Continuous infusion of cholecystokinin and meal pattern in the rat

Purified natural cholecystokinin (CCK-33) was infused continuously for two days at a rate of 5.9 μg/hr in two rats trained to bar-press for food (Noyes pellet 45 mg) on a fixed ratio of five bar presses to obtain one pellet. The animals also received control surgery and were tested in the operant chamber for two days, one prior to and the other following the CCK-33 treatment. CCK-33 suppressed the number of meals, the total amount of food eaten, and the total duration of time spent eating. However, the size of each meal and the rate of intake were not affected. The CCK effect did not interact with the light-dark phases of diurnal cycle. It appears that a major effect of continuous systemic elevation of CCK-33 is to reduce food intake by prolonging the satiety period rather than by decreasing the individual meal size.     

Interoceptive "satiety" signals produced by leptin and CCK

The present studies assessed the extent to which the adiposity signal leptin and the brain-gut hormone cholecystokinin (CCK), administered alone or in combination, give rise to interoceptive sensory cues like those that are produced by a low (1h) level of food deprivation. Rats were trained with cues arising from 1 to 24-h food deprivation as discriminative stimuli. For one group, 24-h food deprivation predicted the delivery of sucrose pellets, whereas 1-h food deprivation did not. Another group received the reversed deprivation level-sucrose contingency. After asymptotic performance was achieved, the effects of leptin and CCK on food intake and on discrimination performance were tested under 24-h food deprivation. In Experiment 1a, leptin administered into the third cerebroventricle (i3vt) at 3.5 or 7.0 microg doses had little effect, compared to saline on food intake or discriminative responding. In Experiment 1b, leptin (7.0 microg, i3vt) combined with CCK-8 (2 microg/kg, i.p.) reduced food intake significantly, but the findings indicated that CCK-8 alone produces interoceptive discriminative cues more like those produced by 1- than 24-h food deprivation. Experiment 2a tested rats with i.p. leptin (0.3 and 0.5mg/kg). Although neither dose suppressed intake, the 0.3mg/kg dose produced interoceptive cues like 1-h food deprivation. Experiment 2b tested two doses of CCK-8 (2 and 4 mg/kg, i.p.) and found significant intake suppression and generalization of discrimination with both doses of CCK-8. These findings suggest a role for both leptin and CCK in the production of sensory consequences that correspond to "satiety".     

Variation of bar-press duration: Where do new responses come from?

Instrumental learning involves both variation and selection: variation of what the animal does, and selection by reward from among the variation. Four experiments with rats suggested a rule about how variation is controlled by recent events. Experiment 1 used the peak procedure. Measurements of bar-press durations showed a sharp increase in mean duration after the time that food was sometimes given. The increase was triggered by the omission of expected food. Our first explanation of the increase was that it was a frustration effect. Experiment 2 tested this explanation with a procedure in which the first response of a trial usually produced food, ending the trial. In Experiment 2, unlike Experiment 1, omission of expected food did not produce a large increase in bar-press duration, which cast doubt on the frustration explanation. Experiments 3 and 4 tested an alternative explanation: a decrease in expectation of reward increases variation. Both used two signals associated with different probabilities of reward. Bar presses were more variable in duration during the signal with the lower probability of reward, supporting this alternative. These experiments show how variation can be studied with ordinary equipment and responses.     

