditions as the result of abnormally high incentive-motivation, i.e., the conditioned motivation to obtain rewards (Henke, 1975, 1977), or stronger approach tendencies (Fried, 1972). Based on the present data, these interpretations may be extended to account for the increase in the rate of responding after lesions in the fornix. In other words, it may be argued that the observed effects of lesions in the fornix system might be due to the cutting of fibres which have a septal origin. One possible explanation might be that these fibers normally transmit information about reductions in incentive-motivation, e.g., during extinction (cf. Henke, 1975).
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The present results show that during the extinction component of the multiple schedule the rats with lesions in the septum, amygdala, and stria terminalis responded at similar relative rates. Lesions in the hippocampus and fornix, however, retarded the extinction behavior (Fig. 2). These findings are interpreted to indicate that the normal learning of not to respond, possibly the result of the anticipation of non-reward (cf. Bolles, 1972), requires an intact hippocampus. Furthermore, the data suggest that the fornix contains fibers which are involved in this process. When absolute response rates are considered (Table 1), the data show that rats with septal lesions emitted more responses in extinction than the controls. On the other hand, as pointed out previously, adjustments for a greater overall response output following the lesions 'removed' these response differences (cf. Henke, 1975; Ross, Grossman, & Grossman, 1975). In line with the 'incentive-model' of septal lesion effects, these data also fit the idea of an abnormal increase of incentivemotivation after septal damage. More specifically, the argument has been made that extinction conditions not only produce response interference due to an anticipation of non-reward (a cognitive process) but also lead to the extinction of incentivemotivation which had been acquired during acquisition (Henke, 1975). The analysis of the escape responses from S— shows that lesions in the septum and the stria terminalis produced an increase in this behavior. Conversely, lesions in the amygdala reduced the escape performance (Fig. 3). These findings indicate that an amygdala-septum axis, through fibers in the stria terminalis, influence responses to the aversive effects of nonreward. Anatomically, such a system might include fibers originating in the medial septal nuclei which then project through the stria terminalis to the basolateral amygdala (Raisman, 1966). Data by Gray and his co-workers have, in fact, implicated the
P.G. Henke
medial septal area in responses to nonreward (Gray, 1972; Gray et al., 1972). It is possible that the presumed reciprocal relationship for 'emotionality' between amygdala and septum (King & Meyer, 1958; Schwartzbaum & Gay, 1966) also exists in regard to aversive non-reward. The incentive-model of septal lesion effects predicts that non-reward experiences are more aversive because of greater expectations of reward. The possibility that septal lesions altered the responsiveness to the white noise stimulus itself was not supported, however, by the findings of a previous study which measured the unconditioned escape responses to the same stimulus. No significant differences between septal and control rats were found (Henke, 1976). Figure 3 shows that barpress responses which turned off S— were significantly reduced following lesions in the amygdala. Wagner (1966) has suggested that the withholding of expected reinforcements is functionally equivalent to other forms of punishment. Based on this interpretation it may be concluded that lesions in the amygdala greatly attenuate the punishing effect of non-reward. In fact, Wagner (ig66) has also asserted that the increase in running behavior in the second alley of a double runway following non-reward in the first goal box, i.e., the FE, may actually be due to an addition of escape responses from the aversive non-reward situation. This analysis also fits the finding that amygdala lesions virtually eliminate the FE in a double runway (Henke & Maxwell, 1973). In summary, it is concluded that the affective responses following non-reward experiences, i.e., the aversive and energizing effects, are related to processes in the amygdala. It is suggested that the inhibitory influences from the septal area, through fibers of the stria terminalis, normally mediate the aversive effects of non-reward relative to current levels of reward expectations. A cognitive mechanism, normally used to learn to anticipate non-reward, is Limbic lesions and non-reward
associated with the hippocampus and fornix system. In other words, the hippocampus is assumed to be important when nonreward inhibits approach responses on which it was made contingent. In addition, the fornix also contains fibers which transmit information about reductions in incentive-motivation. This is suggested to occur following various manipulations to reduce reward availability, e.g., during extinction. RESUME
