changes in 24-hour fluctuations of feeding behavior during hypothalamic hyperphagia in rats

Physiology & Behavior, Vol. 21, pp. 615--622. Pergamon Press and Brain Research Publ., 1978. Printed in the U.S.A.

Changes in 24-Hour Fluctuations of Feeding Behavior During Hypothalamic

Hyperphagia in Rats

W. J. R I E T V E L D , F. TEN HOOR, M. KOOIJ A N D W. F L O R Y

Department of Physiology, Department of Physiological Psychology, University of Leiden Leiden, The Netherlands

(Received 6 October 1977)

RIETVELD, W. J.', F. TEN HOOR, M. KOOIJ AND W. FLORY. Changes in 24-hourfluctuations of feeding behavior during hypothalamic hyperphagia in rats. PHYSIOL. BEHAV. 21(4) 615--622, 1978.--Lesions were electrolytically placed in the ventromedial hypothalamic area in rats. Bilateral lesions appear to be more effective in producing hyperphagia than unilateral lesions. Changes in the circadian pattern during the hyperphagic phase are mainly brought about by a variable degree of increase in eating during the light periods. The fact whether or not a rhythm is present is determined by the localization of the lesion. Symmetrically placed lesions result in loss of rhythmicity in most cases but this does not correlate with the degree of hyperphagia. A possible role of connections between the suprachiasmatic nucleus and the ventromedial hypothalamic area is suggested.

Circadian rhythms Feeding behavior Hypothalamus Hyperphagia

IT has been known for many years that injury or destruction of the hypothalamus will lead to obesity [5, 8, 10, 22], which is the result of an increased food intake (hyperphagia) and is maximal after destruction of the ventromedial region [5,14]. The increased food intake takes place during the so- called dynamic phase of hyperphagia and leads to a rapid increase in body weight. In course of time a static phase is reached in which the hyperphagia disappears and body weight stabilizes at a new level.

To what extent the food pattern during and after hyperphagia is changed, is rather complicated. Brooks and cowor- kers [7] describe how during the dynamic phase the daylight activity level in rats becomes equal to or even slightly higher than the night time level. In the static phase the pattern tends to revert to normal [6,7]. Balagura and Devenport [2] describe the eating pattern of hyperphagic rats in a more de- tailed way. In contrast to the findings of Brooks et al. they found that the circadian pattern in hypothalamic lesioned animals of both sexes shows an increase in feeding during light and dark time as compared to normals. Eating was similar in both sexes during the light period but greater for females at night.

Kakolewski, Deaux, Christensen and Case [15] report that rats with bilateral ventromedial hypothalamic lesions display a permanent loss of the circadian rhythm in food intake not only during the dynamic phase but also during the static phase. They tentatively put forward the question whether this disappearance is caused by substitution of a random eating pattern or by that of a free running oscillatory pattern.

More recently differences in circadian feeding patterns during hyperphagia have been described by Ahlskog, Ran- dall and Hoebel [1]. Following a suggestion by Gold [11] that lesions which produce hypothalamic hyperphagia are not restricted to the ventromedial hypothalamic nucleus, they induced hyperphagia by destruction of the ventral noradrenergic bundle (VNAB) using intramesencephalic injections of 6-hydroxydopamine. Their results suggest that the class- ical ventromedial hypothalamic (VMH) hyperphagia and the VNAB-hyperphagia are two different syndromes. The VMH-lesioned animals show an increased food intake during the day as well as during the night. On the other hand the VNAB-lesioned rats display an increase in food intake at night but a normal level at day. The literature on unilateral VMH lesions and hyperphagia is scarce [20] and no mention is made of whether a circadian pattern is present or not.

Although the data on changes in the 24 hr eating pattern of rats after lesions in the neighbourhood of the ventromedial hypothalamic area thus are rather diverse, they clearly indicate that in this way the 24 hr eating pattern can be desyn- chronized or destroyed. Experiments by Richter [29] already showed that blinding or various types of endocrinological interference are not able to abolish the 24 hr rhythm in eating and locomotor activity; lesions of the hypothalamic area are the only ones which result in complete disappearance of circadian fluctuations. This suggests that a center or network is situated in the hypothalamus, by which the 24 hr eating rhythm is initiated or coordinated.

