Comparison of central administration of corticotropin-releasing hormone and urocortin on food intake, conditioned taste aversion, and c-Fos expression

Corticotropin-releasing hormone (CRH) is a potent regulator of the hypothalamic-pituitary-adrenal axis, and reduces food intake when administered into the third cerebral ventricle (i3vt). However, CRH also promotes conditioned taste aversion (CTA) learning which indicates that its anorectic effects are accompanied by aversive consequences that would reduce food intake independently of energy regulation. Urocortin (Ucn) is a closely related mammalian peptide that binds to both identified CRH receptor subtypes and also reduces food intake when administered i3vt. The present experiments compared the aversive consequences of i3vt administration of CRH and Ucn at doses that produced comparable decrements in food intake. Experiment 1 found that 1.0 microg Ucn and 2.0 microg CRH produced similar reductions in food intake. Experiment 2 demonstrated that, at these doses, CRH but not Ucn promoted robust and reliable CTA learning. A third experiment showed comparable increased c-Fos-like immunoreactivity after Ucn and CRH in forebrain and hindbrain structures associated with food intake. It is concluded that Ucn, at doses that reduce food intake to levels like that observed after administration of CRH, do not produce similarly aversive consequences.     

Dose-dependent effects of peptide YY(3-36) on conditioned taste aversion in rats

We used a conditioned taste aversion test to assess whether PYY(3-36) reduces food intake by producing malaise. Two-hour IV infusion of PYY(3-36) (8, 15, and 30 pmol/kg/min) at dark onset in non-food-deprived rats produced a dose-dependent inhibition of feeding and a conditioned aversion to the flavored chow paired with PYY(3-36) infusion. In food-deprived rats, PYY(3-36) at 2 and 4 pmol/kg/min inhibited intake of a flavored saccharin solution without producing conditioned taste aversion, whereas higher doses (8 and 15 pmol/kg/min) inhibited saccharin intake and produced taste aversion. These results suggest that anorexic doses of PYY(3-36) may produce a dose-dependent malaise in rats, which is similar to that reported for PYY(3-36) infusion in humans. Previous studies have shown that PYY(3-36) potently inhibits gastric emptying, and that gut distention can produce a conditioned taste aversion. Thus, PYY(3-36) may produce conditioned taste aversion in part by slowing gastric emptying.     

Peripheral activation of corticotropin-releasing factor receptor 2 inhibits food intake and alters meal structures in mice

The orexigenic effect of urocortins (Ucns), namely Ucn 1, Ucn 2 and Ucn 3 through activation of corticotropin-releasing factor (CRF) receptors, has been well characterized after injection into the brain but not in the periphery. We examined the role of CRF receptor subtype 2 (CRF₂) in the regulation of food intake using intraperitoneal (ip) injection of Ucns and the selective CRF₂ antagonist, astressin₂-B, and CRF₂ knockout (−/−) mice. Meal structures were monitored using an automated episodic solid food intake monitoring system. Ucn 2 (3, 10 or 30 μg/kg, ip) induced a rapid in onset, long lasting and dose-dependent decrease (38%, 66% and 86%, respectively at 4 h) of cumulative food intake after an overnight fast in mice. Ucn 3 anorexic effect was 10-times less potent. Astressin₂-B (30 or 100 μg/kg) injected ip, but not intracerebroventricularly, blocked the inhibitory effect of ip Ucn 1 and Ucn 2 (10 μg/kg). Fasted CRF_2−/− mice did not respond to ip Ucn 1 (10 μg/kg). Meal microstructure analysis of the 4-h re-feeding response to an overnight fast showed that Ucn 2 (10 μg/kg, ip) decreased meal size and duration, but increased meal frequency. In mice fed ad libitum, Ucn 2 (30 μg/kg) injected ip before the dark phase decreased the 4-h nocturnal meal size and duration without influencing meal frequency while the 10 μg/kg dose had no effect. These data indicate that Ucns, through peripheral CRF₂ receptor-mediated induction of satiation, inhibit the eating response to a fast more potently than the physiological nocturnal feeding in mice.     

