Hippocampal control of behavioral arousal: Duration of lesion effects and possible interactions with recovery after frontal cortical damage

XPEKIIIESTAL

33, 1%170

NEUROLOGY

Hippocampal

(1971)

Control

Duration

of

Lesion

Interactions

School

of Psychlogy,

of Mrdicinc.

Behavioral

Effects

with

Frontal Dcbavtrrrent

of

and

Possible

Recovery

Cortical

BYROE Prim-ctou

Arousal: after

Damage

A. CAMPBELL Unitwsifjt,

Princeton.

PERCY BALLANTINE

11

Ulliwrsit~~

Ci?lcimati,

of Cincimati.

Mew

Ohio

Jersey

08540

4521

AND GARY LYNCH* Dcpavtmwt

of Psycl~oh’ology.

Unizwrsity Received

of California.

Irzlirze,

Cnlifomia

92664

June 12.1971

The effects of gross hippocampal aspiration were compared with those of frontal cortical lesions on wheel running and generalized activity of rats during control and arousal conditions. The rate of recovery of these functions was then measured in animals with hippocampal lesions or with combined hippocampal and frontal lesions. From these studies, it appears that the hippocampus shares arousal control functions with frontal cortex, that it is partially responsible for the recovery that has been reported to occur after frontal lesions, but does not itself recover after damage. introduction

A behavioral syndrome associated with damage to the frontal cortex of the rat was described in earlier papers (3, 12, 13), and the speed with which various aspects of this syndrome disappeared or “recovered” was established (13, 14). Several explanations for recovery have been advanced; one of the best supported is that reorganization takes place in brain structures functionally related to the damaged region (4). It has often been noted that the hippocampus is anatomically and functionally related to the frontal cortex (15). and experiments using various species have shown that damage to this structure produces behavioral deficits similar to those seen after frontal lesions (8, 18, 23). This raises the possibility that the hippocampus may be involved with the recovery seen after frontal lesions. 1 This

research

was

supported

by NSF

Grant 159

GB-16973

and NIH

Grant

MHOS501.

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CAMPBELL,

BALLANTINE,

AND

LYNCH

The present paper compares the syndrome of effects caused by hippocampal lesions with portions of that described earlier for frontal ablations, and then establishesthe rate of recovery for these hippocampal effects. In addition, possible interactions between frontal cortex and hippocampus during recovery were inferentially examined with the use of combined (i.e., frontal and hippocampal) lesions. Experiment

I: Methods

and

Results

Frontal ablation causes two very different changes in the spontaneous activity of rats : daily wheel running is chronically elevated (3) ; and the excitatory effects of starvation, the estrous cycle, and amphetamine on generalized activity (as measured by stabilimeter cages or observation techniques) are greatly potentiated (3). Wheel running and generalized activity have separable neural substrates, both with frontal components (12). The first experiment examined the effects of radical hippocampal lesions on these behaviors, with apparatus and testing procedures identical to those used in the frontal pole studies. Methods. Male rats of the Sprague-Dawley strain (Perfection Breeders) weighing 275-325 g at the beginning of the experiment were used. Surgery was performed under pentobarbital sodium anesthesia with atropine pretreatment. A midline scalp and muscle incision was performed and the appropriate skull area cleared. Aspiration was used for the hippocampal lesions which were directed at the anterior-dorsal hippocampus, including the hippocampal commissure, dorsal fornix, and fimbria. The lesions were made with small diameter pipets which kept cortical damage to a minimum. Sham operations consisted of removing a skull flap but did not damage the dura mater. The running wheels used were standard Wahmann activity cages, and the stabilimeter cages have been described earlier (3). They were wire mesh cages measuring 18 X 20 X 38 cm, mounted on a central axle with a microswitch located at one end, which opened or closed each time the animal moved from one side of the fulcrum to the other. Food and water were suspended in containers over the axle to avoid unbalancing the cage. Both types of cages were kept in temperature-controlled (20 C), soundproof rooms. Servicing of cages,feeding, and weighing of the rat were carried out during a 2-hr interval from 10-12 am during which activity was not recorded. The rooms were kept on a 12-hr day-night cycle (lights on at 6 am) for all experiments. In this situation, the animals were exposed only to sounds indigenous to the rooms (fans, other rats, etc) and were effectively isolated from the laboratory environment and recording equipment. For the stabilimeter studies, naive rats were divided into those