The confusing results on changes in eating pattern after ventromedial hypothalamic lesions may partly be explained

Copyright © 1978 Brain Research Publications Inc.I0031-9384/78/100615-08502.00/0

616 RIETVELD ET AL.

by differences in experimental circumstances or in localization of the lesions. The present experiments were designed to investigate changes in eating activity of rats after uni- or bilateral destruction of the ventromedial hypothalamus and to compare these changes with morphological data. These studies led to a hypothesis on the origin of circadian eating activity which fits in well with recent f'mdings [26,31] which suggest a role of the suprachiasmatic nucleus in the origin of other 24 hr rhythms.

METHOD

Animals

Female Wistar rats (n=37, initial body weight about 250 g) were individually housed in perspex cages in a light and temperature (23°C) controlled room with an average relative air humidity of 60%. Lights were on from 0100-1300 hr and off from 1300-0100 hr. The rats had free access to food and water. Food consisted of pellets (Muracon; mean size 3.5 x 5.0 mm; mean weight 0.125 g) contained in boxes which were constructed as described below. Tap water could be obtained ad lib by licking from bottles hanging in the cage. During light periods the rats were inspected regularly and any food spillage was thus rapidly detected. By weighing the food boxes and correcting for spillage total food intake of each rat was determined weekly. Body weight of the rats was determined twice weekly (Mettler balance, type K5). In addition food and water intake were measured concurrently once a week for 24 hr to detect deviations from the normal pattern in 15 VMH lesioned hyperphagic animals.

Surgery

After anaesthesia (pentobarbitone sodium 60 mg/kg IP or hypnorm ® 1 ml/kg IM) a stainless steel electrode (di- ameter 0.4 mm; bare tip 0.5 mm) was introduced uni- or bilaterally into the brain of the animal with the aid of a home-made stereotaxic apparatus. The coordinates used were 0.0; 0.6; -9 .0 mm according to the atlas of Cushman and Pellegrino. A reference electrode was introduced IM or into the rectum. Lesions were made by applying a dc current of 1.2 mA for 20 sec, the hypothalamic electrode being the cathode. After the lesion had been made and the animal had recovered from the operation, it was returned to its cage and fasted for 24 hr.

Apparatus

A survey of the experimental set-up is schematically shown in Fig. 1. Each cage contained a pellet box constructed in such a way that for each approach to the food a lightweight perspex flap had to be pushed away by the animal. A similar device has been described recently by Bostwick and Porter [4]. The movement of the flap resulted in the activa- tion of an approach indicator (Pepped and Fuchs), the output of which was fed into a home-made Activity Monitor Control Unit (AMC Unit). Using this apparatus eating activity data from 20 animals and a number of items concerning general information (e.g., No. of animal, light or dark period) can be stored simultaneously during a 0.5 hr period, totalled for this period and then recorded on paper tape (Tally 420). The data were analyzed using an IBM 1800 or a PDP 11/45 computer.

In a separate experiment with 9 normal rats it was shown that using this set-up there is for each rat a good correlation between the number of food approaches recorded and food

a n i m a l h o u s e 1 - 2 0

[+nv,+nm°o / Oa:Ou,:O+ j

FIG. I. Schematic survey of the experimental set-up used to record eating activity in rats. Each of 20 animal houses contains a food box with an approach detector which is activated when the animal pushes away a lightweight perspex flap to reach the food. The output of the detector is fed into an Activity Monitor Control Unit (AMC unit) together with data on No. of cage and light or dark period. In the AMC unit these data are stored and totalled during a period which can be adjusted by means of a clock (in the present experiment a 0.5 hr period was chosen). The output of the AMC unit is recorded on paper tape and then analysed using an IBM 1800 or PDP

11/45 computer.

intake measured during the 12 hr light as well as the 12 hr dark periods. The correlation coefficients found range from 0.90 to 0.99 with a mean value of 0.94 (SD 0.04). Taking a mean pellet weight of 0.125 g and assuming an average consumption of 1 pellet per approach, the relative food intake in one rat can roughly be estimated from the number of food approaches recorded.