Tamoxifen produces conditioned taste avoidance in male rats: An analysis of microstructural licking patterns and taste reactivity

Estrogen receptor activation has been shown to reduce body weight and produce a conditioned reduction in food intake in male rats that is putatively mediated by estradiol's suggested aversive effects. Evidence has shown that the selective estrogen receptor modulator tamoxifen used in the prevention and treatment of breast cancer may also produce changes in food intake and body weight, which are known to impact cancer development and survival. The purpose of the present study was to examine whether tamoxifen produces a conditioned reduction in intake similar to estradiol by producing a conditioned aversion. A one bottle lickometer test was used to examine conditioned changes in sucrose drinking, while the taste reactivity test was used to measure rejection reactions, which serve to index aversion in rats. A backward conditioning procedure that consisted of 3 conditioning days and one vehicle test day was used to examine conditioned changes in 0.3 M sucrose intake and taste reactivity. Our results show that tamoxifen produced a conditioned reduction in sucrose drinking in a one bottle fluid intake test that was similar to the effects produced by estradiol (positive control); however, no active rejection reactions were produced by either tamoxifen (1 and 10 mg/kg) or estradiol. The present results suggest that tamoxifen, at the doses used in the present study, acts as an estrogen receptor agonist to regulate food intake and that the conditioned reduction in intake produced by tamoxifen and estradiol reflects conditioned taste avoidance rather than conditioned taste aversion.     

Mechanisms of oleoylethanolamide-induced changes in feeding behavior and motor activity

Oleoylethanolamide (OEA), a lipid synthesized in the intestine, reduces food intake and stimulates lipolysis through peroxisome proliferator-activated receptor-alpha. OEA also activates transient receptor potential vanilloid type 1 (TRPV1) in vitro. Because the anorexigenic effect of OEA is associated with delayed feeding onset and reduced locomotion, we examined whether intraperitoneal administration of OEA results in nonspecific behavioral effects that contribute to the anorexia in rats. Moreover, we determined whether circulating levels of other gut hormones are modulated by OEA and whether CCK is involved in OEA-induced anorexia. Our results indicate that OEA reduces food intake without causing a conditioned taste aversion or reducing sodium appetite. It also failed to induce a conditioned place aversion. However, OEA induced changes in posture and reduced spontaneous activity in the open field. This likely underlies the reduced heat expenditure and sodium consumption observed after OEA injection, which disappeared within 1 h. The effects of OEA on motor activity were similar to those of the TRPV1 agonist capsaicin and were also observed with the peroxisome proliferator-activated receptor-alpha agonist Wy-14643. Plasma levels of ghrelin, peptide YY, glucagon-like peptide 1, and apolipoprotein A-IV were not changed by OEA. Finally, antagonism of CCK-1 receptors did not affect OEA-induced anorexia. These results suggest that OEA suppresses feeding without causing visceral illness and that neither ghrelin, peptide YY, glucagon-like peptide 1, apolipoprotein A-IV, nor CCK plays a critical role in this effect. Despite that OEA-induced anorexia is unlikely to be due to impaired motor activity, our data raise a cautionary note in how specific behavioral and metabolic effects of OEA should be interpreted.     

Urocortin in the lateral septal area modulates feeding induced by orexin A in the lateral hypothalamus

The intermediate portion of the lateral septum (LSi) contains high levels of urocortin (UCN) peptide and type 2 corticotropin-releasing hormone (CRH) receptor (CRHR2) and has anatomic and functional connections with the lateral hypothalamus (LH). We tested the effect of UCN in the LSi on feeding. Injection of 10 or 30 pmol UCN into LSi significantly decreased feeding in food-deprived rats for 24 h without producing conditioned taste aversion (CTA). Pretreatment with a CRH receptor antagonist, alpha-helical CRH (alpha-hCRH), blocked the inhibitory effect of UCN on deprivation-induced feeding at 1 and 2 h postinjection. Furthermore, UCN in the LSi significantly decreased feeding induced by LH-injected orexin A at 2 and 4 h postinjection, and addition of alpha-hCRH blocked the inhibitory effect of UCN on orexin A-induced feeding. In conclusion, UCN significantly inhibits feeding induced by deprivation and LH-injected orexin A without producing a CTA, an effect that is mediated by CRHR2. These data define the LSi as an important site for UCN-induced anorexia and indicate that LSi UCN may influence orexin A feeding signals in the LH.     