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with hippocampal lesions and those with sham operations (groups of seven and six) and housed for 1 week in individual colony cages. Weights were taken daily during this period and food pellets were scattered about the cage floor. Following this, the rats were transferred to the stabilimeter cages and given 3 days of food and water ad Iibitu~z followed by 4 days of food deprivation. Unlike stabilimeter activity, wheel running takes several days to reach a stable level and varies greatly among animals. Therefore, the rats were given 10-14 days of preoperative wheel running, and animals which ran more than 2500 or fewer than 100 revolutions per day were discarded. In this way a relatively homogeneous group of runners was obtained and it was no problem to match the experimental groups for activity. As with the stabilimeter experiments, the groups were also equated for preoperative body weight. After this adaptation period, the wheel-housed animals were divided into two groups: eight with hippocampal lesions and five with sham operations. Following surgery, the rats were returned immediately to the activity wheels, given 14 days of unlimited running with food and water always available, and then deprived of food for 2 days. Deprivation was limited to 2 days because of the higher mortality rate associated with longer periods of starvation in the wheel (3). At the end of the deprivation period, cardiac perfusions with saline solution followed by 10% formalin were performed on all subjects. The brains were removed and serial coronal slices were made at 50 p on a freezing microtome. Representative sections were then stained with cresyl violet and reconstructions made with a microprojector. Results. Reconstructions of representative brains are shown in Fig. 1. Destruction of the dorsal hippocampus was complete and bilateral in the transverse plane with the exception of a small portion of fimbria in the extreme lateral edge of the lateral ventricle. In most animals the lesions extended in the anterior to posterior dimension from the level of the hippocampal commissure in front of the hippocampal cell fields to a point immediately anterior to the habenula. Extra hippocampal damage was in most cases unilateral and restricted to dorsal thalamus (including stria medullaris) and occasionally the stria terminalis. In two rats the thalamic damage was judged to be excessive and their data were not included in the study. General recovery from the lesions was uneventful, and the rats quickly regained their preoperative weights. They demonstrated no obvious impairments in motor control or affective behavior and required no special treatments. As seen in the left panels of Fig. 2, the rats with hippocampal lesions

162

CAMPBELL,

BALLANTINE,

AND

LYNCH

FIG. 1. Succesive coronal section through a representative hippocampal hatched).

lesion (cross-

were considerably more active in the stabilimeter cages during ad libitum feeding periods than sham-operated animals (P
II: Methods

and

Results

The first experiment demonstrated that destruction of the dorsal hippocampus potentiates the arousal effects produced by food deprivation. The following experiment investigated the effects of similar lesions on the stim-

163

AROCSAL STABILIMETERS

‘WHEELS

PRE OP.

I2345

II 13 15 I7

12345678

DAYS

FIG. 2. The effects of hippocampal lesions (0) on median daily wheel and stabilimeter activity during ad /iOitzun and food-deprivation conditions. The double arrows (t) refer to the time at which food was removed from the cages; note also that the last 3 days of preoperative wheel activity are shown. Sham-operated animals, open circles. (0).

ulant properties

of amphetamine

and measured the duration of such effects. Earlier studies (13) have shown that the augmenting effects of frontal lesions on amphetamine hyperactivity gradually disappear over 40 days of postoperative recovery. Methods. Subjects and apparatus were described in Experiment I. ,Aspiration of the dorsal hippocampus was performed on three groups of rats which were then allowed to recover in individual colony cages for 7 (eight rats), 17 (seven rats), and 37 (eight rats) days before being tested. Sham-operated rats (three groups of nine each) were also tested after intervals. The rats were placed in the stabilimeter cages and given a saline solution injection 24 hr later (i.e., days 8, lS, and 35). On the fourth day of testing they received r-amphetamine sulfate (dosage 3.5 mg/kg, ip). Preoperative weights were selected so that weight at the time of injection would be the same in all groups. In this way the animals received approximately the same absolute quantity of amphetamine as well as equal injections (“g/kg). Following the injections, crossings were recorded every 30 min for 4 hr. These procedures, including drug doses, were identical to those followed in earlier studies which established recovery rates after frontal lesion ( 13 ) . Results. Histological analysis revealed that the lesions for these groups