Endogenous amylin contributes to the anorectic effects of cholecystokinin and bombesin

Previous studies indicated that amylin contributes to the anorectic effects of cholecystokinin (CCK) and bombesin (BBS), possibly by enhancing the release of pancreatic amylin or by modulating their anorectic actions within the central nervous system (CNS). To elucidate the interaction between amylin and CCK or BBS, respectively, we investigated the influence of an IP injection of CCK or BBS on feeding in amylin-deficient mice (IAPP(-/-)). The anorectic effects of CCK and BBS were nearly abolished in IAPP(-/-) mice compared to wildtype (WT) mice (e.g. 20 microg/kg CCK, 1-h food intake: WT/NaCl 0.53 +/- 0.03 g; WT/CCK 0.16 +/- 0.03 g (P < 0.001); IAPP(-/-)/NaCl 0.49 +/- 0.05 g; IAPP(-/-)/CCK 0.39 +/- 0.04 g). Acute amylin replacement restored the anorectic effect of CCK in IAPP(-/-) mice.To find out whether CCK or BBS enhance the feeding-induced release of pancreatic amylin, we injected rats with CCK-8 (0.5-50 microg/kg) or BBS (5 microg/kg) and measured plasma amylin levels after injections. Neither CCK nor BBS increased the plasma amylin level in rats. We suggest that the mediation of the anorectic effects of CCK and BBS by amylin is not dependent on a CCK- or BBS-induced release of pancreatic amylin, but may rather be due to a modulation of their effects by amylin within the CNS.     

Capsaicin does not attenuate bombesin-induced suppression of operant responding for food reward.

Systemic treatment with capsaicin, a neurotoxin which damages unmyelinated peptide-containing sensory neurons, has been shown to attenuate bombesin (BBS)-induced suppression of food intake. To determine whether capsaicin-sensitive fibers mediate the effect of BBS on appetitive motivation, we examined BBS-induced suppression of operant responding in rats pretreated neonatally with capsaicin (50 mg/kg; SC) or control vehicle. At 8-10 weeks of age, rats were trained to bar press for food. After achieving a stable level of performance, the animals were injected with BBS (10 micrograms/kg), normal saline, or prefed with 20 Noyes 45-mg pellets. Animals were then tested in an operant chamber on an FR 5 schedule of reinforcement for one hour. The results indicated that BBS suppressed bar pressing, regardless of whether animals were pretreated with capsaicin or control vehicle. These findings are inconsistent with the hypothesis that BBS induces satiety via capsaicin-sensitive neurons. The results suggest the possibility that more than one mechanism may mediate the effects of BBS: a neural mechanism involved in consummatory responses and a humoral mechanism involved in the operant response.     

The Effect of Ratio and Interval Training on Pavlovian-Instrumental Transfer in Mice

Conditional stimuli (CS) that are paired with reward can be used to motivate instrumental responses. This process is called Pavlovian-instrumental transfer (PIT). A recent study in rats suggested that habitual responses are particularly sensitive to the motivational effects of reward cues. The current experiments examined this idea using ratio and interval training in mice. Two groups of animals were trained to lever press for food pellets that were delivered on random ratio or random interval schedules. Devaluation tests revealed that interval training led to habitual responding while ratio training produced goal-directed actions. The presentation of CSs paired with reward led to positive transfer in both groups, however, the size of this effect was much larger in mice that were trained on interval schedules. This result suggests that habitual responses are more sensitive to the motivational influence of reward cues than goal-directed actions. The implications for neurobiological models of motivation and drug seeking behaviors are discussed.     

High-fat diet is associated with blunted splanchnic sympathoinhibitory responses to gastric leptin and cholecystokinin: implications for circulatory control