Urocortin 1 administered into the hypothalamic supraoptic nucleus affects open-field behaviour in rats

The presence of both Urocortin 1 (Ucn1) and corticotropin-releasing factor 2 receptors (CRF₂R) in the hypothalamic supraoptic nucleus (SON) suggests that endogenous Ucn1 released within this brain area acts as a local signal that might be involved in the regulation of not only endocrine but also behavioural stress responses. To test this hypothesis, we monitored the effects induced by the administration of a range of doses of synthetic Ucn1 (0.001–1.0 μg) bilaterally into the SON of rats in the open field test (OFT). Ucn1 administration produced an inverted U-shaped dose–response curve on OFT behaviour, in particular the dose of 0.01 μg of Ucn1 significantly increased the number of rearing and grooming episodes without affecting locomotion. In addition, this dosage augmented also the latency to visit the centre of the open field. Pre-treatment with the CRF₂R antagonist, astressin-2B (0.1 μg) normalized Ucn1 treatment-induced effects. These results suggest that Ucn1 released within the SON area interacts with CRF₂R to control the state of arousal.     

Brain-derived neurotrophic factor in the ventromedial nucleus of the hypothalamus reduces energy intake

Recent studies show that brain-derived neurotrophic factor (BDNF) decreases feeding and body weight after peripheral and ventricular administration. BDNF mRNA and protein, and its receptor TrkB, are widely distributed in the hypothalamus and other brain regions. However, there are few reports on specific brain sites of actions for BDNF. We evaluated the effect of BDNF, given into the ventromedial nucleus of the hypothalamus (VMH), on normal and deprivation- and neuropeptide Y (NPY)-induced feeding behavior and body weight. BDNF injected unilaterally or bilaterally into the VMH of food-deprived and nondeprived rats significantly decreased feeding and body weight gain within the 0- to 24-h and the 24- to 48-h postinjection intervals. Doses effectively producing inhibition of feeding behavior did not establish a conditioned taste aversion. BDNF-induced feeding inhibition was attenuated by pretreatment of the TrkB-Fc fusion protein that blocks binding between BDNF and its receptor TrkB. VMH-injected BDNF significantly decreased VMH NPY-induced feeding at 1, 2, and 4 h after injection. In summary, BDNF in the VMH significantly decreases food intake and body weight gain, by TrkB receptor-mediated actions. Furthermore, the anorectic effects of BDNF in this site appear to be mediated by NPY. These data suggest that the VMH is an important site of action for BDNF in its effects on energy metabolism.     

Comparison of the anorexigenic activity of CRF family peptides

Corticotropin releasing factor (CRF) family peptides have an important role in the control of food intake. We investigated the effects of CRF family peptides on food intake and body weight gain in mice. Of the CRF family peptides, including CRF, urocortin1 (Ucn1), urocortin2 (Ucn2) and urocortin3 (Ucn3), peripherally administered Ucn1 was shown to have the most potent inhibitory effect on the food intake and body weight gain of both lean and high fat fed obese mice. In addition, repeated administration of Ucn1 lowered blood glucose and acylated ghrelin, and decreased the visceral fat weight of high fat fed obese mice.     

Cholecystokinin produces conditioned place-aversions,not place-preferences,in food-deprived rats: Evidence against involvement in satiety

The net reinforcing/aversive properties of cholecystokinin (CCK) were measured using a conditioned place-preference paradigm. Both sated and food-deprived rats showed a dose dependent aversion to an environment previously paired with CCK that was correlated with the effects of CCK on feeding. In contrast, all rats showed a conditioned preference for an environment previously paired with food. These results demonstrate the aversive properties of CCK and suggest that the decrease in feeding that follows peripheral administration of CCK results from a CCK-induced malaise.     