164

CAMPBELL,

BALLANTINE,

AND

LYNCH

varied more than those used in Experiment I, and it was necessary to discard two animals from the ZO-day and three from the 40-day groups leaving five rats in each. It was noted that lesions in animals used for the longer recovery intervals tended to be larger than those in the lo-day group. However, the description and statistical significance of the results given below were essentially the same if the data from the rejected animals were included. Table 1 summarizes the daily activity of the different groups for the first and second days after being placed in the stabilimeter cages. It is clear that the increase in daily activity produced by the hippocampal lesions was permanent, since the rats given over 5 weeks of postoperative recovery were at least as active as those tested a week after surgery. The decline in activity from the first to second days reflects adaptation to the cage. None of the differences between hippocampal groups was statistically significant while all hippocampal versus sham comparisons were significant (P
1

DAILY ACTIVITY OF RATS WITH HIPPOCAMPAL LESIONS AND SHAM OPERATIONS TESTED AFTER VARIOUS RECOVERY PERIODS a Test

Operation

Recovery (days)

day

1

2

3

Hippocampal Sham

10 10

442 202

482 118

462 114

Hippocampal Sham

20 20

7.51 291

597 144

830 1.51

Hippocampal Sham

40 40

785 230

465 126

398 112

at each interval

and all animals

s Different mine injection

groups were on day 3.

tested

received

an ampheta

163

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Experiment

III: Methods

and

Results

On the basis of the previous experiment it appears that the hippocampus (like the frontal cortex) acts to inhibit the increases in spontaneous activity caused by starvation or amphetamine, a finding which strengthens the possibility that it might be involved in the recovery which takes place after frontal lesions. If this were so, removal of both frontal tortes and hippocampus would be expected to produce a larger effect than damage to either alone, and the interaction would be permanent since the structure normally involved in recovery from frontal lesions (i.e., hippocampus) would be destroyed. If, on the other hand, an estrahippocampal region were involved in recovery from frontal lesions, the rats with combined lesions would gradually return to the level of rats with hippocampai lesions only. Xrt1zods. Frontal poles and hippocampus were removed by aspiration in one operation, following the general procedures outlined in Experiment I. In aspirating the frontal poles, the intent was to remove all tissue rostra1 to anterior pole of the caudate nucleus with the exception of tissue on the floor of the cranial activity related to the olfactory tubercle and prepyriform cortex (3 ) . The postoperative procedures followed those described in Experiment I. Two recovery periods (20 and 40 days) were used. Groups of sham, combined frontal and hippocampal lesions, and hippocampal only lesions were tested at each period : at 20 days there were 11 rats with sham operations, eight with hippocampal, and 13 with combined lesions, while at 40 days the group consisted of six with sham, five with hippocampal. and six with 400

-

300

-

20

IO B w 8

200-

100 -

o-

h & &*a-** 0

2

4

k --***a* 6

8

0 TIME

2

SUCCESSIVE

4

6 30

8 MINUTE

FIG. 3. Effects of varying postoperative intervals on the median response of hippocampal (80) and sham-operated rats to a 3.5 mg/kg r-amphetamine injection. Saline solution: dotted lines. Activity at successive 30-min periods after the injection is shown and different groups were tested at each recovery interval.

