Medial versus lateral parabrachial nucleus lesions in the rat: effects on cholecystokinin- and -fenfluramine-induced anorexia

The two major components of the pontine parabrachial nucleus (PBN), the medial (gustatory) and lateral (visceral) subdivisions, have been implicated in a variety of ingestive behaviors. The present study examined the influence of bilateral ibotenic acid lesions of the medial or lateral PBN on the anorectic effects of two systemically administered drug treatments. In Experiment 1, 24-h food-deprived rats where injected with sulfated cholecystokinin_26-33 (CCK; 0, 4.0, or 8.0 μg/kg) and then given 60 min access to food. In Experiment 2, the influence of -fenfluramine (DFEN; 0, 0.5, 1.0, or 2.0 mg/kg) on deprivation-induced feeding was examined in the same rats using the same behavioral procedure as in Experiment 1. Lesions of the lateral PBN abolished CCK-, but not DFEN-induced anorexia whereas lesions of the medial PBN augmented DFEN-, but had no influence on CCK-induced anorexia. The results suggest that the satiating effects of CCK and DFEN are mediated through different mechanisms involving, respectively, visceral and orosensory processing.     

Medial versus lateral parabrachial nucleus lesions in the rat: effects on cholecystokinin- and D-fenfluramine-induced anorexia

The two major components of the pontine parabrachial nucleus (PBN), the medial (gustatory) and lateral (visceral) subdivisions, have been implicated in a variety of ingestive behaviors. The present study examined the influence of bilateral ibotenic acid lesions of the medial or lateral PBN on the anorectic effects of two systemically administered drug treatments. In Experiment 1, 24-h food-deprived rats where injected with sulfated cholecystokinin (26-33) (CCK; 0, 4.0, or 8.0 microg/kg) and then given 60 min access to food. In Experiment 2, the influence of D-fenfluramine (DFEN; 0, 0.5, 1.0, or 2.0 mg/kg) on deprivation-induced feeding was examined in the same rats using the same behavioral procedure as in Experiment 1. Lesions of the lateral PBN abolished CCK-, but not DFEN-induced anorexia whereas lesions of the medial PBN augmented DFEN-, but had no influence on CCK-induced anorexia. The results suggest that the satiating effects of CCK and DFEN are mediated through different mechanisms involving, respectively, visceral and orosensory processing.     

Capsaicin pretreatment attenuates multiple responses to cholecystokinin in rats

Systemic injection of the peptide hormone cholecystokinin (CCK) is known to inhibit food intake and gastric emptying, and to stimulate neurohypophyseal secretion of oxytocin (OT) in rats. Previous studies also have shown that surgical destruction of afferent fibers in the gastric vagus eliminates the inhibitory effects of CCK on food intake. The present experiments used capsaicin to destroy peripheral sensory fibers in rats, and confirmed the failure of CCK to inhibit food intake. Similarly, capsaicin pretreatment markedly attenuated the stimulatory effect of CCK on OT secretion and the inhibitory effect of CCK on gastric emptying in rats. These and other results suggest that in rats CCK acts on receptors located on afferent fibers in the gastric vagus and stimulates inhibition of gastric emptying predominantly via a vagovagal reflex arc through the brainstem.     

C-fos protein expression in the nucleus of the solitary tract correlates with cholecystokinin dose injected and food intake in rats

C-fos protein expression was investigated in the nucleus of the solitary tract (NTS) in response to increasing cholecystokinin (CCK) doses and food intake in rats by counting the number of c-fos protein positive cells in the NTS. C-fos protein expression in the NTS dose-dependently increased in response to CCK, the lowest effective dose being 0.1 μg/kg. The ED₅₀ for c-fos protein expression in the NTS in response to CCK was calculated to be 0.5 to 1.8 μg/kg, depending on the anatomical level of the NTS investigated. Food intake increased c-fos protein expression in the NTS, the maximum number of c-fos protein positive cells being reached at 90 min after the start of food intake. Regression analysis identified a positive correlation between c-fos protein expression and the amount of food intake. Our data indicate that subpopulations of the NTS that are activated by CCK or food intake are involved into the short-term regulation of food intake and the neural control of feeding by the caudal brainstem.     