Paraventricular hypothalamic alpha-melanocyte-stimulating hormone and MTII reduce feeding without causing aversive effects

alpha-Melanocyte-stimulating hormone (alpha-MSH) appears to play a tonic inhibitory role in feeding and energy storage. MTII, a specific synthetic MC3-R/MC4-R agonist, has similar effects on feeding in rats. The current studies demonstrate that PVN administration of alpha-MSH or MTII decreases nocturnal and NPY-stimulated food intake without causing aversive effects. Co-administration with NPY of 600 pmol alpha-MSH or 1 pmol MTII into the PVN caused a significant decrease in NPY-induced feeding. PVN administration of MTII or alpha-MSH at doses effective to suppress feeding did not cause conditioned taste aversion (CTA). ICV administration of alpha-MSH, however, did cause weak CTA. These results indicate that the potent effects on feeding of MC3-R and MC4-R agonists when injected into the PVN are not due to aversive effects.     

Hypothalamic neuronal histamine signaling in the estrogen deficiency-induced obesity

Menopause is one of the triggers that induce obesity. Estradiol (E2), corticotropin-releasing hormone (CRH), and hypothalamic neuronal histamine are anorexigenic substances within the hypothalamus. This study examined the interactions among E2, CRH, and histamine during the regulation of feeding behavior and obesity in rodents. Food intake was measured in rats after the treatment of E2, α-fluoromethyl histidine, a specific suicide inhibitor of histidine decarboxylase that depletes hypothalamic neuronal histamine, or CRH antagonist. We measured food intake and body weight in wild-type mice or mice with targeted disruption of the histamine receptors (H1-R) knockout (H1KO mice). Furthermore, we investigated CRH content and histamine turnover in the hypothalamus after the E2 treatment or ovariectomy (OVX). We used immunohistochemical staining for estrogen receptors (ERs) in the histamine neurons. The E2-induced suppression of feeding was partially attenuated in rats pre-treated with α-fluoromethyl histidine or CRH antagonist and in H1KO mice. E2 treatment increased CRH content and histamine turnover in the hypothalamus. OVX increased food intake and body weight, and decreased CRH content and histamine turnover in the hypothalamus. In addition, E2 replacement reversed the OVX-induced changes in food intake and body weight in wild-type mice but not in H1KO mice. Immunohistochemical analysis revealed ERs were expressed on histamine neurons and western blotting analysis and pre-absorption study confirmed the specificity of ER antiserum we used. These results indicate that CRH and hypothalamic neuronal histamine mediate the suppressive effects of E2 on feeding behavior and body weight.     

Tests of adipsia and conditioned taste aversion following the intrahypothalamic injection of amylin.

Intrahypothalamic injection of amylin (AMY) was shown to reduce the intake of rat chow and water for 8 and 4 h, respectively, in schedule-fed rats. Amylin also reduced water intake to a much lesser degree in 24-h water-deprived rats. A test of the ability of AMY to form a conditioned taste aversion yielded no change in saccharin preference, as compared to controls treated with vehicle. These results suggest that although AMY has adipsic effects, the reduction in water is not of sufficient magnitude to cause the anorexia. In addition, the failure of AMY to support a conditioned taste aversion suggests that AMY does not cause anorexia by inducing malaise. Therefore, in addition to other metabolic effects, AMY may be involved in the control of food and water intake.     

Pharmacological stimulation of brain carnitine palmitoyl-transferase-1 decreases food intake and body weight

