Elevated ghrelin alters the behavioral effects of perinatal acetaminophen exposure in rats.

A growing body of evidence links prenatal and early postnatal acetaminophen (APAP) exposure to atypical development of brain and behavior. In adult rodents, APAP is known to produce oxidative stress and lower anxiety-related behavior following acute exposure. In models of early-life exposure, APAP has also been shown to alter anxiety-related and other behaviors. Since the neuropeptide ghrelin has been recently shown to reduce oxidative stress markers and act as a neuroprotectant, we hypothesized that exposure to ghrelin prior to exposure to APAP would mitigate the behavioral effects of APAP exposure. On postnatal day 7, pups were administered doses of either APAP (51.97 mg/kg), ghrelin (1 mg/kg/ml), ghrelin + APAP, or vehicle only. As adults, anxiety-related behavior was assessed in the open field and elevated plus maze. Behavior differed based upon treatment condition. In rats unexposed to ghrelin, APAP treatment resulted in increased exploration (i.e., reduced anxiety) in the open field relative to controls. Rats co-administered APAP and ghrelin did not differ from vehicle-only controls. No significant effects of APAP or interactions between APAP and ghrelin exposures were observed in the elevated plus maze. These results are the first to demonstrate that ghrelin can mitigate the effects of perinatal APAP exposure in rats.

Alcohol Binge Drinking and Anxiety-Like Behavior in Socialized Versus Isolated C57BL/6J Mice.

BACKGROUND: Binge alcohol drinking has been characterized as a key feature of alcoholism. The drinking-in-the-dark (DID) preclinical model, a procedure that promotes high levels of ethanol (EtOH) intake in short periods of time, has been extensively used to investigate neuropharmacological and genetic determinants of binge-like EtOH consumption. Using DID methodology, alcohol-preferring strains of mice such as C57BL/6J (B6) mice consume enough EtOH to achieve blood concentrations (≥1.0 mg/ml) associated with behavioral intoxication (i.e., motor incoordination). DID procedures typically involve the use of socially isolated animals (single-housed prior to and during the experiment). Previous research indicates that stress associated with social isolation can induce anxiety-like behavior and promote increases in EtOH intake. The present study investigates the role of housing conditions in anxiety-like behavior and binge-like EtOH intake using a DID procedure. METHODS: Male and female B6 mice were isolated or pair-housed for a period of 6 weeks prior to evaluation of anxiety-like (elevated plus maze, light and dark box, open field) and drinking (water, 10% sucrose, 10 to 30% EtOH) behavior. In order to measure intake, a variation of the standard DID procedure using a removable, transparent, and perforated plastic barrier strip (designed to temporarily divide the cage in 2) was introduced. This allowed for individual intake records (2-hour test) of isolated and socially housed animals. RESULTS: Increased anxiety-like behavior and reduced sucrose consumption were found in isolated mice. The effects of housing conditions on EtOH intake were sex- and concentration-dependent. In male mice, isolation increased 20 and 30% EtOH intake. In females, however, an increased intake of EtOH (30%) was found in socialized animals. No effects of housing or sex were found at EtOH 10%. CONCLUSIONS: Together with previous literature, the present study suggests that social isolation can promote anxiety-associated behavior and produce sex-dependent changes in binge-like EtOH consumption.

Brain Site-Specific Inhibitory Effects of the GLP-1 Analogue Exendin-4 on Alcohol Intake and Operant Responding for Palatable Food.

Approximately 14.4 million Americans are experiencing alcohol use disorder (AUD) and about two-thirds of people who experience drug addiction will relapse, highlighting the need to develop novel and effective treatments. Glucagon-like peptide-1 (GLP-1) is a peptide hormone implicated in the mesocorticolimbic reward system and has become a peptide of interest with respect to its putative inhibitory effects on drug reward. In order to further develop treatments for those diagnosed with AUD, the interplay between GLP-1 receptor signaling and ethanol consumption must be elucidated. In the present study, we investigated the ability of the GLP-1 analogue, exendin-4 (Ex-4), to alter alcohol intake and operant responding for sucrose pellets in order to further understand the role of this compound in mediating reward. We selected multiple sites throughout the prosencephalic and mesencephalic regions of the brain, where we directly administered various doses of Ex-4 to male Sprague Dawley rats. In alcohol investigations, we utilized a two-bottle choice intermittent access protocol. In separate groups of rats, we adopted an operant paradigm in order to examine the effect of Ex-4 on motivated responding for palatable food. Results indicated that GLP-1 receptor signaling effectively suppressed voluntary alcohol intake when injected into the ventral tegmental area (VTA), the accumbens core (NAcC) and shell (NAcS), the dorsomedial hippocampus (DMHipp), and the lateral hypothalamus (LH), which are all structures linked to brain reward mechanisms. The arcuate nucleus (ARcN) and the paraventricular nucleus (PVN) of the hypothalamus were unresponsive, as was the basolateral amygdala (BLA). However, Ex-4 treatment into the ArcN and PVN suppressed operant responding for sucrose pellets. In fact, the VTA, NAcC, NAcS, LH, and the DMHipp all showed comparable suppression of sucrose responding. Overall, our findings suggest that these central structures are implicated in brain reward circuitry, including alcohol and appetitive motivation, which may be mediated by GLP-1 receptor mechanisms. GLP-1, therefore, may play a critical role in modifying addictive behaviors via activation of multiple GLP-1 systems throughout the brain.