Gastric leptin and cholecystokinin (CCK) act on vagal afferents to induce cardiovascular effects and reflex inhibition of splanchnic sympathetic nerve discharge (SSND) and may act cooperatively in these responses. We sought to determine whether these effects are altered in animals that developed obesity in response to a medium high-fat diet (MHFD). Male Sprague-Dawley rats were placed on a low-fat diet (LFD; n = 8) or a MHFD (n = 24) for 13 wk, after which the animals were anesthetized and artificially ventilated. Arterial pressure was monitored and blood was collected for the determination of plasma leptin and CCK. SSND responses to leptin (15 μg/kg) and CCK (2 μg/kg) administered close to the coeliac artery were evaluated. Collectively, MHFD animals had significantly higher plasma leptin but lower plasma CCK levels than LFD rats (P < 0.05), and this corresponded to attenuated or reversed SSND responses to CCK (LFD, -21 ± 2%; and MHFD, -12 ± 2%; P < 0.05) and leptin (LFD, -6 ± 2%; and MHFD, 4 ± 1%; P < 0.001). Alternatively, animals on the MHFD were stratified into obesity-prone (OP; n = 8) or obesity-resistant (OR; n = 8) groups according to their weight gain falling within the upper or lower tertile, respectively. OP rats had significantly higher resting arterial pressure, adiposity, and plasma leptin but lower plasma CCK compared with LFD rats (P < 0.05). The SSND responses to CCK or leptin were not significantly different between OP and OR animals. These results demonstrate that a high-fat diet is associated with blunted splanchnic sympathoinhibitory responses to gastric leptin and CCK and may impact on sympathetic vasomotor mechanisms involved in circulatory control.     

Gastrin releasing peptide reduces meal size in rats

The satiety-eliciting effect of gastrin-releasing peptide (GRP), a putative mammalian counterpart of bombesin (BBS), was examined in mildly food-deprived rats. Intraperitoneal injections of GRP resulted in a significant decrease of 30-minute food intake at 2, 4, 8 and 16 μg/kg, while 1 μg/kg had no reliable effect. Intraperitoneal GRP at 4 and 8 μg/kg did not suppress 30-minute water consumption by thirsty rats. When the dose-effect curves of GRP and BBS are compared on a molar scale, GRP is approximately 30% less potent than BBS in suppressing food intake. The two dose-effect curves are similar in shape and their regression lines have parallel slopes. These data lend further support to the hypothesis that GRP is a mammalian counterpart of BBS and strengthen the argument that they may function as endogenous satiety factors.     

Instrumental responding remains under the control of the consequent outcome after extended training

The effect of extensive training on the contribution of response--outcome learning to instrumental performance in rats was re-examined in two experiments using a transfer test. In each experiment, two discriminative stimuli were established as signals for different response--outcome combinations (e.g. light: nose poke-pellets and noise: handle pull-sucrose). Then, two different responses (lever press and chain pull) were concurrently trained with those outcomes. In Experiment 1, these responses underwent extensive training, each with a different outcome (e.g. lever press-pellets and chain pull-sucrose). In Experiment 2, these responses were trained moderately with one outcome (e.g. lever press-pellets and chain pull-sucrose) and extensively with a different outcome (e.g. lever press-sucrose and chain pull-pellets). Finally, transfer tests were conducted in which the discriminative stimuli, noise and light, were tested periodically with the lever and chain. In Experiment 1, the stimuli consistently and preferentially elevated performance of the response trained with the same outcome relative to that trained with a different outcome. In Experiment 2, the stimuli elevated both responses nondifferentially. However, a drive manipulation (thirst) designed to increase the value of sucrose relative to pellets revealed a significant preference for the response that had been extensively trained with sucrose. Overall, the results of these experiments confirm previous findings that instrumental behaviors do not become increasingly independent of their consequent outcomes with extended training. However, the transfer results of Experiment 2 highlight a potential limitation of the transfer test for assessing variations in the strength of R--O associations.     

Effect of peripheral administration of cholecystokinin on food intake in apolipoprotein AIV knockout mice