Area postrema: site where cholecystokinin acts to decrease food intake

Administration of cholecystokinin (CCK) reduces food intake in rats. This effect of CCK was attenuated in rats with thermal lesions of the area postrema. This result was specific to CCK, as area postrema lesions had no effect on the reduction in food intake produced by amphetamine. The effect of the lesions was not due to changes in taste sensation as the lesioned rats showed no deficits in morphine induced conditioned taste aversions. Thus, the area postrema may be the major site where CCK acts to decrease food intake.     

Loss of acute satiety response to cholecystokinin in pregnant rats

During pregnancy, food intake and fat mass are increased to meet the energy demands of the growing conceptus and to prepare for the subsequent demands of lactation. A state of leptin resistance develops during pregnancy in the rat, which can facilitate the increase in food intake despite pregnancy-induced increases in leptin concentrations. Cholecystokinin (CCK) is a satiety factor that is released from the gut during feeding and acts to terminate short-term food intake. Circulating leptin concentrations can modulate the anorexic response to CCK; low leptin concentrations decrease the potency of CCK to reduce food intake. Because rats are leptin resistant by day 14 of pregnancy, it was hypothesised that the feeding response to CCK would be attenuated at that time. Nonpregnant and day 14 pregnant rats received an i.p. injection of CCK-8 (3 μg/kg body weight) or vehicle directly before the start of the dark phase. Food intake was measured 30 min after lights out. Approximately 90 min after receiving either CCK-8 or vehicle, rats were transcardially perfused with 4% paraformaldehyde. Food intake was significantly decreased in CCK-treated nonpregnant rats, although similar treatment did not reduce food intake in day 14 pregnant rats. CCK treatment lead to significant increased in c-Fos expression in the nucleus of the solitary tract (NTS) in both nonpregnant and pregnant rats compared to vehicle treatment, although the number of CCK-induced c-Fos positive cells was significantly less in pregnant rat compared to nonpregnant rats. Although CCK treatment increased the number of c-Fos positive cells in the hypothalamic paraventricular nucleus and supraoptic nucleus in nonpregnant rats, no significant increase was observed in these areas during pregnancy. These results indicate that pregnant rats are no longer responsive to the actions of CCK on short-term food intake and that CCK action in the NTS is reduced during pregnancy.     

Fos expression in the brain induced by peripheral injection of CCK or leptin plus CCK in fasted lean mice

We previously reported a synergistic interaction between leptin and cholecystokinin (CCK) to reduce food intake through CCK-A receptors in lean mice fasted for 24 h. To identify the activated neuronal pathways, we investigated changes in Fos expression in brain nuclei 2 h after single or combined intraperitoneal (i.p.) injections of leptin (120 μg/kg) and sulfated CCK-8 (3.5 μg/kg) in male lean mice (C57BL/6) fasted for 24 h using immunohistochemistry for Fos, the protein product of the early gene, c-fos. Leptin did not increase Fos expression in the brain compared with vehicle-treated mice. CCK increased the numbers of Fos-positive neurons in the nucleus of the solitary tract (NTS)/area postrema (AP), central nucleus of the amygdala (CeA) and, to a smaller extent, in the paraventricular nucleus of the hypothalamus (PVN) (5.2-, 2.3- and 0.3-fold respectively). Injections of leptin–CCK further enhanced Fos expression by 40% in the PVN compared with that induced by CCK alone, but not in the other nuclei. Devazepide (a CCK-A receptor antagonist, 1 mg/kg, i.p.) prevented the increase in Fos expression induced by leptin–CCK in the PVN and by CCK alone in the PVN, CeA and NTS/AP. These results indicate that in fasted mice, i.p. injection of CCK increases Fos expression in specific brain nuclei through CCK-A receptors while leptin alone had no effect. Leptin in conjunction with CCK selectively enhanced Fos expression in the PVN. The PVN may be an important site mediating the synergistic effect of leptin–CCK to regulate food intake.     