Exendin-4 antagonizes the metabolic action of acylated ghrelinergic signaling in the hypothalamic paraventricular nucleus.

In the current study we investigated the interaction of hypothalamic paraventricular nucleus (PVN) glucagon-like peptide-1 (GLP-1) and ghrelin signaling in the control of metabolic function. We first demonstrated that acylated ghrelin injected directly into the PVN reliably altered the respiratory exchange ratio (RER) of adult male Sprague Dawley rats. All testing was carried out during the initial 2 h of the nocturnal cycle using an indirect open circuit calorimeter. Results indicated that acylated ghrelin induced a robust increase in RER representing a shift toward enhanced carbohydrate oxidation and reduced lipid utilization. In contrast, treatment with comparable dosing of des-acyl ghrelin failed to significantly impact metabolic activity. In separate groups of rats we subsequently investigated the ability of exendin-4 (Ex-4), a GLP-1 analogue, to alter acylated ghrelin's metabolic effects. Rodents were treated with either systemic or direct PVN Ex-4 followed by acyl ghrelin microinjection. While our results showed that both systemic and PVN administration of Ex-4 significantly reduced RER, importantly, Ex-4 pretreatment itself reliably inhibited the impact of ghrelin on RER. Overall, these findings provide increasingly compelling evidence that GLP-1 and ghrelin signaling interact in the neural control of metabolic function within the PVN.

Glucagon-Like Peptide-1 (GLP-1) and 5-Hydroxytryptamine 2c (5-HT2c) Receptor Agonists in the Ventral Tegmental Area (VTA) Inhibit Ghrelin-Stimulated Appetitive Reward.

Current literature indicates that the orexigenic peptide ghrelin increases appetitive motivation via signaling in the mesolimbic reward system. Another gastric peptide, glucagon-like peptide-1 (GLP-1), and the neurotransmitter 5-hydroxytryptamine (5-HT), are both known to suppress operant responding for food by acting on key mesolimbic nuclei, including the ventral tegmental area (VTA). In order to investigate the interaction effects of ghrelin, GLP-1, and 5-HT within the VTA, we measured operant responding for sucrose pellets after the administration of ghrelin, the GLP-1 receptor agonist exendin-4 (Ex-4), and the 5-HT2c receptor agonist Ro60-0175 in male Sprague-Dawley rats. Following training on a progressive ratio 3 (PR3) schedule, animals were first injected with ghrelin into the VTA at doses of 3 to 300 pmol. In subsequent testing, separate rats were administered intraperitoneal (IP) Ex-4 (0.1⁻1.0 µg/kg) or VTA Ex-4 (0.01⁻0.1 µg) paired with 300 pmol ghrelin. In a final group of rats, the 5-HT2c agonist Ro60-0175 was injected IP (0.25⁻1.0 mg/kg) or into the VTA (1.5⁻3.0 µg), and under both conditions paired with 300 pmol ghrelin delivered into the VTA. Our results indicated that ghrelin administration increased operant responding for food reward and that this effect was attenuated by IP and VTA Ex-4 pretreatment as well as pre-administration of IP or VTA Ro60-0175. These data provide compelling evidence that mesolimbic GLP-1 and serotonergic circuitry interact with the ghrelinergic system to suppress ghrelin's effects on the mediation of food reinforcement.

Central effects of insulin detemir on feeding, body weight, and metabolism in rats.