Data Analysis

After the data had been put on disk (DEC RP04), they were for each animal divided into the pre- and postoperative period and a plot of activity against time was made contain- ing also information about the light/dark regime (Fig. 2). This plot was inspected and according to the findings data reduc- tion performed resulting in 6 histograms per animal, 3 representing the eating activity during the preoperative phase and 3 representing the eating activity during the postoperative period (Fig. 3). As the postoperative period was usually rather long, it was divided into parts of 22 days from which the histograms were made. These show the total number of half-hour intervals in which the rat displayed each level of eating activity as quantified by the number of times food was taken. With the aid of these histograms differences between the eating activity before and that after the operation could be observed, and an indication obtained of differences between day and night activity for each animal. To test the presence of a difference in eating activity between day and night the eating activities in the light periods and those in the dark periods were totalled for the 22 day periods. In this way 22 matched pairs of light and dark eating activity were obtained and a Wilcoxon test for matched pairs was carried out [30] at a significance level ofp <0.01 in all cases. Additionally histograms of the average activity during the light period as well as during the dark period were made for the same 22 day periods.

Histology

At the end of the experiment the animals were killed and

CIRCADIAN RHYTHM A N D VMH LESIONS 617

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eating rhythm.

the brains successively flushed with a 0.9% NaCI and a 10% Formalin solution. The brains were then removed and stored in a 10% Formalin solution for 24 hr. From the hypothalamic part of the brain transverse 8 /xm sections were then made, mounted on microscope slides, dried at 37°C for 24 hr and stained according to K10ver-Barrtra. Sections were studied and classified with respect to s ize and symmetry of the lesions. To detect whether VMH lesions had any influence on the oestrus cycle , vaginal smears were made on 5 consecutive days in 5 control and 15 VMH lesioned hyperphagic rats. The smears were stained according to Papanicolaou and the phase of the cycle determined.

Procedure

At the start of the experiment the rats were placed in a cage and allowed to get accustomed to the environment and the special food box for a period of about 2 weeks. During this period eating activity was monitored and animals han- dled as described. The rats were then lesioned, put back into the cages, fasted for 24 hr and monitored for periods ranging from 15 to 35 weeks , the length of this period depending on the time it took to reach the static phase of hyperphagia.

P R E O P E R A T W E P E R I O D

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FIG. 3. Frequency diagrams of half-hourly eating activity before and after lesioning the ventromedial hypothalamus. In each histogram the first column represents the number of half hour periods (totalled for a period of 25 days) in which the rat takes food 1--4 times; the second column represents the number of half hour periods (totalled for a period of 25 days) in which the rat takes food 5-8 times and so on. The left panel shows histograms of a rat with an increased total eating activity after operation. The histograms from the dark and light periods show that there is a 24 hr eating rhythm in the preoperative as well as in the postoperative period. In the right panel the histograms show an increased total eating activity and a disap-

pearance of the 24 hr eating rhythm after operation.

RESULTS

A survey of the results of the experiments is given in Table 1. Out of 24 animals with bilateral VMH lesions 20 became hyperphagic, the criterion for this being that body weight gain during the first 4 postoperative weeks was > 10% of the body weight at operation. This criterion is based on growth curves (3-30 weeks) obtained in 25 normal animals of the same strain as used in the experiments described. In the female animals the mean rate of growth once they had reached a body weight of 250 g was on the average 4% in 4 weeks. In the bilaterally VMH lesioned animals body weight gain ranged from 48-639 g and was on the average 48.4% of body weight at operation. The dynamic phase of hyperphagia lasted between 2 and 30 weeks with a mean value of 10.0 weeks. As it is difficult to give exact criteria for the moment at which the dynamic phase comes to an end, the time span of the dynamic phase was estimated from growth curves which were obtained for each animal; Fig. 4 shows two ex- tremes.