Apolipoprotein AIV (apo AIV) and cholecystokinin (CCK) are satiation factors secreted by the small intestine in response to lipid meals. Apo AIV and CCK-8 has an additive effect to suppress food intake relative to apo AIV or CCK-8 alone. In this study, we determined whether CCK-8 (1, 3, or 5 μg/kg ip) reduces food intake in fasted apo AIV knockout (KO) mice as effectively as in fasted wild-type (WT) mice. Food intake was monitored by the DietMax food system. Apo AIV KO mice had significantly reduced 30-min food intake following all doses of CCK-8, whereas WT mice had reduced food intake only at doses of 3 μg/kg and above. Post hoc analysis revealed that the reduction of 10-min and 30-min food intake elicited by each dose of CCK-8 was significantly larger in the apo AIV KO mice than in the WT mice. Peripheral CCK 1 receptor (CCK1R) gene expression (mRNA) in the duodenum and gallbladder of the fasted apo AIV KO mice was comparable to that in WT mice. In contrast, CCK1R mRNA in nodose ganglia of the apo AIV KO mice was upregulated relative to WT animals. Similarly, upregulated CCK1R gene expression was found in the brain stem of apo AIV KO mice by in situ hybridization. Although it is possible that the increased satiating potency of CCK in apo AIV KO mice is mediated by upregulation of CCK 1R in the nodose ganglia and nucleus tractus solitarius, additional experiments are required to confirm such a mechanism.     

Stimulation of either cholecystokinin receptor subtype reduces while antagonists potentiate or sensitize a morphine-induced excitatory response.

Cholecystokinin peptides (CCK) have been shown to antagonize many opioid-mediated effects. The present study was undertaken to determine whether peripheral injections of cholecystokinin sulphated octapeptide (CCK8), cholecystokinin tetrapeptide (CCK4), the CCK(1) (lorglumide) and the CCK(2) (PD-135,158 and LY-225910) receptor antagonists can influence a classic morphine excitatory effect, i.e. the display of Straub tail reaction in mice (STR). A total of 570 female Balb/C mice were tested. Experiment 1 was undertaken to determine whether i.p. injections of CCK8 or CCK4 can influence STR. Each animal was treated with i.p. injections of saline or CCK8 (10 and 20 nmol/kg) or CCK4 (20 and 40 nmol/kg). After 30 min all animals received an i.p. injection of morphine hydrochloride (10.0 mg/kg). The highest doses of both CCK8 (35% STR) and CCK4 (40% STR) significantly reduced STR as compared to saline (85% STR) treated mice (Fisher test; P < 0.01). In experiment 2 each animal was treated with ip injections of saline or 1.0 mg/kg lorglumide or PD-135,158 fifteen minutes before an injection of morphine at doses ranging from 1.0 to 50.0 mg/kg. In experiment 3 animals were treated with injections of saline, 0.1 or 10.0 mg/kg lorglumide or LY-225910 before an injection of a fixed MC dose (2.0 mg/kg). Both lorglumide and PD-135,158 induced a significant shift to the left in the morphine dose-response curves as well as a significant decrease in ED50 of the STR. ED50 for lorglumide was significantly lower than ED50 for PD-135,158. Both doses of lorglumide and the highest dose of LY-225910 significantly increased the percent of animals displaying STR. Experiment 4 was undertaken to determine whether repeated peripheral injections of morphine or the morphine-potentiating agents CCK(1) (lorglumide) and the CCK(2) (LY-225910) receptor antagonists can induce morphine sensitization. Each animal was treated with 5 daily i.p. injections of saline (control group), 1.5 mg/Kg morphine hydrochloride (group morphine), and 1.0 mg/Kg lorglumide (group LOR) or LY-225910 (group LY). One, two, three and four weeks after the last treatment day, all animals were challenged with one i.p. injection of morphine (1.5 mg/Kg). The morphine, LOR groups and group LY showed a significant increase in percentage of animals displaying STR. These data demonstrate that the blockade of endogenous CCK actions leads to morphine sensitization probably through both CCK receptors. The present data are consistent with the antagonistic effects of CCK and opioids in the control of morphine-induced STR. In addition, these results suggest that both CCK receptors are involved in the modulatory effects of CCK on this morphine effect.     

