Physiology & Behavior, Vol. 22, pp. 661-667. PergamonPress and BrainResearch Publ., 1979.Printedin the U.S.A.
The Effect of Various Hunger Manipulations on Self-Stimulation and the Feeding Elicited by It I M. R U S S E K 2 D e p a r t m e n t o f Physiology, National School o f Biological Sciences, National Polytechnic Institute, M~xico, 17 D.F. ( R e c e i v e d 7 A p r i l 1978) RUSSEK, M. The effect of various hunger manipulations on self-stimulation and the feeding elicited by it. PHYSIOL. BEHAV. 22(4) 661--667, 1979.--Out of 16 rats self-stimulating in the lateral hypothalamic feeding area, 11 ingested a liquid food consisting of chocolate milk (stimulation-bound feeding) during self-stimulation even though they had been satiated, while the other five did not. The first group was called "feeders" and the second "non-feeders." In "feeders," adrenaline (0.1-0.2 mg/kg) injected intraperitoneally and stomach loading (20-25 ml) with either 30% glucose or a liquid food (egg-nog) produced a similar marked reduction in the rate of self-stimulation at all but the highest stimulus intensity and an even greater reduction of stimulation-bound feeding. No effect or even an increase in self-stimulation was observed in "nonfeeders." Therefore, the anorexigenic effects of adrenaline appear similar to natural satiation by stomach loading. On the other hand, amphetamine (2 mg/kg) increased the self-stimulation of "non-feeders" and produced a peculiar effect in "feeders": the threshold was higher, but once they started to self-stimulate the rates were higher and they continued even when the stimulus intensity was decreased below the basal threshold. This was interpreted as a mixture of anorexia and a central excitatory effect. Insulin (8-12 U/kg) and 24 hr fasting increased self-stimulation of "non-feeders." However, in "feeders" fasting increased self-stimulation, while insulin decreased it markedly. Self-stimulation
Anorexia
Adrenaline
Amphetamine
T H E RATE of bar pressing for electrical stimulation of the lateral hypothalamus has been shown by several authors [1, 2, 3, 4, 5] to increase in food-deprived animals. These changes are also observed when the animals are injected with substances affecting hunger. Mogenson, Russek and Stevenson [9] observed that rats self-stimulating in the lateral hypothalamus could be assigned to four different categories: (1) those in which selfstimulation induced drinking of either chocolate milk or water when given in a single choice (feeders-drinkers); (2) those that would drink chocolate milk but not water (feeders); (3) those that drank water but very little or no chocolate milk (drinkers); and, (4) those that would not drink either (nonfeeders-nondrinkers). In the first two groups, intraperitoneal adrenaline significantly decreased both the rate of self-stimulation and the amount of water ingested. In the last two groups, the same dose produced no significant change. Therefore, it seemed that intraperitoneal adrenaline affected self-stimulation only when that stimulation induced feeding. In these experiments, however, the rats were implanted with bipolar electrodes, aimed at the site where Mogenson and Stevenson [8] had found optimum electrically-elicited water drinking. The rats were trained and tested for the effect of adrenaline in the presence of water only, and the chocolate milk test was done afterwards in an attempt to find out why some rats re-
Stomach loading
sponded to adrenaline and some did not. The adrenaline decreased self-stimulation rate only in those rats that exhibited stimulation-bound ingestion of chocolate milk. Moreover, the stimulus intensity used for self-stimulation was considerably above the threshold. Thus, a stimulus eliciting a response close to the maximum would not be affected so much by adrenaline, which would explain why the average effect was rather moderate and the variability quite large. In order to analyze the effects of hunger and satiety on self-stimulation and stimulation-bound feeding, the present study was performed. In this work, several parameters were modified from Mogenson et al. [9]. First, monopolar electrodes were used, aimed at a more lateral region where feeding was more likely to be obtained. Second, the rats were offered a double choice between water and chocolate milk while self-stimulating. Third, three or more electrical intensities were used to elicit self-stimulation, the lower one as close to the threshold as possible. Under these conditions the effects of the same doses of adrenaline used previously [9] were tested again. In addition, the effects of amphetamine, insulin, stomach loading, and fasting were also tested. This permitted a comparison between the anorexigenic effects of adrenaline and amphetamine on the one hand, and natural satiation on the other. Furthermore, a comparison could also be made between insulin-induced hunger and natural hunger.
JThis work was sponsored by Tecnofarmaci, S.p.A. (Corporation for the Development of Pharmaceutical Research) Rome, Italy. 2The author is fellow of the COFAA-IPN.