Insulin detemir (DET) is a basal insulin analog that, in contrast to other long-acting forms of insulin, has significant weight-gain-sparing effects in diabetic patients. We hypothesized that this effect of DET may be due to its enhanced catabolic action in the central nervous system. We investigated the long-term effects of single third ventricular (3V) microinjections of equimolar doses of DET and regular insulin in normal male rats on feeding, body weight, energy expenditure (EE), and respiratory quotient (RQ). Also, in acute testing, we assessed the ability of lower doses of DET to alter feeding, EE, and RQ when microinjected directly into the paraventricular nucleus (PVN). The anabolic peptide ghrelin served as a positive control in acute testing. 3V administration of both DET (0.5-2.0 mU) and regular insulin (2.0-8.0 mU) significantly reduced feeding and body weight over 48 and 120 h, respectively, with DET yielding greater inhibitory effects. DET also stimulated greater elevations of EE and reductions of RQ over 72 and 48 h postinjection, respectively. In acute (4 h) testing, microinjections of DET (0.5 mU) into the PVN reduced feeding, increased EE, and reduced RQ, while ghrelin (100 pmol) had the opposite effects. When administered sequentially into the PVN, DET (0.25 and 0.5 mU) reversed ghrelin-induced feeding, EE, and RQ effects. These data support the notion that the weight-sparing effect of DET is at least in part based on its central catabolic action and that enhanced EE and reduced RQ may participate in this effect.

Ghrelin enhances food intake and carbohydrate oxidation in a nitric oxide dependent manner.

In the present study we sought to investigate interactions between hypothalamic nitric oxide (NO) and ghrelin signaling on food intake and energy substrate utilization as measured by the respiratory exchange ratio (RER). Guide cannulae were unilaterally implanted in either the arcuate (ArcN) or paraventricular (PVN) nuclei of male Sprague-Dawley rats. Animals were pretreated with subcutaneous (2.5-10mg/kg/ml) or central (0-100pmol) N-nitro-l-Arginine methyl ester (l-NAME) followed by 50pmol of ghrelin administered into either the ArcN or PVN. Both l-NAME and ghrelin were microinjected at the onset of the active cycle and food intake and RER were assessed 2h postinjection. RER was measured as the ratio of the volume of carbon dioxide expelled relative to the volume of oxygen consumed (VCO2/VO2) using an open-circuit indirect calorimeter. Our results demonstrated that peripheral and central l-NAME pretreatment dose-dependently attenuated ghrelin induced increases in food intake and RER in either the ArcN or PVN. In fact the 100pmol dose largely reversed the metabolic effects of ghrelin in both anatomical regions. These findings suggest that ghrelin enhancement of food intake and carbohydrate oxidation in the rat ArcN and PVN is NO-dependent.

Mesolimbic exendin-4 attenuates reward salience evoked by neuropeptide Y and ghrelin.

In the present study, we investigated the effects of the glucagon-like peptide-1 (GLP-1) agonist exendin-4 (Ex-4) on the stimulatory action of neuropeptide Y (NPY) and ghrelin. These effects were examined in relation to operant responding for palatable food or voluntary ethanol intake in a two-bottle limited access paradigm. Male Sprague Dawley rats, each with ventral tegmental area (VTA) unilateral guide cannulae, were used. Ex-4 was paired with either NPY, ghrelin, or combined NPY and ghrelin treatment. Our results indicated that while NPY and ghrelin reliably stimulated operant responding for sucrose pellets and increased ethanol intake, Ex-4 suppressed intake and, most importantly, significantly reduced the effects of NPY and ghrelin. Overall, this work provides compelling evidence that VTA GLP-1, NPY, and ghrelin systems interact within the brain to modulate reward salience.

Differential effects of intra-ventral tegmental area ghrelin and glucagon-like peptide-1 on the stimulatory action of D-amphetamine and cocaine-induced ethanol intake in male Sprague Dawley rats.

In order to further elucidate the role of mesolimbic peptides in the expression of ethanol reward, the present study investigated the effects of ghrelin and glucagon-like peptide-1 (GLP-1) on ethanol intake, in addition to ethanol intake stimulated by systemic d-amphetamine or cocaine treatment. While a number of studies suggest that ghrelin plays an important role in mesolimbic reward, emerging data now indicate that GLP-1 receptor mechanisms inhibit reward signaling, possibly by directly or indirectly inhibiting ghrelinergic activity within the mesolimbic system. In the present study all rats were initially habituated to a 6% ethanol solution. We then demonstrated that intraperitoneal injections of d-amphetamine and cocaine increased ethanol intake compared to the vehicle condition. In subsequent testing we examined the effects of ventral tegmental area (VTA) ghrelin or vehicle paired with a fixed dose of d-amphetamine or vehicle. In separate rats we then investigated the impact of the GLP-1 agonist exendin-4 (Ex-4), injected into the VTA, on ethanol intake alone, or when Ex-4 was co-administered with d-amphetamine or cocaine. Our results indicated that VTA ghrelin significantly increased ethanol intake, and most importantly, potentiated the effect of d-amphetamine and cocaine on ethanol consumption. Conversely, VTA Ex-4 inhibited ethanol intake and antagonized the stimulatory effect of d-amphetamine and cocaine on ethanol consumption. In a final study we further demonstrated that VTA Ex-4 treatment significantly inhibited the combined stimulatory effects of ghrelin paired with d-amphetamine or ghrelin paired with cocaine. Overall our findings are consistent with a critical role for both ghrelin and GLP-1 receptor mechanisms in mesolimbic ethanol reward circuitry. Moreover, our results further suggest that ghrelin and GLP-1 modulate the stimulatory effect of psychostimulants on ethanol intake.