Effects of cholecystokinin octapeptide (CCK-8) on food intake in adult and aged rats under different feeding conditions

The effects of CCK on food intake were investigated under fixed feeding conditions in comparison to a test meal taken after 16 h of food deprivation. The experiments were performed on young adult rats (8 weeks old) as well on aged rats (23 months old). Intraperitoneal CCK-8 (8 and 40 μg/kg) significantly reduced the size of a test meal following 16-h food deprivation. This effect was independent of the age of the rats. However, under fixed feeding conditions neither of the doses used in this study reduced food intake in the young adult rats, whereas the highest dose of 40 μg/kg did so in the aged rats. These results suggest that the hypophagic effect of exogenous CCK-8 depends on experimental conditions, food intake being reduced after a period of food deprivation but not under a fixed feeding regimen in adult animals. Furthermore, the data suggest that age is a factor contributing to the complex behavioral actions of CCK, because only old animals were more susceptible to an anorectic action of CCK under the fixed feeding schedule. An explanation may lie in an interaction of other known behavioral effects of CCK (e.g., anxiogenic, mnemonic action) with its effects under the different feeding schedules.     

The inhibitory effect of pretrial reinforcement on instrumental conditioning

It has been shown previously that pretrial feeding retards acquisition of an instrumental response. The present experiment tested three hypotheses for this effect: priming, overshadowing and conditioned inhibition. Rats were trained in a runway task with a partial schedule of food reinforcement. Rats that were prefed immediately prior to rewarded trials showed a decrement in acquisition when compared with subjects prefed 180 seconds pretrial. Other subjects prefed immediately prior to nonrewarded trials showed a similar decrement in acquisition, even though the pretrial and goal reinforcers occurred on separate trials. Thus, the adverse effects of prefeeding occurred in the absence of conditions necessary for priming or overshadowing to occur. The results suggest there may be an inhibitory aftereffect of food rewards which can become conditioned to subsequent stimuli.     

Bar-pressing for water reward: effects of nootropic drugs and peptides on discrimination learning in rats

Rats maintained on 23-hr water deprivation were first trained to bar-press for continuous water reinforcement and then to discriminate between regularly alternating periods (24 sec) during which time a light signal was either on and each response was reinforced or the light was off and bar-presses were not rewarded. The following drugs were injected s. c. prior to the sessions of discriminative learning: piracetam, 1-(4-Methyl-piperazinocarbonylmethyl)-2-pyrrolidone/hydrogen maleate (VUFB 13763), N alpha-glycyl-glycyl[8-lysine]des-9-glycinamide-vasopressin (DG-Trigly-LVP) and an analog of MIF, EUC-Leu-beta-Ala-NH2 (EUC, 2-oxoimidazolidine-1-carboxylic acid). None of the drugs influenced the total number of bar-pressing (sum of reinforced and non reinforced responses). Piracetam (100 mg.kg-1), VUFB 13763 (40 mg.kg-1) and EUC-Leu-beta-Ala-NH2 (1 mg.kg-1) improved the performance of rats on the discrimination learning task, DG-Trigly-LVP slowed the rate of acquisition.     

CCK-resistance in Zucker obese versus lean rats

Obese Zucker rats are less sensitive to the satiety effect of CCK than lean litter mates. The present studies further characterised this CCK resistance. Subcutaneous injection of the CCK agonist caerulein dose-dependently decreased food intake in Zucker obese and lean rats whereas the CCK-B agonist gastrin-17 did not. Caerulein at 4 μg/kg, which resulted in CCK plasma bioactivity slightly above postprandial levels, decreased food intake in lean rats but not in obese rats. The decrease in food intake was also more marked at higher caerulein doses (20–100 μg/kg) in lean versus obese rats. In lean animals the satiety effects of the “near physiological” 4 μg/kg caerulein dose was abolished after blockade of vagal afferents with capsaicin, whereas the effects of higher caerulein doses were not. CCK-stimulated amylase secretion from pancreatic acini and binding capacity of ¹²⁵I- labelled CCK-8 were decreased in obese versus lean rats. The CCK-A antagonist loxiglumide at 20 mg/kg, a dose which abolished the action of all caerulein doses on food intake, failed to alter the food intake either in obese or in lean rats when given without an agonist. The results suggest that the satiety effects of “near physiological” doses of caerulein in lean rats are mediated by vagal afferents whereas pharmacological doses act via non-vagal mechanisms. The differences in CCK's satiety effect between lean and obese rats may be due to differences in CCK-receptor binding and action at peripheral vagal sites. However, the failure of the CCK-A antagonist to increase food intake questions whether any of the effects of exogenous CCK are of physiological relevance.     