A 24 hr eating rhythm was present in all rats before the

618 R I E T V E L D ET AL

T A B L E 1

SURVEY OF EXPERIMENTAL DATA

Criterion Bilateral lesions Unilateral lesion

No. of animal operated 24 13 No. of animals with hyperphagia* 20 7 Mean maximal body weight gain of 48.4 ± 22.2 15.6 _+ 3.4 hyperphagic animals (% of initial weight ± SD)

Mean duration of dynamic phase 10.0 ± 6.6 n.d. (weeks ± SD)

No. of hyperphagic animals in which a 24 hr eating rhythm was - present 8 7 - absent 5 - -

- variably present 6 - - - not determinable 1 - -

*An animal was considered to be hyperphagic when body weight gain during the first 4 postoperative weeks was > 10% of the body weight at operation (initial weight).

n.d. not determined

1000

900

800

700

600

500

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300~

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5 10 15 20 25 30 35

FIG. 4. Two examples of the course of bodyweight (g) of rats made hyperphagic by bilateral electrolytic lesions of the ventromedial

hypothalamus.

' ' ' - - ' ' ~ o d a ~ per,o~ [ . food ,ntake{ . . . . l T (grams) .l~gnt pet',or

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FIG. 5. Persistence of 24 hr eating rhythm in a rat after bilateral electrolytic lesions in the ventromedial hypothalamus. Food intake was estimated assuming a pellet weight of 0.125 g and an average

consumption of one pellet per approach.

V M H lesions were made. In 8 of the bilaterally lesioned animals this rhythm remained unchanged during the dynamic as well as the static phase of hyperphagia (Fig. 5). In 6 animals a 24 hr rhythm was present during part of the dynamic and/or the static phase and in 1 animal it was impossible to determine whether or not a rhythm was present. In 5 animals the rhythm comple te ly disappeared immedia te ly after the lesions had been made and this remained so during the whole exper imental period (Fig. 6). The observat ion that a 24 hr eating rhythm remains present during hyperphagia does not exclude " a t t e n u a t i o n " of rhythmicity. To give an impression o f the variations observed be tween comple te loss of rhy thm and " n o r m a l " rhythmici ty, Fig. 7 shows typical examples of day and night eating act ivi ty of V M H lesioned and control rats.

Out of the 4 rats which did not become hyperphagic after bilateral V M H lesions, 2 appeared to have a unilateral lesion, in 1 animal the lesions had been placed too far caudally and in 1

CIRCADIAN RHYTHM AND VMH LESIONS 619

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Food consumption was estimated as described in Fig. 5.

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FIG. 7. Typical examples of types of day (open bars) and night (diagonal hatching) eating activity, expressed as a percentage of total eating activity, of VMH lesioned and control rats. The bars indicate the highest, lowest and mean value of the mean eating activity observed during at least 7 consecutive 22 day periods. © rhythm present; • rhythm absent; * rhythm variably present. To test whether a rhythm was present or absent, a Wilcoxon test for matched pairs was carried out (see text). Rats No. 2-9, 14--3, 5-9, 7-6, 1-9 and 8-7 were bilaterally lesioned, Rat No. 3-9 was unilat-

erally lesioned and Rat No. 4-8 was a control animal.

animal the lesion was supposed to have been on the fight side, but appeared to be placed somewhere on the left.

Out of 13 animals in which unilateral VMH lesions were made, 7 became hyperphagic. In all unilaterally lesioned hyperphagic animals a 24 hr eating rhythm remained present in the postoperative period.

FIG. 8. Average day and night (cross hatched) eating activity (%) of all hyperphagic animals. I. Before operation. II. Immediately after bilateral VMH lesions were made. III. Half-way through the postoperative observation period. IV. At the end of the observation period. The eating activity during the dark period before the operation was put at 100%. Bar indicates SEM. In all postoperative periods there is a relative increase of eating activity during the light

periods.