Role of mesolimbic ghrelin in the acquisition of cocaine reward.

The present study examined the ability of ghrelin administration into either the ventral tegmental area (VTA) or nucleus accumbens (NAc) to potentiate cocaine-induced conditioned place preference (CPP). Additionally, we examined the impact of co-injection of the ghrelin 1a antagonist JMV 2959 with ghrelin in order to evaluate the potential attenuation of ghrelin's effects on cocaine-induced CPP. Adult male Sprague-Dawley rats were allowed simultaneous access to either side of the CPP apparatus to establish baseline chamber preferences. The rats were then restricted to either their non-preferred or preferred side over the course of conditioning which lasted for a total of 8 consecutive days. On days in which rats were restricted to their non-preferred side, systemic cocaine (0.5-5.0 mg/kg IP) followed by central ghrelin (300 pmol), or co-administration of ghrelin (300 pmol) with JMV 2959 (10 µg), was administered either into the VTA or NAc immediately prior to the conditioning period. On alternate days rats were treated with vehicle then placed into what was initially determined to be their preferred side. CPP was calculated as the difference in the percentage of total time spent in the treatment-paired compartment during the post-conditioning session and the pre-conditioning session. Our results indicated that ghrelin potentiated cocaine-induced CPP and that this effect was attenuated by JMV 2959. Overall, these findings provide further evidence that mesolimbic ghrelin signaling is indeed critically involved in mediating the rewarding effects of cocaine.

Accumbal ghrelin and glucagon-like peptide 1 signaling in alcohol reward in female rats.

The present study investigated the relationship between accumbal ghrelin and glucagon-like peptide 1 (GLP-1) signaling in alcohol reward in female rats. Animals with guide cannulae targeting the nucleus accumbens core (NAcC) and shell (NAcS) were habituated to alcohol for 12 weeks through a two-bottle intermittent access paradigm. JMV2959, a ghrelin antagonist, and exendin-4 (Ex-4), a GLP-1 agonist, were microinjected at the onset of the nocturnal cycle. Alcohol, food, water, and total fluid intake were measured 2, 6, and 24 h postinjection. Results indicated that JMV2959 reduced alcohol consumption when injected into both the NAcC and NAcS. Ex-4 administration as well as combined JMV2959 and Ex-4 treatment reduced intake when injected into the NAcS, but not the NAcC. These effects were time-dependent. JMV2959 had no effect on food intake when administered into either the NAcC or the NAcS, whereas Ex-4 decreased food intake when injected separately into both structures. The combination of JMV2959 and Ex-4 decreased food intake when administered only into the NAcC. These effects were also time-dependent. No estrous-related effects on alcohol or food intake were found. However, water and total fluid intake were increased during the metestrus and diestruses phases of the estrous cycle compared with the proestrus and estrus phases. Overall, these findings demonstrate the importance of accumbal ghrelin and GLP-1 signaling in alcohol reward and appetitive motivation.

Opioidergic and dopaminergic modulation of cost/benefit decision-making in Long Evans Rats.

Eating disorders are associated with impaired decision-making and dysfunctional reward-related neurochemistry. The present study examined the potential contributions of dopamine and opioid signaling to these processes using two different decision-making tasks. In one task, Long Evans Rats chose between working for a preferred food (high-carbohydrate banana-flavored sucrose pellets) by lever pressing on a progressive-ratio schedule of reinforcement vs. obtaining less preferred laboratory chow that was concurrently available. In a second (effort-free) task, rats chose between the same two reinforcers when they were both available freely. Rats were trained in these tasks before receiving haloperidol (0.00, 0.05, 0.10mg/kg, intraperitoneally (i.p.)) or naloxone (0.0, 1.5, 3.0mg/kg, i.p.). In the first task, haloperidol decreased breakpoint, lever presses, number of reinforcers earned, and increased chow intake, whereas naloxone decreased breakpoint and number of reinforcers earned but had no effect on chow consumption. In the effort-free task, haloperidol reduced intakes of both foods without affecting preference, whereas naloxone selectively reduced the consumption of banana-pellets. The present findings support converging evidence suggesting that DA signaling affects processes more closely related to appetitive motivation, leaving other components of motivation unchanged. By contrast, opioid signaling appears to mediate aspects of hedonic feeding by selectively altering intakes of highly palatable foods. For preferred foods, both appetitive and consummatory aspects of food intake were altered by opioid receptor antagonism. Our findings argue against a general suppression of appetite by either compound, as appetite manipulations have been shown to unselectively alter intakes of both types of food regardless of the task employed.