Satiety: the roles of peptides from the stomach and the intestine

Rats were surgically prepared to allow perfusions of anatomically limited portions of the gastrointestinal (GI) surface during test meals. The results demonstrated that at least one potent satiety signal was generated when ingested food accumulated in the stomach and did not enter the small intestine. This gastric satiety signal did not require the vagus nerve for its operation. In addition, at least one other potent satiety signal was generated when food perfused the small intestine. This intestinal satiety signal did not require gastric distension for its operation. We tested a variety of GI peptides to determine whether any met the criteria imposed by this evidence for regionally specific satiety signals. Bombesin (BBS), a peptide present in high concentration in the stomach, was a potent and behaviorally specific inhibitor of food intake. Its satiating effect was not altered by subdiaphragmatic vagotomy. Cholecystokinin (CCK), a peptide hormone that is released from the small intestine by food, was also a potent and behaviorally specific inhibitor of food intake; its satiating effect did not require gastric distension for its expression, but its satiating effect was markedly reduced or abolished by subdiaphragmatic vagotomy. Thus, BBS and CCK may mediate at least part of the satiating effect of food acting in the stomach and in the small intestine, respectively.     

Abrogation of peripheral cholecystokinin-satiety in the capsaicin treated rat

Cholecystokinin (CCK) is a peripheral and central mediator of short-term satiety. When given i.p., CCK decreases food intake in previously fasted rats for a period of 30 min. The effect has been previously shown to be abolished by vagotomy and more specifically by severing of vagal sensory rootlets. These studies were designed to determine the effects on rat feeding behavior, and in particular CCK-satiety, of the sensory neurotoxin capsaicin. In neonates, capsaicin selectively and permanently destroys unmyelinated sensory fibers including those in the vagus nerve. Rat neonates were treated with capsaicin, 50 mg/kg or vehicle, and surviving females studied at 8-10 weeks of age. The weights, 24-h food intake, and feeding responses to insulin were the same in adult capsaicin treated (Cap Rx) and vehicle treated (Veh Rx) rats. CCK (8 micrograms/kg i.p.) reduced 30 min food intake 61 +/- 18% in Veh Rx animals (mean +/- S.D., P less than 0.01). In capsaicin denervated animals, CCK also significantly reduced 30 min food intake from 5.09 +/- 1.10 to 3.92 +/- 0.84 g (P less than 0.01), but the mean reduction, 23 +/- 6%, was significantly less than in Veh Rx rats (P less than 10(-4]. A separate group of females, similarly treated as neonates with capsaicin or vehicle, were subjected to bilateral lesioning of the ventromedial hypothalamus. Both Cap Rx and Veh Rx animals gained significantly and equally more than non-lesioned controls. 24 h vagal transport of substance P was reduced 70% in age matched capsaicin treated animals compared to controls. These studies demonstrate that peripheral CCK-satiety is partly mediated by capsaicin sensitive fibers, presumably in the vagus nerve. Substance P is one possible transmitter mediating this reflex. Further conclusions are that active inhibition of an intact peripheral CCK-stimulated reflex arc is not necessary for full expression of central inducers of feeding, e.g., insulin or lesioning of the ventromedial hypothalamus, and that destruction of these fibers does not alter long-term weight regulation in rats receiving a normal diet.     

Do peptide-induced changes in feeding occur because of changes in motivation to eat?