C o p y r i g h t © 1979 B r a i n R e s e a r c h P u b l i c a t i o n s Inc.--0031-9384/79/040661-07502.00/0
662
RUSSEK METHOD
The experiments were carried out in 16 adult male Wistar rats, 250-350 g, that were trained to self-stimulate in either or both lateral hypothalamic areas (A=6; L = 2 ; V=7.5 from cortex, see [13]) where monopolar platinum electrodes were implanted previously under pentobarbital anesthesia (45 mg/kg). The indifferent electrode was wrapped around the skull screws used to anchor the cranioplastic cement to the skull. The stimulation delivered when the animals pressed the bar was a 1.2 sec train of cathodal monophasic rectangular waves (duration 1 msec; frequency I00 Hz; intensity 0.04--0.15 mA from a constant-current stimulator). The lowest intensity that induced prolonged, sometimes intermittent, self-stimulation was considered the threshold. Other intensities were applied, in steps of 0.02 mA. When the animals achieved a relatively constant rate of selfstimulation at each stimulus intensity, they were presented with two spouts, located just above the lever. These were connected to graduated cylinders containing commercial chocolate milk and water, respectively. The rate of selfstimulation was measured by the slope of the line drawn by a Gerbrands cumulative recorder. Each stimulus intensity was given for at least 5 rain, and the amount of chocolate milk drunk during this period (none of the rats drank water) was measured on the graduated cylinder. In addition, the drinking pattern was detected with a drinkometer and recorded on the cumulative record of bar presses by vertical pen deflections. The threshold intensity and 2 to 4 higher intensities were applied before and after the procedure being tested, repeating each intensity for periods of 5 rain or more, at several time intervals. The self-stimulation and stimulation-bound feeding at each intensity were recorded before and after the procedure under study. Priming stimuli were applied at the beginning of each intensity. If the animal did not selfstimulate, the priming was repeated every minute, for the next 5 min. Then the intensity was increased. The effects of the following substances on the selfstimulation and stimulation-bound feeding were tested: intraperitoneal injections of isotonic saline (control), adrenaline 0.1-0.2 mg/kg (Epinephrine hydrochloride, Parke Davis), amphetamine 2 mg/kg (D-amphetamine hydrochloride, National Biochemical Corp.), or insulin 8-12 u/kg (crystalline Zn insulin F . E . U . Eli Lilly and Co.). The effects on self-stimulation and stimulation-bound feeding elicited by the following manipulations were also tested: tube stomach loading with 20-25 ml (90 ml/kg) of water (control), 30% (w/v) glucose, or liquid diet [13], or food-deprivation for 24 hr with water ad lib. The intubation was performed by the procedure used currently to feed aphagic rats [13]. The doses of adrenaline varied two-fold because we tried to obtain a dose/effect curve using 0.1, 0.15 and 0.2 mg/kg. As we did not obtain enough data for each dose, and as the average effect of the three doses was not so much different, we decided to pile all the results together. The same happened with the two doses of insulin. RESULTS
General Of the 16 rats that self-stimulated in one or both lateral hypothalamic areas, 9 were " f e e d e r s . " They consistently exhibited stimulation-bound feeding of chocolate milk during the self-stimulation. They ingested 30 to 40 ml during one
experimental session, which they did not do in the absence of self-stimulation (Fig. 1). Five of the remaining 7 rats were "non-feeders." They self-stimulated with the same range of thresholds and rates but did not feed during it (Fig. 2). There were no rats that drank water during sell-stimulation. The last 2 rats, behaved on some days as " f e e d e r s " and on other days as "non-feeders." When the electrical intensity of the stimulus was increased above a certain level, the stimulation-bound ingestion of the " f e e d e r s " stopped, while the self-stimulation continued at a somewhat lower rate. On the other hand, when the "non-feeders" were stimulated continuously with the same stimulus parameters used for self-stimulation, after 20 to 45 see they began to ingest chocolate milk. Thus, the term "non-feeders" means only that they did not feed during self-stimulation. The thresholds of both types of rats varied slightly from day to day and between animals, but all were in the range of 0.05 to 0.11 mA. The average for the " f e e d e r s " was 0.079 -+ 0.007 and for the "non-feeders" 0.080 _+ 0.009 (~ + cry). The standard errors of the mean for the averages of bar presses per minute ranged from 1.5 to 7 BP/min, varying with the " n " of the sample, so no statistical treatment was necessary to assess the significance of the large changes observed.