Figure 8 shows the combined histograms of the average activity during the light and the dark periods measured over all hyperphagic animals. Measurements were taken preoperatively (I), immediately postoperatively (II) and half way through the postoperative period (III) as well as at the end of the observation period (IV). Increase of activity is most marked during the light periods.

When studying the histological preparations, it was observed that in the posterior-anterior direction all lesions lay in between +2.0 and-2 .0 mm with respect to the bregma and that all included the VMH area. Some lesions were situated somewhat more anteriorly, others somewhat more posteriorly. All but 4 lesions were located ventral as well as caudal to the paraventricular nucleus (PVN).

However, there was no correlation between a more anterior location of the lesion and increase in bodyweight or loss of rhythmicity. The rats with more posteriorly situated lesions did not show a higher food intake or more loss of rhythmicity as compared to the whole group. As to the left- fight variations, none of the lesions reached the lateral hypothalamic areas. The ventricle was often dilated most probably due to tissue retraction. Studying size and locus of the lesions, it was observed that a number of these showed a symmetrical aspect, while a number of other lesions were clearly asymmetrically placed with respect to the median line. Fig. 9 gives typical examples of these two types of lesions. To identify the degree of symmetry or asymmetry of the lesions, histological preparations of all lesions were classified as asymmetric (score 0) or symmetric (score 1) by 10 independent observers. The mean score for each indi- vidual animal was correlated with the presence or absence of a 24 hr eating rhythm during the postoperative period divided into parts of 22 days.

620 R I E T V E L D ET AL

FIG. 9. Typical examples of the types of lesions observed. Top: lesion which is symmetrically placed with respect to the median line. Level according to Cushman and Pellegrino + 0.0. Bottom: example of asymmetrical lesion. Level 0.0.

CIRCADIAN RHYTHM AND VMH LESIONS 621

The latter was expressed as a percentage, 100% indicating absence and 0% presence of rhythm during all parts of the whole postoperative period. For all data (bi- and unilaterally lesioned hyperphagic animals) a correlation coefficient of .70 (p<0.01) was found; with data obtained from bilaterally lesioned hyperphagic animals only, the correlation coefficient was .54 (p<0.01); Pearson's rank correlation test).

To investigate whether loss of 24 hr rhythm after the operation was related to the degree of hyperphagia, the per- centual increase in weight was correlated with the percentage of rhythm as indicated above (Pearson's rank correlation test). The correlation coefficient was 0.23 (not signifi- cant).

In 3 out of 15 hyperphagic animals in which water intake was measured for24 hr per week, the water intake was much higher than might be expected to be the result of their increased food intake. However, no relationship was observed between this increased water intake and the degree of hyperphagia or the presence or absence of rhythmicity. Six of 15 hyperphagic animals in which vaginal smears were studied showed a completely abnormal oestrus cycle. Again no correlation was found between this abnormality and the degree of hyperphagia or the presence or absence of eating rhythmicity.

DISCUSSION

In contrast with the method described by Kissileff [16] and with a number of other operant methods [3, 19, 33], our set-up used to record the rat's eating activity does not meas- ure meal size and pattern, nor does it give precise information on the amount of food consumed. However, as it was the intention to study the influence of VMH lesions on 24 hr eating activity and to limit intervention with the normal laboratory rat eating situation to a minimum, we developed the described simple, but reliable procedure, which in addition offers the possibility to roughly estimate the animals' food intake.

The results obtained with respect to hyperphagia and loss of rhythmicity compare fairly well with those described by a number of other investigators [2, 5, 7, 14, 15] and can be summarized as follows.

(1) Bilateral, electrolytically made, lesions of the ventromedial hypothalamic area in rats often lead to hyperphagia (20 out of 24 operated animals).

(2) Unilateral VMH lesions can lead to hyperphagia; however, the probability of a positive result is much lower than in bilateral lesions (7 out of 13 operated animals).

(3) After bilateral VMH lesions resulting in hyperphagia, a 24 hr eating rhythm can be present or absent during the whole or part of the observation period (up to 30 weeks).