Responses to cholecystokinin in the ventromedial nucleus of the rat hypothalamus in vivo

The peptide cholecystokinin (CCK) is a short‐term satiety signal released from the gastrointestinal tract during food intake. From the periphery, CCK signalling travels via the vagus nerve to reach the brainstem from which it is relayed higher into the brain. The hypothalamus is a key integrator of appetite‐related stimuli and the ventromedial nucleus of the hypothalamus (VMN) is thought to have an important role in the regulation of satiety. We investigated the effect of intravenous injections of CCK on the spontaneous firing activity of single VMN neurons in urethane‐anaesthetised rats in vivo. We found that the predominant effect of CCK on the electrical activity in the VMN is inhibitory. We analysed the responses to CCK according to electrophysiologically distinct subpopulations of VMN neurons and found that four of these VMN subpopulations were inhibited by CCK, while five were not significantly affected. Finally, CCK‐induced inhibitory response in VMN neurons was not altered by pre‐administration of intravenous leptin.     

Examining the effects of lipopolysaccharide and cholecystokinin on water ingestion: comparing intake and palatability

Lipopolysaccharide (LPS) and cholecystokinin (CCK) have been shown to have anorectic properties in a variety of species. The present study examined the effects of LPS and CCK, both alone and in combination, on two different aspects of water ingestion, water intake and palatability. On test days, animals were first injected intraperitoneally (i.p.) with either LPS (200 microg/kg) or NaCl vehicle, and 2 h later received a second injection of either CCK (8 microg/kg) or NaCl vehicle. In Experiment 1, water intake was monitored for 1 h on 3 separate test days 72 h apart; while in Experiment 2, water palatability was assessed using the taste reactivity test (TRT), on two separate test days 72 h apart. Both LPS and CCK significantly (p<0.05) reduced water intake, with the effects of combined LPS with CCK being more pronounced than either agent injected alone. Rats developed a rapid tolerance to the effects of LPS on water intake on subsequent exposures to LPS. Results from the TRT indicated that LPS enhanced water palatability (p<0.05), as evidenced by a high level of ingestive responding, whereas CCK produced a pattern of responding indicative of satiety. LPS plus CCK reduced ingestive responding on the first test day, but these responses were significantly increased on the second test day (p<0.05). These results demonstrate that although LPS reduces water intake, it enhances water palatability. The results further underscore the necessity for examining palatability changes in addition to intake measures when studying the regulation of feeding and drinking.     

Intracerebroventricular administration of mouse leptin does not reduce food intake in the chicken

Recently, it has been suggested that leptin plays an important role in regulation of food intake and metabolism in rats and mice, however, the effect of central administration of leptin on food intake in chicks has not been reported. We have investigated the anorexigenic effect of leptin administered by intracerebroventricular (i.c.v.) injection in chicks using mouse leptin, which shows 97% homology to chicken leptin. Three experiments were conducted. After being deprived of food for 3 h, male broiler chicks were administered leptin by i.c.v. injection at dose levels of 0, 0.2, 1.0 and 5.0 μg (Experiment 1) or 0, 2.5 and 5.0 μg (Experiment 2). The birds were allowed free access to the diet for 2 h (Experiment 1) and 24 h (Experiment 2) after treatment. Male Single Comb White Leghorn chicks were used in Experiment 3 and were treated in the same manner as in Experiment 1. In all experiments, central administration of mouse leptin did not influence food intake in the time periods examined. It appears that either mouse leptin does not bind to the chicken leptin receptor or in the chicken brain the leptin receptor may be absent.     