The Action of Intraperitoneal Adrenaline and Stomach Loading Intraperitoneal adrenaline elicited in the " f e e d e r s " a substantial reduction in the rate of self-stimulation at all but the highest intensity (Fig. 3), and an even greater reduction in stimulation-bound feeding (Fig. 1). At the threshold intensity (T) the rate was decreased 80% in average (most rats did not bar press at all). At a stimulus intensity of 0.02 mA above threshold the rate decreased about 60% of that obtained before adrenaline (p
The Effect of Various Hunger Manipulations on Self-Stimulation and the Feeding Elicited by It I M. R U S S E K 2 D e p a r t m e n t o f Physiology, National School o f Biological Sciences, National Polytechnic Institute, M~xico, 17 D.F. ( R e c e i v e d 7 A p r i l 1978) RUSSEK, M. The effect of various hunger manipulations on self-stimulation and the feeding elicited by it. PHYSIOL. BEHAV. 22(4) 661--667, 1979.--Out of 16 rats self-stimulating in the lateral hypothalamic feeding area, 11 ingested a liquid food consisting of chocolate milk (stimulation-bound feeding) during self-stimulation even though they had been satiated, while the other five did not. The first group was called "feeders" and the second "non-feeders." In "feeders," adrenaline (0.1-0.2 mg/kg) injected intraperitoneally and stomach loading (20-25 ml) with either 30% glucose or a liquid food (egg-nog) produced a similar marked reduction in the rate of self-stimulation at all but the highest stimulus intensity and an even greater reduction of stimulation-bound feeding. No effect or even an increase in self-stimulation was observed in "nonfeeders." Therefore, the anorexigenic effects of adrenaline appear similar to natural satiation by stomach loading. On the other hand, amphetamine (2 mg/kg) increased the self-stimulation of "non-feeders" and produced a peculiar effect in "feeders": the threshold was higher, but once they started to self-stimulate the rates were higher and they continued even when the stimulus intensity was decreased below the basal threshold. This was interpreted as a mixture of anorexia and a central excitatory effect. Insulin (8-12 U/kg) and 24 hr fasting increased self-stimulation of "non-feeders." However, in "feeders" fasting increased self-stimulation, while insulin decreased it markedly. Self-stimulation
Anorexia
Adrenaline
Amphetamine
T H E RATE of bar pressing for electrical stimulation of the lateral hypothalamus has been shown by several authors [1, 2, 3, 4, 5] to increase in food-deprived animals. These changes are also observed when the animals are injected with substances affecting hunger. Mogenson, Russek and Stevenson [9] observed that rats self-stimulating in the lateral hypothalamus could be assigned to four different categories: (1) those in which selfstimulation induced drinking of either chocolate milk or water when given in a single choice (feeders-drinkers); (2) those that would drink chocolate milk but not water (feeders); (3) those that drank water but very little or no chocolate milk (drinkers); and, (4) those that would not drink either (nonfeeders-nondrinkers). In the first two groups, intraperitoneal adrenaline significantly decreased both the rate of self-stimulation and the amount of water ingested. In the last two groups, the same dose produced no significant change. Therefore, it seemed that intraperitoneal adrenaline affected self-stimulation only when that stimulation induced feeding. In these experiments, however, the rats were implanted with bipolar electrodes, aimed at the site where Mogenson and Stevenson [8] had found optimum electrically-elicited water drinking. The rats were trained and tested for the effect of adrenaline in the presence of water only, and the chocolate milk test was done afterwards in an attempt to find out why some rats re-
Stomach loading
sponded to adrenaline and some did not. The adrenaline decreased self-stimulation rate only in those rats that exhibited stimulation-bound ingestion of chocolate milk. Moreover, the stimulus intensity used for self-stimulation was considerably above the threshold. Thus, a stimulus eliciting a response close to the maximum would not be affected so much by adrenaline, which would explain why the average effect was rather moderate and the variability quite large. In order to analyze the effects of hunger and satiety on self-stimulation and stimulation-bound feeding, the present study was performed. In this work, several parameters were modified from Mogenson et al. [9]. First, monopolar electrodes were used, aimed at a more lateral region where feeding was more likely to be obtained. Second, the rats were offered a double choice between water and chocolate milk while self-stimulating. Third, three or more electrical intensities were used to elicit self-stimulation, the lower one as close to the threshold as possible. Under these conditions the effects of the same doses of adrenaline used previously [9] were tested again. In addition, the effects of amphetamine, insulin, stomach loading, and fasting were also tested. This permitted a comparison between the anorexigenic effects of adrenaline and amphetamine on the one hand, and natural satiation on the other. Furthermore, a comparison could also be made between insulin-induced hunger and natural hunger.
JThis work was sponsored by Tecnofarmaci, S.p.A. (Corporation for the Development of Pharmaceutical Research) Rome, Italy. 2The author is fellow of the COFAA-IPN.