Inhibition of brain carnitine palmitoyl-transferase-1 (CPT-1) is reported to decrease food intake and body weight in rats. Yet, the fatty acid synthase (FAS) inhibitor and CPT-1 stimulator C75 produces hypophagia and weight loss when given to rodents intracerebroventricularly (icv). Thus roles and relative contributions of altered brain CPT-1 activity and fatty acid oxidation in these phenomena remain unclarified. We administered compounds that target FAS or CPT-1 to mice by single icv bolus and examined acute and prolonged effects on feeding and body weight. C75 decreased food intake rapidly and potently at all doses (1-56 nmol) and dose dependently inhibited intake on day 1. Dose-dependent weight loss on day 1 persisted through 4 days of postinjection monitoring. The FAS inhibitor cerulenin produced dose-dependent (560 nmol) hypophagia for 1 day, weight loss for 2 days, and weight regain to vehicle control by day 3. The CPT-1 inhibitor etomoxir (32, 320 nmol) did not alter overall day 1 feeding. However, etomoxir attenuated the hypophagia produced by C75, indicating that CPT-1 stimulation is important for C75's effect. A novel compound, C89b, was characterized in vitro as a selective stimulator of CPT-1 that does not affect fatty acid synthesis. C89b (100, 320 nmol) decreased feeding in mice for 3 days and produced persistent weight loss for 6 days without producing conditioned taste aversion. Similarly, intraperitoneal administration decreased feeding and body weight without producing conditioned taste aversion. These results suggest a role for brain CPT-1 in the regulation of energy balance and implicate CPT-1 stimulation as a pharmacological approach to weight loss.     

The role of putative 5-HT1A and 5-HT1B receptors in the control of feeding in rats

8-hydroxy-2(di-n-propylamino)tetraline (8-OH-DPAT) and 5-methoxy-3(1,2,3,6-tetrahydro-4-pyridinyl)1H indole succinate (RU 24969), two agonists on the putative serotonin 1A and serotonin 1B receptors, were used for exploring the role of these sites in the inhibitory effect of serotonin (5-HT) on feeding. In free-feeding rats, 2.5-5 mg/kg RU 24969 significantly reduced food intake while doses of 8-OH-DPAT ranging from 0.125 to 0.5 mg/kg increased eating. The effects of the highest doses were associated with hyperlocomotion and hyperreactivity for RU 24969 and a typical motor syndrome (flat body posture and forepaw treading) for 8-OH-DPAT. The motor syndrome caused by 0.5 mg/kg 8-OH-DPAT was much more obvious in food-deprived rats in which food intake was also markedly reduced. RU 24969 1.25 and 5 mg/kg reduced food intake by food-deprived rats and caused hyperlocomotion not different from that in free-feeding animals. Pretreatment with metergoline (2 mg/kg i.p.) prevented the effect of 5 mg/kg RU 24969 on food intake by food-deprived rats but had no effect on the reduction of eating caused by 0.5 mg/kg 8-OH-DPAT. The motor syndrome caused by 8-OH-DPAT was not changed by metergoline but the hyperlocomotion caused by RU 24969 was potentiated. Haloperidol (0.1 mg/kg i.p.) completely blocked the hyperlocomotion but did not change the reduction of food intake caused by RU 24969 in food-deprived rats. It is suggested that the putative serotonin 1B receptors specifically mediate the inhibitory effect of 5-HT on feeding whereas serotonin 1A sites act by enhancing eating only in free-feeding animals.     

Intravenous injection of urocortin 1 induces a CRF2 mediated increase in circulating ghrelin and glucose levels through distinct mechanisms in rats

Urocortins (Ucns) injected peripherally decrease food intake and gastric emptying through peripheral CRF₂ receptors in rodents. However, whether Ucns influence circulating levels of the orexigenic and prokinetic hormone, ghrelin has been little investigated. We examined plasma levels of ghrelin and blood glucose after intravenous (iv) injection of Ucn 1, the CRF receptor subtype involved and underlying mechanisms in ad libitum fed rats equipped with a chronic iv cannula. Ucn 1 (10 μg/kg, iv) induced a rapid onset and long lasting increase in ghrelin levels reaching 68% and 219% at 0.5 and 3 h post injection respectively and a 5-h hyperglycemic response. The selective CRF₂ agonist, Ucn 2 (3 μg/kg, iv) increased fasting acyl (3 h: 49%) and des-acyl ghrelin levels (3 h: 30%) compared to vehicle while the preferential CRF₁ agonist, CRF (3 μg/kg, iv) had no effect. <!-- no-mfc -->Ucn 1's<!-- /no-mfc --> stimulatory actions were blocked by the selective CRF₂ antagonist, astressin₂-B (100 μg/kg, iv). Hexamethonium (10 mg/kg, sc) prevented Ucn 1-induced rise in total ghrelin levels while not altering the hyperglycemic response. These data indicate that systemic injection of Ucns induces a CRF₂-mediated increase in circulating ghrelin levels likely via indirect actions on gastric ghrelin cells that involves a nicotinic pathway independently from the hyperglycemic response.     