Effects of baclofen on feeding behaviour examined in the runway

The motivational mechanisms underlying the effects of systemic administration of the GABA-B agonist baclofen on feeding were examined using a runway. Food-deprived male hooded Lister rats were trained to traverse a runway for food reinforcement. Baclofen (1 mg/kg i.p.) significantly increased food intake and this was most evident on the final two blocks of testing. The 2 mg/kg dose of baclofen increased running speed without significantly altering intake. At the highest dose tested (4 mg/kg), no significant effects on either consummatory or appetitive measures were observed. These data suggest that low doses of baclofen enhance the consummatory phase of ingestion by attenuating the natural signals associated with onset of satiation. The data also suggest that baclofen has complex effects on appetitive behaviour that may interfere with its effects on consumption.     

Attenuation of drinking sweetened water following calcium channel blockade.

Recent reports cite results that both cocaine-induced conditioned place preference and activity stimulation are attenuated by pretreatment with the calcium channel blocker isradipine (ISR) in rats. By blocking voltage-dependent L-type calcium channels, ISR may regulate neural dopamine release that, in turn, decreases the putative rewarding effects mediated by dopaminergic mechanisms. It is known that nonfluid deprived rats avidly consume sweetened fluids; this suggests that the sweet taste is rewarding. Three experiments were conducted to determine the effects of ISR on drinking sweetened and nonflavored water. Experiment 1 was designed to test whether ISR would attenuate the intake of a palatable solution in a dose-dependent manner. To this end, ISR was administered both peripherally (3.0-30 mg/kg) and centrally (0.3-30 micrograms/rat) prior to a solution of saccharin and d-glucose (S + G) being made available to rats (15 min/day) and intake was recorded. ISR produced dose-dependent decreases (38%-81%) in S + G intake dependent on the route of administration. In Experiment 2, water intake was measured in 18 h water-deprived rats following ISR (10 mg/kg) administration as well as comparing S + G drinking. The effect of two ISR vehicles, dimethyl sulfoxide and Tween 80, upon fluid intake was also determined. ISR injection did not attenuate water intake in 18 h water-deprived rats and the choice of vehicle did not affect the ISR-induced attenuation of S + G drinking. In Experiment 3, a single dose (30 micrograms) of ICV administered ISR, that attenuated S + G intake by approximately 44%, did not attenuate water intake in 18 h water-deprived rats.(ABSTRACT TRUNCATED AT 250 WORDS)     

Reversal of fenfluramine and fluoxetine anorexia by 8-OH-DPAT is attenuated following raphe injection of 5,7-dihydroxytryptamine

Drugs that enhance serotonergic neurotransmission reduce food intake by directly or indirectly activating serotonergic receptors. In contrast drugs that inhibit serotonergic neurotransmission such as the 5-HT_1A agonist 8-hydroxy-2-(di-n-propyl-amino)tetralin (8-OH-DPAT) stimulate food intake. The present study examined the effects of 8-OH-DPAT on the feeding suppressant action of the indirect 5-HT agonists fenfluramine (FEN; 0.63–2.5 mg/kg) and fluoxetine (FLU; 2.5–10 mg/kg), as well as the 5-HT_1B/2C agonist 1-(3-trifluoromethylphenyl)piperazine (TFMPP; 0.5–2 mg/kg). 8-OH-DPAT (62.5–250 μg/kg) was administered 5 min prior to FEN, FLU or TFMPP, injected 30 min before food access. While FEN, FLU and TFMPP dose-dependently reduced 2 h food intake, 8-OH-DPAT stimulated eating behavior. 8-OH-DPAT (62.5–250 μg/kg) pretreatment reversed the anorectic action of FEN (1.25 mg/kg) and FLU (5 mg/kg) but not TFMPP (1 mg/kg). Separate groups of rats were injected with 5,7-dihydroxytryptamine (5,7-DHT; 3 μg free base) into both the dorsal and median raphe, which resulted in extensive 5-HT depletion in hypothalamus (80%), striatum and hippocampus (90%). In both 5,7-DHT and vehicle-injected rats, FEN (1.25 mg/kg) and FLU (5 mg/kg) suppressed feeding. In 5,7-DHT treated rats, however, the ability of 8-OH-DPAT (125 μg/kg) to block FEN and FLU induced anorexia was attenuated. That is, 8-OH-DPAT pretreatment did not reverse the feeding inhibitory effects of either FEN or FLU. Further, the ability of FEN and FLU to suppress food intake was not altered by the 5,7-DHT lesion. These findings suggest that the reversal of FEN and FLU anorexia by 8-OH-DPAT is partially dependent on the integrity of brain 5-HT systems since their disruption compromises the ability of this 5-HT_1A agonist to antagonize the feeding suppressant action of either FEN or FLU. However, the ability of treatments which impair 5-HT neurotransmission to reverse FEN and FLU induced suppression of food intake may depend upon whether this impairment is acute and reversible (8-OH-DPAT), or chronic and irreversible (5,7-DHT).     