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CAMPBELL,

BALLANTINE,

AND

LYNCH

combined lesions. Test procedures were identical in all cases. At the end of the appropriate recovery periods, the rats were placed for the first time in the stahilimeter cages and allowed 3 days to habituate to the cage. They were then given a randomized sequence of injections which included 1, 2, 3, and 4 ml d-amphetamine sulfate (prepared in a volume of 1 ml/kg of body wt) with at least 24 hr between injections. Activity was recorded in the same manner described in Experiment II. Results. Examples of the type of frontal lesions used in these studies have been published (3, 13). Briefly, the injury was confined to a region anterior to the corpus callosum, although the rostra1 tip of the caudate nucleus may have been included on occasion. Lesions were extremely consistent between animals and no relationship between morphological characteristics of the extirpation and behavioral changes was observed. Hippocampal lesions in both hippocampal and combined frontal plus hippocampal groups were accurately placed and little extrahippocampal damage was done. The dose-response curves for both 20- and 40-day recovery groups are shown in Fig. 4. The combined lesions greatly amplified the stimulant effects of amphetamine at all dosages tested : the effect was large and seen in all but two animals. Hippocampal lesion produced similar effects but these were of lesser magnitude at the lower three dosages. Although the total activity over 4 hr increased as a function of dosage in the 4O-day groups, the 4 mg/kg dose was in fact an overdose. This is indicated by the relative depression (insert, Fig. 4) produced by this dose in the 30-90-min postinjection period, the time of maximal effectiveness of amphetamine at lower doses intraperitoneally. Discussion

From these experiments, it appears that frontal cortical and hippocampal lesions in rats affect some behaviors similarly but there are a number of very prominent differences. The major similarity between the two types of lesion is a potentiation of conditions (amphetamine and food deprivation) which normally increase stabilimeter activity. One explanation for this is suggested by the electrophysiological literature. Both hippocampus and frontal cortex have been shown to have pronounced inhibitory effects on the brain stem reticular system (1, 6, 19). In addition, both are particularly responsive to reticular stimulation (7, 22), suggesting that each receives extensive innervation from the reticular formation. To the extent that the reticular formation is linked to behavioral arousal (as it is commonly held to be), it is not surprising that both hippocampal and frontal lesions increase the nonspecific effects of conditions which induce arousal. The present results also point up a number of discrepancies between the

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24002400

20 DAYS

FORTY

TWENTY

/

2000 1600-

OI 1.0

I 20

I 30

I 40

d-AMPHETAMINE

I IO

I 2.0

I 30

1 40

(mg/kg)

FIG. 4. Median response of rats with sham operations CO), (0 --0) or combined frontal cortical and hippocampal lesions ous doses of d-amphetamine sulfate, tested 20 or 40 days after hand panels summarize the response over time of the various with 4.0 mg/kg d-amphetamine.

hippocampal lesions (0-O) to varisurgery. The rightgroups when tested

“syndromes” associated with hippocampal and frontal damage. Hippocampal lesions increase stabilimetric activity during ad lib&m feeding conditions while the frontal ablation does not. On the other hand, frontal lesions increase daily wheel activity and the hippocampus does not appear to participate in this behavior. Taking this latter result first, in an earlier study it was found that only the ventral portion of the frontal pole was related to wheel activity (12). The present data suggest that the discrete ventral frontal-hypothalamic system delineated in that research is regulating behaviors with which the hippocampus is not involved. Anatomical studies in cats and primates link the dorsolateral aspects of the frontal lobes with the hippocampus and the inferior or orbital frontal surfaces to amygdala, magnocellular dorsomedial thalamic nucleus, and hypothalamus-the “ATO” system of Nauta (15). This is not to suggest that these relationships are totally formed in the relatively primitive rat brain, but these data do indicate that some of the circuitry of higher forms may be operative in rats. The hippocampus, in contrast to frontal cortex, inhibits generalized activity in addition to reactively suppressing increases in arousal level. There are little data to suggest that the same neuronal system was involved in both effects. In view of the recent neuroanatomical findings dissociating the rat hippocampus into medial and lateral divisions, according to efferent relationships (17), it is conceivable that different subdivisions are respon-