Des rats ayant subi des lesions bilaterales de l'amygdale, du septum, de l'hyppocampe, de la stria terminalis et du fornix sont soumis a un regime multiple de renforcement dans lequel la pression d'un levier, dans l'une des composantes, est associee a un renforcement (S+) a intervalles variables et la seconde composante associee a l'extinction (S—). Les reponses liees a un second levier eliminent le S— pendant des periodes de 5s au cours de l'extinction. Tous les groupes, exception faite des animaux ayant subi des lesions dans l'amygdale, manifestent un contraste comportemental. Les rats ayant subi des lesions dans l'hyppocampe ou dans le fornix montrent plus de resistance a l'extinction. Le taux des reponses liees au levier qui elimine le S— est plus eleve quand les lesions affectent le septum et la stria terminalis que quand elles affectent l'amygdale. La conclusion affirme que les lesions de l'amygdale attenuent les effets dynamiques et aversifs du non-renforcement, les lesions de l'aire septale et de la stria terminalis augmentent les effets aversifs, et les lesions de l'hyppocampe et du fornix interferent avec les effets inhibiteurs du non-renforcement. REFERENCES BOLLES, R.c. Reinforcement, expectancy, and learning. Psychol. Rev., 1972, 79, 394—409 DICKINSON, A. Septal damage and response output under frustrative non-reward. In R.A. BOAKES and M.S. HALLIDAY (Eds.), Inhibition and learning. London: Academic Press, 1972 DOUGLAS, R.J., PRIBRAM, K.H. Learning and limbic le-
sions. Neuropsycholog., 1966,4, 197-220 FRIED, P.A. The septum and behavior: A review. Psychol. Bull. 1972,78,292—310 GAFFAN, D. Inhibitory gradients and behavioral contrast in rats with lesions in the fornix. Physiol. Behav., 1973,ii,215—220 GRAY.J.A. Effects of septal driving of the hippocampal
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theta rhythm on resistance to extinction. Physiol. Behav., 1972, 8,481—490 GRAY.J.A., QUINTAO, L., & ARAUJO-SILVA, M.T. The partial reinforcement extinction effect in rats with medial septal lesions. Physiol. Behav., 1972, 8,491-496 HENKE, p.G. Effects of reinforcement omission on rats with lesions in the amygdala./, comp. physiol. Psychol., 1973. 8 4. 187-193 HENKE, p.G. Persistence of runway performance after septal lesions in rats.J. comp. physiol. Psychol., 1974, 86, 760-767 HENKE, p.G. Septal lesions and the extinction of incentive-motivation. Physiol. Behav., 1975, 15, 537-542 HENKE, p.G. Septal lesions and aversive nonreward. Physiol. Behav., 1976,17,483-488 HENKE, P.G. Dissociation of the frustration effect and the partial reinforcement extinction effect after limbic lesions in rats./, comp. physiol. Psychol., 1977, 91, 1032-1038 HENKE, P.G., ALLEN, J.D., & DAVISON, c. Effects of lesions
in the amygdala on behavioral contrast. Physiol. Behav., 1972,8, 173—176 HENKE, P.G., & BUNNELL, B.N. Reinforcement and ex-
tinction interactions after limbic lesions in rats. Communic. behav. Biol., Part A, 1971,6, 329—333 HENKE, P.G., & MAXWELL, D. Lesions in the amygdala
and the frustration effect. Physiol. Behav., 1973, 10, 647-650 JARRARD, L.E., ISAACSON, R.L., & WICKELGREN, W.O. Ef-
fects of hippocampal ablation and intertrial interval on running acquisition and extinction./, comp. physiol. Psychol., 1964, 57,442—444 KIMBLE, D.P. Hippocampus and internal inhibition. Psychol. Bull., 1968, 70, 285-295 KING, F.A., & MEYER, P.M. Effects of amygdaloid lesions
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upon septal hyperemotionality in the rat. Science, 1958, 1*8, 655-656 MABRY, P.D., & PEELER, D.F. Effect of septal lesions on response to frustrative non-reward. Physiol. Behav., 1972,8,909-913 RAISMAN, G. The connexions of the septum. Brain, 1966,89,317-348 REYNOLDS, G.s. Behavioral contrast./, exp. Anal. Behav., 1961,4,57-71 ROSS, J.F., GROSSMAN, L., 8c GROSSMAN, s.p. Some be-
havioral effects of transecting ventral or dorsal fiber connections of the septum in the rat./, comp. physiol. Psychol., 1975,89,5-19 SCHWARTZBAUM, j.s., & GAY, P.E. Interacting behavioral effects of septal and amygdaloid lesions in the rat./. comp. physiol. Psychol., 1966, 61, 59-65 SKULL, j . DA VIES, K., & AMSEL, A. Behavioral contrast and
frustration effect in multiple and mixed fixedinterval schedules in the rat./, comp. physiol. Psychol., 1970,71,478-483 SWANSON, A.M., & ISAACSON, R.L. Hippocampal lesions
and the frustration effect in rats./, comp. physiol. Psychol., 1969, 68, 562—567 TERRACE, H.S. Conditioned inhibition in successive discrimination learning. In R.A. BOAKES and M.S. HALLI-
DAY (Eds.), Inhibition and learning. London: Academic Press, 1972 WAGNER, A.R. Frustration and punishment. In R.N. HABER (Ed.), Current research in motivation. New York: Holt, Rinehait and Winston, 1966 WINER, B.J. Statistical principles in experimental design. New York: McGraw-Hill, 1962 (First received 10 November 1978) (Date accepted 8 March 1979)
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