Intraventricular GLP-1 reduces short- but not long-term food intake or body weight in lean and obese rats

Glucagon-like-peptide-1 (7–36) amide (GLP-1), when infused into the third ventricle (IVT), reduces short-term food intake. In the present experiments, we assessed whether IVT administration of GLP-1 could influence long-term food intake and body weight of lean Long Evans rats and of fatty Zucker (fa/fa) rats. In Experiment 1, we replicated the observation that 10 μg GLP-1, given IVT, reduces one and 2 h food intake, and extended the observation to fatty Zucker rats. However, in both rat strains, 24 h food intake and body weight were unchanged by this acute treatment. In Experiment 2, GLP-1 (30 μg/day) was infused IVT continuously for 4 days via an osmotic mini-pump. This treatment also had no effect on food intake or body weight in either Long–Evans or fatty Zucker rats. A control experiment verified that the GLP-1 remained biologically active over the duration of the infusion period. In a final experiment, Long-Evans rats were restricted to two 2 h periods of access to food each day for 6 days. Prior to each of these access periods, rats received either 15 μg of GLP-1 IVT or a vehicle control injection. While GLP-1 significantly reduced food intake on the first day of treatment, this effect of GLP-1 rapidly disappeared such that it was reduced on the second day and absent on the third day; and there was no effect on body weight at any time. Collectively, the present experiments do not support the hypothesis that GLP-1, acting in the CNS, is an important regulator of long-term food intake and body weight.     

Alteration of CCK-induced satiety in post-Nippostrongylus brasiliensis-infected rats

In rats, the nematode Nippostrongylus brasiliensis induces an intestinal inflammation, but after the inflammation had resolved and the worm burden eliminated, morphological alterations of the intestinal wall, mainly consisting of mast cell hyperplasia and enteric nerve remodeling, persist for several weeks. Intestinal signals reaching the brain through the vagus nerve and involving neuropeptides such as CCK, play a role in the control of food intake. Our hypothesis was that neuroimmune alterations of the intestine may alter this control. This work was aimed to evaluate whether post-infection alterations of the intestinal wall may affect the satiety effects of CCK and further, the role of mast cells and their mediators, histamine and serotonin, in post-N. brasiliensis-infected rats. In basal conditions, food intake was not different in control and post-infected groups of rats. Post-infected rats were characterized by prolonged satiety effects of both CCK and histamine but not serotonin. The prolonged effect of CCK was reduced when mast cells were previously stabilized by ketotifen, which had no effect per se on food intake. No difference was observed in the increase of food intake induced by CCK-A and CCK-B receptor antagonists in both control and post-infected rats. Mast cell degranulation with compound 48/80 induced severe anorectic effects that lasted for less than 24h in post-infected rats and as long as 6 days in controls. Thus, in our experimental conditions, i.e., within 30-50 days post-N. brasiliensis infection, we observed an enhancement of the anorectic effect of exogenous CCK involving mast cell degranulation and histamine.     