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CAMPBELL,

BALLANTINE,

AND

LYNCH

sibIe for the chronic and reactive aspects of hippocampal inhibition of activity. Preliminary behavioral data (Lynch and Campbell, unpublished data) suggest that this may be the case. In addition, the combined lesions revealed a number of possible behavioral interactions between frontal cortex and and hippocampus. These lesions made the animals more active during ad libituvn feeding conditions than hippocampal lesions alone, occasionally vicious, and moderately aphagic. Neither of the last two effects was seen with frontal or hippocampal lesions alone. Using response to amphetamine as a test, hippocampally lesioned rats show no evidence of recovery after postoperative periods that are adequate for the complete disappearance of frontal lesion effects. There are a number of neurological theories to explain recovery or failure of recovery; e.g. axonal growth (16) and supersensitivity (21), but more global theories usually incorporate the idea of reorganization in the brain structures functionally related to the damaged area (4). There are numerous experimental demonstrations that something of this sort occurs; for example, after damage to primary motor (9) or affective/motivational (5, 24) centers, various cortical regions appear to gain a degree of regulation in these functions that apparently is not present in the normal animal. Since the present data and earlier work indicate that frontal cortex and hippocampus “share” arousal control functions, it seems plausible that changes in hippocampal activity could account for the recovery that takes place after frontal lesions. The results from the present experiments provide at least indirect support for this idea. If an extrahippocampal structure is responsible for the disappearance of the potentiation of arousal caused by frontal lesions, some recovery should have occurred in the rats with combined f rontal-hippocampal lesions. Since the dose-response curves for the animals with combined lesions at 40 days demonstrate that the effect of the frontal lesions is still very much present at that time, it follows that the hippocampus is at least in part responsible for the complete recovery reported for rats with frontal lesions alone. Further research will be required before these conclusions can be accepted, (particularly with 40day intervals separating frontal and hippocampal surgery), but these results do provide an explanation for the fact that the arousal potentiation caused by frontal lesions shows recovery while the wheel activity increases appear to be permanent ( 14). In light of these findings, it is puzzling that there was no evidence of recovery after hippocampal lesions. In its simplest terms, this implies that hippocampus (and probably other structures) can offset the changes caused by frontal lesions but cortex is unable to reciprocally adjust for the arousal imbalances produced by hippocampal damage. This is particularly

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surprising since there are numerous examples of the involvement of cortical areas in the recovery which occurs after damage to various &cortical and brain-stem regions (24). This emphasizes the necessity of eatablishing the parameters of the recovery process for various neurological systems and the behaviors associated with them. It seems not unlikely that a number of mechanisms are involved with the recovery process and generalized hypotheses deduced from a limited number of cases will prove to be inadequate. Finally, while the electrophysiological and behavioral data indicate that cortex and hippocampus share inhibitory functions over the reticular formation and behavioral arousal, neuroanatomical research shows that they are accomplishing this by very different mechanisms. The frontal cortex’is linked with the thalamic reticular system via the inferior thalamic peduncle (12,2(J) and appears to suppress the facilitory pontine reticular formation via a relay in the bulbar nucleus reticularis gigantocellularis (19). Hippocampal innervation from the reticular formation appears to travel chiefly in the medial forebrain bundle (7). Therefore it is possible that the hippocampus and frontal poles are inhibiting different aspects of the reticular formation and arousal response: greater use of observational techniques should be of great help in detailing the nature of the inhibitory deficits caused by these lesions. References 1. ADEY, W. R., J. P. SEGUNDO, and R. B. LIVIWXTOX. 1957. Corticofugal influences on intrinsic brain-stem conduction in cat and monkey. J. Ne~~rophysiol. 20 : 1-16. 2. BATINI, C., and 0. POMPEIANO. 1957. Chronic fastigial lesions and their compensation in cat. .4rch. Ital. Biol. 95 : 147-165. 3. CAMPBELL, B. -4., and G. LYNCH. 1%9. Cortical modulation of spontaneous activity during hunger and thirst. J. Camp. Pltysio2. Psychol. 67: 115-122. 4. CHOW, K. 1967. Effects of ablations, pp. 705-715. In “The Neurosciences.” G. Quarton, T. Melnechuk, and F. Schmitt [eds.]. Rockefeller Univ. Press, New York. 5. CYTAIVA, J.. and P. TEITELBAUM. 196’7. Spreading depression and recovery of subcortical functions. Acta Biol. Exp. Warsaw 27: 345-353. 6. DELL, P. 1963. Reticular homeostasis and cortical reactivity, pp. 82-103. 11~ “Brain Mechanisms.” G. Morizzi, A, Fessard, and H. Jasper [eds.]. Elsevier, New York. 7. GREEN, J., and A. ARDUIXI. 1954. Hippocampal electrical activity in arousal. J. Nmrophysiol. 17 : 533-538. 8. GROSS, C. G., S. L. CHOROVER, and S. M. COHEN. 1965. Caudate, cortical, hippocampal and dorsal thalamic lesions in rats: alternation and Hebb-Williams maze performance. NezrroQsychologia 3 : 53&6X 9. KENNAKD, M. 1942. Cortical reorganization of motor function: Studies on series on monkeys of various ages from infancy to maturity. .4rrh. Nettrol. Psyckiat. 48 : 227-241.