(4) After unilateral VMH lesions resulting in hyperphagia the normal 24 hr eating rhythm always remains present.

The fact that lesions of the ventromedial hypothalamic area often result in hyperphagia is generally accepted. Ob- servations on changes in 24 hr eating activity after VMH lesions are not unanimous. Although it cannot be excluded that the rhythmicity observed in the present experiment was a passive response to the LD cycle, the changes of the 24 hr rhythm in eating activity after VMH lesions have a number of more probable explanations. Several investigators [9, 19, 28] indicate that especially more posteriorly situated VMH lesions which cause serious neuroendocrine disturbances, produced attenuated or abolished feeding rhythms. We ob-

served changes in drinking behavior or oestrus cycle in only few VMH lesioned animals and could not find any correlation with presence or degree of hyperphagia or rhythmicity. The number of observations being small, it seems not allowed to draw def'mite conclusions. A second explanation may be that the rhythm change is caused by lesions of the ventromedial hypothalamic nucleus itself, assuming that the cell group which determines food intake also governs the 24 hr rhythm. One would then expect a good correlation between the degree of hyperphagia and the change in rhythmicity, but this was not found in our results. Moreover, Gold et al [12], using parasagittal knife cuts to induce hyperphagia, came to the conclusion that hypothalamic obesity cannot be attributed to disrupted eating rhythms, and that satiety and eating activity may have separate though adja- cent substrates, the paraventricular nucleus (PVN) playing an important role in the satiety neurocircuitry [13].

This leads to the next possibility that a separate neuronal substrate which lies in the direct vicinity of the VMH and which generates and synchronizes the 24 hr eating activity, might be damaged to a lesser or greater extent when lesioning the VMH. Circadian fluctuations in neuronal activity of the VMH and LH areas have been reported by Koizumi and Nishino [17]. However, they conclude from earlier findings [25,26] that these fluctuations originate in the suprachiasmatic nucleus and not in more caudal cells of the hypothalamus. There are no reports about circadian fluctuation in neural activity of the PVN. Stephan and Zucker [31] report that in rats bilateral electrolytical lesions of the suprachiasmatic nucleus result in elimination of the circadian rhythm in eating and locomotor activity. Close inspection of our histological data showed that the suprachiasmatic nucleus was intact in all preparations.

A fourth possibility is that connections are severed between a more distantly located rhythm generating and syn- chronizing centre and the VMH when inducing hyperphagia with the electrolytical technique; this rhythm centre could be situated in nuclei from which activity reaches the VMH via the ventral noradrenergic bundle (VNAB).

If lesions of this bundle were the underlying cause of the rhythm changes observed after electrolytical VMH lesions, one might expect that these changes are similar to those found after destruction of the VNAB, where there is an increase in food intake at night, but a normal level at day [1]. When screening our data this pattern was not found (Fig. 8), indicating that lesions of the VNAB are not involved.

Using autoradiographic methods Swanson and Cowan [32] describe nervous connections between the suprachiasmatic nucleus and the periventricular area, reaching as far as the caudal ventromedial hypothalamic area. Especially in the area between the ventromedial hypothalamus and the ar- cuate nucleus, and in the ventral part of the VMH, they find a large number of labeled fibres. Millhouse [21 ] reports that a large number of VMH cells have ventrally running dendrites which make contact with fibres from the suprachiasmatic nucleus. Recording antidromic action potentials in the suprachiasmatic nucleus after electrical stimulation of the VMH, Kreisel [18] established the existence of suprachiasmatic efferents to the VMH electrophysiologically. Ret- rochiasmatic surgical knife cuts eliminate corticosterone rhythms [23] as well as pineal N-acetyl transferase rhythm [24]. On the other hand elimination of circadian rhythms in drinking activity requires a more extensive isolation of the suprachiasmatic nucleus, indicating the presence of dorso- lateral efferents as well [27].