Effects of nocturnal intraperitoneal administration of cholecystokinin in rats: simultaneous increase in sleep, increase in EEG slow-wave activity, reduction of motor activity, suppression of eating, and decrease in brain temperature

Rats received an i.p. injection of cholecystokinin-octapeptide sulfate ester (CCK; 4, 10 or 50 micrograms/kg) or physiological saline at dark onset, and the 24-h sleep-wake cycle (12-h-dark and 12-h-light phases), spontaneous motor activity and brain temperature (Tbr) were recorded. EEG activity was studied through spectral analysis for 2.5 h, and food intake was measured at the end of postinjection hour 1. In response to CCK, non-REM sleep increased at the expense of wakefulness, and the sleep-promoting effect was substantiated by an increase in EEG slow-wave activity. Motor activity, Tbr and food intake decreased. The effects vanished in postinjection hour 2; the diurnal rhythms were not modified. The changes varied as a function of the dose: the effects were significant following 10 micrograms/kg, and even higher in response to 50 micrograms/kg CCK. The results indicate that i.p. CCK definitely promotes non-REM sleep. This effect may belong to the behavioral sequence elicited by the peptide, which is often attributed to satiety. As evidenced by the reduction of Tbr, CCK also exerts strong autonomic actions, which might interfere with the behavioral responses.     

Somatodendritic Release of Cholecystokinin Potentiates GABAergic Synapses Onto Ventral Tegmental Area Dopamine Cells

BackgroundNeuropeptides are contained in nearly every neuron in the central nervous system and can be released not only from nerve terminals but also from somatodendritic sites. Cholecystokinin (CCK), among the most abundant neuropeptides in the brain, is expressed in the majority of midbrain dopamine neurons. Despite this high expression, CCK function within the ventral tegmental area (VTA) is not well understood.MethodsWe confirmed CCK expression in VTA dopamine neurons through immunohistochemistry and in situ hybridization and detected optogenetically induced CCK release using an enzyme-linked immunosorbent assay. To investigate whether CCK modulates VTA circuit activity, we used whole-cell patch clamp recordings in mouse brain slices. We infused CCK locally in vivo and tested food intake and locomotion in fasted mice. We also used in vivo fiber photometry to measure Ca²⁺ transients in dopamine neurons during feeding.ResultsHere we report that VTA dopamine neurons release CCK from somatodendritic regions, where it triggers long-term potentiation of GABAergic (gamma-aminobutyric acidergic) synapses. The somatodendritic release occurs during trains of optogenetic stimuli or prolonged but modest depolarization and is dependent on synaptotagmin-7 and T-type Ca²⁺ channels. Depolarization-induced long-term potentiation is blocked by a CCK₂ receptor antagonist and mimicked by exogenous CCK. Local infusion of CCK in vivo inhibits food consumption and decreases distance traveled in an open field test. Furthermore, intra-VTA–infused CCK reduced dopamine cell Ca²⁺ signals during food consumption after an overnight fast and was correlated with reduced food intake.ConclusionsOur experiments introduce somatodendritic neuropeptide release as a previously unknown feedback regulator of VTA dopamine cell excitability and dopamine-related behaviors.     

Characteristics of feeding induced by the serotonin agonist 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT)

The effects of the putative serotonin agonist 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) on food intake in freely-feeding and food deprived rats were examined. In freely-feeding rats, low doses of 8-OH-DPAT (15-60 micrograms/kg) significantly increased food intake without affecting drinking, grooming, rearing or locomotion. Higher drug doses (125-4000 micrograms/kg) also produced feeding and caused locomotor stimulation and serotonin-related stereotyped behaviour (i.e., forepaw padding, headweaving, wet dog shakes, flat body posture). When feeding and stereotypy were observed concurrently, response competition was evident and feeding behaviour was fragmented into numerous short eating bouts. As drug-induced stereotypy declined with time, this fragmented pattern of eating was succeeded by long bouts of eating which were similar to those observed at doses of 15-60 micrograms/kg 8-OH-DPAT. In 24 hr food deprived rats, low doses of 8-OH-DPAT had no effect on food intake. However, high doses of 8-OH-DPAT (250-4000 micrograms/kg) decreased feeding in food deprived animals, an effect which was probably secondary to the induction of stereotypy. It is proposed that the behavioural effects of 8-OH-DPAT may be explained by a dual effect on brain serotonergic mechanisms, which is dose dependent. Thus, low doses of the drug may preferentially activate inhibitory presynaptic serotonin receptors (autoreceptors), decrease serotonin metabolism and thereby increase feeding. In contrast, high doses of 8-OH-DPAT appear to stimulate postsynaptic serotonin receptors and thus produce stereotypy. Alternatively, it is possible that 8-OH-DPAT may elicit feeding by postsynaptic serotonin receptor blockade.     