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10. KIMBLE, D. 1963. The effects of bilateral hippocampal lesions in rats. J. Co)rtp. Physiol. Psychol. 56 : 273-283. 11. LIU, C., and W. CHAMBERS. 1958. Intraspinal sprouting of dorsal root axons. Arch. Neural. Psychiat. 79 : 4661. 12. LYNCH, G. S. 1970. Separable forebrain mechanisms controlling different manifestations of spontaneous activity. J. Camp. Physiol. Psychol. 70,: 48-59. 13. LYNCH, G. S., P. BALLANTINE, and B. CAMPBELL. 1969. Potentiation of behavioral arousal following cortical damage and subsequent recovery. Exp. Neurol. 23: 195-206. 14. LYNCH, G. S., P. BALLANTINE, and B. CAMPBELL. 1971. Differential rates of recovery following frontal cortical lesions in rats. Physiology Behazfior. in press. 15. NAUTA, W. J. H. 1964. Some efferent connections of the prefrontal cortex in the monkey, pp. 3-31. Ztz “Frontal Granular Cortex and Behavior.” J. Warren and K. Akert reds.]. McGraw-Hill, New York. 16. RAISMAN, G. 1969. Neuronal plasticity in the septum. Brabt Res. 14: 25-48. 17. RAISMAN, G., W. COWAN, and T. POWELL. 1965. An experimental analysis of the efferent projection of the hippocampus. Brain 99: 82-108. 18. ROSVOLD, H. E., and M. K. SZWARCBART. 1964. Neural structures involved in delayed response performance, pp. 1-16. 11% “The Frontal Granular Cortex and Behavior.” J. M. Warren and K. Akert [eds.]. McGraw-Hill, New York. 19. SAUERLAND, E. K., Y. NAKAMURA, and C. D. CLEMEXTE. 1967. The role of the lower brain stem in cortically induced inhibition of somatic reflex in the cat. Brain Res. 6 : 164-179. 20. SCHEIBEL, M. E., and A. G. SCHEIBEL. 1967. Anatomical basis of attention mechanisms in vertebrate brains, pp. 577-602. In “The Neurosciences.” G. C. Quarton, T. Melnechuck, and F. 0. Schmitt [eds.]. Rockefeller Univ. Press, New York. 21. SHARPLESS, S. K. 1964. Reorganization of function in the nervous system-use and disuse. Ann. Rev. Physiol. 26 : 357-388. 22. STARZL, T., C. TAYLOR, and H. MAGOUN. 1951. Ascending conduction in the reticular activating system with special reference to the diencephalon. J. Ncurophysiol. 14 : 461. 23. TEITELBAUM, H. 1964. A comparison of orbitofrontal and hippocampal lesions upon discrimination learning and reversal in the cat. Exp. Neurol. 9: 452-462. 24. TEITELBAUM, P., and J. CYTAWA. 1965. Spreading depression and recovery from lateral hypothalamic damage. Science 147 : 61-63.