622 R I E T V E L D E T A L .

When studying our morphological preparat ions, we observed that especial ly in the bilateral lesions which were judged to be symmetr ica l with respect to the median line, the per iventr icular area as well as the area lying ventral ly to the V M H was bilaterally damaged. On the contrary, in the lesions qualified as asymmetr ica l a part of the ventrolateral area is usually intact; in unilateral lesions this is a lways the case.

These data, the high correlat ion which exists be tween symmetry of a lesion and loss of 24 hr rhythmici ty of eating act ivi ty ( r=.70; p<0 .01) , and the fact that the suprachias- mat ic nucleus is involved in the control o f a number of o ther circadian rhythms [31], suggest that the loss of circadian rhythm in eating act ivi ty after electrolyt ical ly made V M H lesions depends on damage of connect ions be tween the suprachiasmatic nucleus and the vent romedia l hypothalamic

area. The degree of damage could then be responsible for the observed variability in loss of rhythm.

Whether terminals of the dorsal per iventr icular ly running efferents are involved or a more dorso-lateral , yet un- specified, system remains to be investigated.

In order to test this hypothesis further it will be necessary to study the effect o f lesioning the suprachiasmatic nucleus on 24 hr eating activity. These invest igat ions are now being under taken in our laboratory.

ACKNOWLEDGEMENTS

We thank Miss H. M. A. G. van der Loo, H. Duindam and L. Leeflang for handling the animals and making the lesions, J. L. den Hoed for making the histological preparations, Mrs. H. M. M. Willemse-van der Geest for general assistance and Miss M. de Rooy for typing the manuscript.

REFERENCES

1. Ahlskog, J. E., P. K. Randall and B. G. Hoebel. Hypothalamic hyperphagia: Dissociation from hyperphagia following destruction of noradrenergic neurons. Science 190: 399-401, 1975.

2. Balagura, S. and L. D. Devenport. Feeding patterns of normal and ventromedial hypothalamic lesioned male and female rats. J. comp. physiol. Psychol. 71: 357-364, 1970.

3. Becker, E. E. and H. R. Kissileff. Inhibitory controls of feeding by the ventromedial hypothalamus. Am. J. Physiol. 226: 383- 396, 1974.

4. Bostwick, A. D. and J. J. Porter. An efficient, inexpensive food hopper for monitoring feeding habits of rats. Behav. Res. Meth. lnstrum. 9: 471--472, 1977.

5. Brobeck, J. R., J. Tepperman and C. N. H. Long. Experimental hypothalamic hyperphagia in the albino rat. Yale J. Biol. Med. 15: 831-853, 1943.

6. Brooks, C. M. C. The relative importance of changes in activity in the development of experimentally produced obesity in the rat. Am. J. Physiol. 147: 708--716, 1946.

7. Brooks, C. M. C., R. A. Lockwood and M. L. Wiggins. A study of the effect of hypothalamic lesions on the eating habits of the albino rat. Am. J. Physiol. 147: 735-741, 1946.

8. Erdheim, J. Ober Hypophysenganggeschwulste und Hirm cholesteatome. In: Sitzungsb. d.k. Akad. d. Wissensch., Wien 113: 537-726, 1904.

9. Friedman, M. I. and E. M. Stricker. The physiological psychology of hunger. A physiological perspective. Psychol. Rev. 83: 409--431, 1976.

10. FrBhlich, A. Dr. Alfred Fr6hlich stelit einen Fall von Tumor der Hypophyse ohne Akromegalie vor. Wien. Klin. Rundschau 15: 883--886, 1902.

11. Gold, R. M. Hypothalamic obesity: the myth of the ventromedial nucleus. Science 182: 488--489, 1973.

12. Gold, R. M., G. Sumprer, H. M. Ueberacher and G. Kapatos. Hypothalamic hyperphagia despite imposed diurnal or noctur- nal feeding and drinking rhythms. Physiol. Behav. 14: 861-865, 1975.

13. Gold, R. M., A. P. Jones and P. E. Sawchenko. Paraventricular area: critical focus of a longitudinal neurocircuitry mediating food intake. Physiol. Behav. 18: 1111-1119, 1977.