The usual paradigm in which peptides are tested for their effect on food intake involves measuring intake of readily available food. In the lever press apparatus, the subjects must "work" to get food. Such work has traditionally been used as a means of measuring motivation. Mice were trained to press a lever for milk reinforcement. After achieving a stable level of performance, we tested the effects of gastrin-related peptide (GRP), bombesin (BBS) and cholecystokinin octapeptide (CCK-8) on lever pressing. All three peptides suppressed lever pressing for milk reinforcement. Prefeeding mice with milk increased the suppression of lever pressing to a greater extent in peptide-treated mice than in saline-treated mice. As the duration of prefeeding increased, lever pressing decreased. When mice were required to make more lever presses to obtain milk, both saline- and CCK-8-treated mice increased their lever pressing. However, saline-treated mice pressed at a higher rate than CCK-8-treated mice. Unlike the results obtained with saline and CCK-8, administration of a known gustatory adversant, lithium chloride, suppressed lever pressing to the same degree in mice fed or not fed prior to training. The results are consistent with the hypothesis that these peptides act as satiety agents.     

Effect of short-term prefeeding and body weight on wheel running and responding reinforced by the opportunity to run in a wheel

A biobehavioural analysis of activity anorexia suggests that the motivation for physical activity is regulated by food supply and body weight. In the present experiment, food allocation was varied within subjects by prefeeding food-deprived rats 0, 5, 10 and 15 g of food before sessions of lever pressing for wheel-running reinforcement. The experiment assessed the effects of prefeeding on rates of wheel running, lever pressing, and postreinforcement pausing. Results showed that prefeeding animals 5 g of food had no effect. Prefeeding 10 g of food reduced lever pressing for wheel running and rates of wheel running without a significant change in body weight; the effect was, however, transitory. Prefeeding 15 g of food increased the animals' body weights, resulting in a sustained decrease of wheel running and lever pressing, and an increase in postreinforcement pausing. Overall the results indicate that the motivation for physical activity is regulated by changes in local food supply, but is sustained only when there is a concomitant change in body weight.     

Combined stimulation of glucagon-like peptide-1 receptor and inhibition of cannabinoid CB1 receptor act synergistically to reduce food intake and body weight in the rat

Pharmacological activation of the glucagon-like peptide-1 (GLP-1) receptor and inhibition of the cannabinoid CB1 receptor were found to reduce food intake and body weight in humans and animals. Since earlier studies revealed that endocannabinoids may interact with other neurotransmitters to affect feeding behavior, we have examined whether a stable GLP-1 agonist, exendin-4 and a CB1 receptor antagonist, AM 251, may reciprocally enhance their inhibitory effects on food consumption in the rat. Additionally, we have tested whether the blockade of the GLP-1 receptor by exendin (9-39) modifies AM 251-dependent effects on energy balance. In a dose-response study, male Wistar rats were injected intraperitoneally with either 1.5-6.0 μg/kg exendin-4, 0.5-2 mg/kg AM 251, 80-320 μg/kg exendin (9-39) or their vehicle and the daily food and water intake as well as body weight changes were monitored two days before and two days after the injection. Exendin-4 at a dose of 3.0 and 6.0 μg/kg and AM 251 at a dose 2 mg/kg decreased significantly 24-hour food intake and body weight. Therefore, in the next study, the effects of lower doses of exendin-4 (1.5 μg/kg) and AM 251 (1.0 mg/kg) administered alone or together on food consumption were compared. As opposed to being injected alone, the co-administration of the two resulted in a marked decrease in both daily food intake and body weight. Exendin (9-39) did not modify the suppressory effect of the highest AM 251 dose on food consumption. Apparently, the effect of AM 251 on the appetite is not mediated by GLP-1. The concomitant stimulation of GLP-1 receptor and blockade of CB1 receptor, however, may act synergistically to inhibit appetite in the rat.