Neuropeptide K suppresses feeding in the rat

We studied the effects of neuropeptide K (NPK), a 36 amino acid residue peptide of the tachykinin family, on latency to onset of feeding and cumulative 1 and 2 h food intake in three experimental paradigms. Intraperitoneal injection of NPK (1.25 and 3.14 nmol) to food-deprived rats delayed the onset of feeding and significantly decreased the cumulative food intake. Intraperitoneal injection of NPK (1.25 and 3.14 nmol) to water-deprived rats produced no effect on subsequent drinking behavior. Similarly, intraperitoneal injection of NPK (3.14 nmol) 15 min before onset of the dark phase (of the light-dark cycle) significantly delayed the occurrence of ingestive behavior and the cumulative food intake was markedly suppressed. Furthermore, administration of NPK intraperitoneally (0.5-3.14 nmol) 15 min before intraventricular (i.c.v.) injection of neuropeptide Y (NPY 0.47 nmol) to satiated rats significantly suppressed NPY-induced feeding and delayed the onset of ingestive behavior. However, when administered centrally prior to NPY injection, NPK delayed the onset of feeding response only. Collectively, these findings show that NPK can acutely and consistently suppress feeding behavior.     

Involvement of opioid receptor subtypes in rat feeding behavior

The short-acting opiate antagonist naloxone decreases food intake in three models of ingestive behavior: free feeding, food-deprivation induced feeding and deoxyglucose-induced feeding. Twenty-four hours after administration, the long-acting, mu1 selective antagonist naloxonazine inhibits food intake to the same extent as naloxone in freely feeding and food-deprived rats, but not in animals treated with 2-deoxyglucose. These results indicate that 1) opiates modulate feeding through multiple opioid receptor mechanisms, one of which is the mu subtype, and 2) the feeding observed in various experimental paradigms are modulated by different receptor subtypes. Furthermore, these results illustrate the usefulness of naloxone in defining a behavior as opioid but point out its limitations in discriminating between opioid receptor subtypes.     

Urocortin has cell-proliferative effects on cardiac non-myocytes

Urocortin (Ucn) is a member of the corticotropin-releasing hormone (CRH)-related peptides that has been reported to have cardiac inotropic and hypertrophic effects. In addition, Ucn mRNA was expressed in cardiac myocytes (MCs) and Ucn was suggested to have cardioprotective effects. Recently, it was reported that Ucn mRNA was expressed in cardiac non-myocytes (NMCs). Based on these facts, Ucn is assumed to affect not only MCs but also NMCs in an autocrine fashion. The present study was designed to elucidate a pathophysiological role of Ucn on NMCs. NMCs were prepared by the discontinuous Percoll gradient and adhesion method. Ucn increased [(3)H]-thymidine uptake into NMCs. Ucn also enhanced endothelin-1-induced increase of [(3)H]-thymidine uptake into NMCs. Effects of Ucn on [(3)H]-thymidine uptake into NMCs were significantly abolished by the protein kinase A inhibitor, H89 (10(-5) M), but not by a competitive antagonist of CRH receptors, astressin (10(-5) M). Ucn also increased intracellular cAMP accumulation more potently than CRH on a molar basis. Finally, both MCs and NMCs also secreted Ucn. Together with the recent findings, at least in NMCs, these data suggest that Ucn could exert its own actions via the cAMP signaling pathway, but not through known CRH type 2 receptors, in an autocrine fashion. In conclusion, the present study indicated that Ucn was secreted not only from MCs but also from NMCs and that the primary source of Ucn acting on heart was the heart itself. On the other hand, Ucn could proliferate NMCs as well as MCs, suggesting that Ucn could be involved in cardiac hypertrophy and fibrosis, i.e., cardiac remodeling, in spite of its putative cardioprotective actions.