14. Hetherington, A. W. and S. W. Ranson. The spontaneous activity and food intake of rats with hypothalamic lesions. Am. J. Physiol. 136: 609-617, 1942.

15. Kakolewski, J. W., E. Deaux, J. Christensen and B. Case. Di- urnal pattern in water and food intake and body weight changes in rats with hypothalamic lesions. Am. J. Physiol. 221:711-718, 1971.

16. Kissileff, H. R. Free feeding in normal and "recovered lateral" rats monitored by a pellet detecting eatometer. Physiol. Behav. 5: 163-173, 1970.

17. Koizumi, K. and H. Nishino. Circadian and other rhythmic activity of neurons in the ventromedial nuclei and lateral hypothalamic area. J. Physiol. 263: 331-356, 1976.

18. Kreisel, B., N. Conforti, M. Gutnick and S. Feldman. Anti- dromic responses of suprachiasmatic neurons following mediobasal hypothalamic stimulation. Israel J. Med. Sci. 9: 925-927, 1975.

19. Le Magnen, J., M. Deros, J. Gaudilli~re, J. Louis-Sylvestre and S. Tallon. Role of a lipostatic mechanism in regulation by feeding of energy balance in rat. J. comp. physiol. Psychol. 84: 1-23, 1973.

20. Mayer, J. and R. J. Barrnett. Obesity following unilateral hypothalamic lesions in rats. Science 121: 599-600, 1955.

21. Millhouse, O. E. The organization of the ventromedial hypothalamic nucleus. Brain Res. 55: 71-87, 1973.

22. Mohr, B. Hypertrophie der Hypophyse cerebri und dadurch bedingter Druck auf die H/)hergrund fl~iche ins besonders auf die Sehnerven, das Chiasma derselben und dem L/ingseitigen Hfhenschenkel. Wschr. f.d. ges. Heilk. 6: 565-571, 1840.

23. Moore, R. Y. and V. B. Eichler. Loss of a circadian adrenal corticosterone rhythm following suprachiasmatic lesions in the rat. Brain Res. 42: 201-206, 1972.

24. Moore, R. Y. and D. C. Klein. Visual pathways and the central neural control of a circadian rhythm in pineal serotonin N-acetyltransferase activity. Brain Res. 71: 17-33, 1974.

25. Nishino, H. and K. Koizumi. Rhythmic activity of neurons in the lateral hypothalamic area and ventromedial nuclei. Fedn Proc. 34: 424-427, 1975.

26. Nishino, H., K. Koizumi and C. M. C. Brooks. The role of suprachiasmatic nuclei of the hypothalamus in the production of circadian rhythm. Brain Res. 112: 45-59, 1976.

27. Nunez, A. A. and F. K. Stephan. The effects of hypothalamic knife cuts on drinking rhythms and the oestrus cycle of the rat. Behav. Biol. 20: 224--234, 1977.

28. Panksepp, J. Hypothalamic regulation of energy balance and feeding behavior. Fedn Proc. 33:1150-1165, 1974.

29. Richter, C. P. Sleep and activity: their relation to the 24-hour clock. In: Sleep and Altered States of Consciousness. Associa- tion for Research in Nervous and Mental Disease 45: 8-29, 1967.

30. Siegel, S. In: Non-parametric Statistics for the Behavioral Sci- ences. New York: McGraw-Hill, 1956.

31. Stephan, F. K. and I. Zucker. Circadian rhythms in drinking behavior and locomotor activity of rats are eliminated by hypothalamic lesions. Proc. hath. Acad. Sci. U.S.A. 69: 1583- 1586, 1972.

32. Swanson, L. W. and W. M. Cowan. The efferent connections of the suprachiasmatic nucleus of the hypothalamus. J. comp. Neurol. 160: 1-12, 1975.

33. Thomas, D. W. and J. Mayer. Meal taking and regulation of food intake by normal and hypothalamic hyperphagic rats. J. comp. physiol. Psychol. 66: 642-653, 1968.

changes in 24-hour fluctuations of feeding behavior during hypothalamic hyperphagia in rats

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