A cholecystokinin agonist/antagonist according to dose and time of action: Effect on food intake

A new CCK pseudopeptide (Boc-Tyr (SO3)-Nle-psi-(COCH2) Gly-Trp-Nle-Asp-Phe-NH2) has been described in a previous study as a potent CCK agonist in the peripheral system and as an antagonist in the central nervous system. When administered alone by a peripheral route, this pseudopeptide was found to decrease food intake in free/feeding rats, and thus behaved as a CCK agonist. However, it reversed the decreased feeding effect induced by a full potent agonist (Boc(Nle 28-Nle 31)-CCK26-33). This antagonistic action occurred 3-4 h after treatment, at a time when the agonistic properties of the pseudopeptide alone had disappeared, but when the effect of the full potent agonist remained. The dose-response curve of the antagonistic action was bell-shaped. These results suggest that this new pseudopeptide is not only agonistic in the periphery and antagonistic in the central nervous system as shown in the previous study, but is also both agonistic and antagonistic on the same paradigm according to the dose and time of action.     

Role of leptin in the control of feeding of goldfish Carassius auratus: interactions with cholecystokinin,neuropeptide Y and orexin A,and modulation by fasting

To assess the role of leptin on food intake regulation in goldfish, we examined the effects of central (intracerebroventricular, ICV) and peripheral (intraperitoneal, IP) injections of recombinant murine leptin on feeding behavior. Centrally (100 ng/g) and peripherally (300 ng/g) injected leptin both caused a significant decrease in food intake, compared to the saline-treated controls. To test the hypothesis that leptin influenced orexigenic neuropeptide systems in goldfish, fish were co-injected with neuropeptide Y (NPY) or orexin A and leptin. Both NPY (5 ng/g) and orexin A (10 ng/g) significantly increased food intake. Fish co-injected ICV with NPY (5 ng/g) or orexin A (10 ng/g) and leptin (1 or 10 ng/g) had a food intake lower than that of fish treated with NPY or orexin A alone. NPY mRNA expression in goldfish brain was reduced 2 and 6 h following central injection of leptin. To test the hypothesis that the cholecystokinin (CCK) mediates the effects of leptin in goldfish, fish were simultaneously injected ICV with an ineffective dose of leptin (10 ng/g) and either ICV or IP with an ineffective doses of CCK (1 ng/g ICV or 25 ng/g IP). These fish had a food intake lower than vehicle-treated fish, suggesting that leptin potentiates the satiety actions of CCK. CCK hypothalamic mRNA expression was increased 2 h following central treatment with leptin. The CCK receptor antagonist proglumide blocked both central and peripheral CCK satiety effects. Blockade of CCK brain receptors by proglumide resulted in an inhibition of the leptin-induced decrease in food intake and an attenuation of the inhibiting action of leptin on both NPY- and orexin A-induced feeding. These data suggests that CCK has a role in mediating the effects of leptin on food intake. Fasting potentiated the actions of leptin and attenuated the effects of CCK. Whereas fasting had no effects on the brain mRNA expression of CCK, it increased the brain mRNA expression of NPY and decreased the expression of CART. These changes in neuropeptide expression were partially reversed when fish were treated ICV with leptin. These results provide strong evidence that, in goldfish, leptin influences food intake, in part by modulating the orexigenic effects of NPY and orexin and that its actions are mediated, at least in part, by CCK.