C o p y r i g h t © 1979 B r a i n R e s e a r c h P u b l i c a t i o n s Inc.--0031-9384/79/040661-07502.00/0
662
RUSSEK METHOD
The experiments were carried out in 16 adult male Wistar rats, 250-350 g, that were trained to self-stimulate in either or both lateral hypothalamic areas (A=6; L = 2 ; V=7.5 from cortex, see [13]) where monopolar platinum electrodes were implanted previously under pentobarbital anesthesia (45 mg/kg). The indifferent electrode was wrapped around the skull screws used to anchor the cranioplastic cement to the skull. The stimulation delivered when the animals pressed the bar was a 1.2 sec train of cathodal monophasic rectangular waves (duration 1 msec; frequency I00 Hz; intensity 0.04--0.15 mA from a constant-current stimulator). The lowest intensity that induced prolonged, sometimes intermittent, self-stimulation was considered the threshold. Other intensities were applied, in steps of 0.02 mA. When the animals achieved a relatively constant rate of selfstimulation at each stimulus intensity, they were presented with two spouts, located just above the lever. These were connected to graduated cylinders containing commercial chocolate milk and water, respectively. The rate of selfstimulation was measured by the slope of the line drawn by a Gerbrands cumulative recorder. Each stimulus intensity was given for at least 5 rain, and the amount of chocolate milk drunk during this period (none of the rats drank water) was measured on the graduated cylinder. In addition, the drinking pattern was detected with a drinkometer and recorded on the cumulative record of bar presses by vertical pen deflections. The threshold intensity and 2 to 4 higher intensities were applied before and after the procedure being tested, repeating each intensity for periods of 5 rain or more, at several time intervals. The self-stimulation and stimulation-bound feeding at each intensity were recorded before and after the procedure under study. Priming stimuli were applied at the beginning of each intensity. If the animal did not selfstimulate, the priming was repeated every minute, for the next 5 min. Then the intensity was increased. The effects of the following substances on the selfstimulation and stimulation-bound feeding were tested: intraperitoneal injections of isotonic saline (control), adrenaline 0.1-0.2 mg/kg (Epinephrine hydrochloride, Parke Davis), amphetamine 2 mg/kg (D-amphetamine hydrochloride, National Biochemical Corp.), or insulin 8-12 u/kg (crystalline Zn insulin F . E . U . Eli Lilly and Co.). The effects on self-stimulation and stimulation-bound feeding elicited by the following manipulations were also tested: tube stomach loading with 20-25 ml (90 ml/kg) of water (control), 30% (w/v) glucose, or liquid diet [13], or food-deprivation for 24 hr with water ad lib. The intubation was performed by the procedure used currently to feed aphagic rats [13]. The doses of adrenaline varied two-fold because we tried to obtain a dose/effect curve using 0.1, 0.15 and 0.2 mg/kg. As we did not obtain enough data for each dose, and as the average effect of the three doses was not so much different, we decided to pile all the results together. The same happened with the two doses of insulin. RESULTS
General Of the 16 rats that self-stimulated in one or both lateral hypothalamic areas, 9 were " f e e d e r s . " They consistently exhibited stimulation-bound feeding of chocolate milk during the self-stimulation. They ingested 30 to 40 ml during one
experimental session, which they did not do in the absence of self-stimulation (Fig. 1). Five of the remaining 7 rats were "non-feeders." They self-stimulated with the same range of thresholds and rates but did not feed during it (Fig. 2). There were no rats that drank water during sell-stimulation. The last 2 rats, behaved on some days as " f e e d e r s " and on other days as "non-feeders." When the electrical intensity of the stimulus was increased above a certain level, the stimulation-bound ingestion of the " f e e d e r s " stopped, while the self-stimulation continued at a somewhat lower rate. On the other hand, when the "non-feeders" were stimulated continuously with the same stimulus parameters used for self-stimulation, after 20 to 45 see they began to ingest chocolate milk. Thus, the term "non-feeders" means only that they did not feed during self-stimulation. The thresholds of both types of rats varied slightly from day to day and between animals, but all were in the range of 0.05 to 0.11 mA. The average for the " f e e d e r s " was 0.079 -+ 0.007 and for the "non-feeders" 0.080 _+ 0.009 (~ + cry). The standard errors of the mean for the averages of bar presses per minute ranged from 1.5 to 7 BP/min, varying with the " n " of the sample, so no statistical treatment was necessary to assess the significance of the large changes observed.
The Action of Intraperitoneal Adrenaline and Stomach Loading Intraperitoneal adrenaline elicited in the " f e e d e r s " a substantial reduction in the rate of self-stimulation at all but the highest intensity (Fig. 3), and an even greater reduction in stimulation-bound feeding (Fig. 1). At the threshold intensity (T) the rate was decreased 80% in average (most rats did not bar press at all). At a stimulus intensity of 0.02 mA above threshold the rate decreased about 60% of that obtained before adrenaline (p