The glucagon-like peptide-1 analog exendin-4 antagonizes the effect of acyl ghrelin on the respiratory exchange ratio.

The present study investigated the interaction of hypothalamic arcuate nucleus (ArcN) ghrelin and glucagon-like peptide-1 (GLP-1) signaling on metabolic function. Using indirect calorimetry, we first showed that acylated ghrelin, administered into the ArcN, significantly increased the respiratory exchange ratio (RER) in male Sprague-Dawley rats, representing a shift in fuel utilization toward enhanced carbohydrate oxidation and reduced lipid utilization. In contrast, treatment with similar doses of des-acyl ghrelin failed to induce reliable changes in RER. We then examined the ability of exendin-4 (Ex4) to alter acyl ghrelin's energetic effects. Ex4 is a GLP-1 agonist and has been reported previously to suppress food intake in rodent models. Rats were treated with either systemic or direct ArcN Ex4, followed by acyl ghrelin. Our results indicated that both systemic and central injections of Ex4 alone significantly reduced RER and, importantly, Ex4 pretreatment reliably attenuated the impact of ghrelin on RER. Overall, these findings provide compelling evidence that ghrelin and GLP-1 signaling interact in the hypothalamic control of metabolic function.

Impact of [d-Lys(3)]-GHRP-6 and feeding status on hypothalamic ghrelin-induced stress activation.

Ghrelin administration directly into hypothalamic nuclei, including the arcuate nucleus (ArcN) and the paraventricular nucleus (PVN), alters the expression of stress-related behaviors. In the present study we investigated the effect of feeding status on the ability of ghrelin to induce stress and anxiogenesis. Adult male Sprague Dawley rats were implanted with guide cannula targeting either the ArcN or PVN. In the first experiment we confirmed that ArcN and PVN ghrelin treatment produced anxiety-like behavior as measured using the elevated plus maze (EPM) paradigm. Ghrelin was administered during the early dark cycle. Immediately after microinjections rats were placed in the EPM for 5min. Both ArcN and PVN treatment reduced open arm exploration. The effect was attenuated by pretreatment with the ghrelin 1a receptor antagonist [d-Lys(3)]-GHRP-6. In a separate group of animals ghrelin was injected into either nucleus and rats were returned to their home cages for 60min with free access to food. An additional group of rats was returned to home cages with no food access. After 60min with or without food access all rats were tested in the EPM. Results indicated that food consumption just prior to EPM testing reversed the avoidance of the open arms of the EPM. In contrast, rats injected with ghrelin, placed in their home cage for 60min without food, and subsequently tested in the EPM, exhibited an increased avoidance of the open arms, consistent with stress activation. Overall, our findings demonstrate that ghrelin 1a receptor blockade and feeding status appear to impact the ability of ArcN and PVN ghrelin to elicit stress and anxiety-like behaviors.

Midbrain raphe 5-HT1A receptor activation alters the effects of ghrelin on appetite and performance in the elevated plus maze.

Prior research suggests that midbrain serotonergic signaling and hypothalamic ghrelinergic signaling both play critical roles in appetitive and emotional behaviors. In the present study, we investigated the effects of median raphe nucleus (MRN) somatodentritic 5-HT1A receptor activation on the feeding-stimulant and anxiogenic action of paraventricular nucleus (PVN) ghrelin. In an initial experiment, adult male Sprague-Dawley rats were injected with either ghrelin (200-800 pmol) into the PVN or 8-OH-DPAT (2.5-10 nmol), a 5-HT1A receptor agonist, into the MRN. Performance on the elevated plus maze (EPM) was then assessed. In separate rats, MRN 8-OH-DPAT (2.5-5 nmol) was administered 5 min prior to PVN injection of ghrelin (400 pmol) followed by EPM testing. The orexigenic effects of MRN 8-OH-DPAT (0.1-1.6 nmol) paired with PVN ghrelin (50 pmol) were also examined. When administered alone into the PVN, ghrelin significantly decreased the number of entries and time spent in the open arms of the EPM. This anxiogenic effect was blocked if rats were allowed to eat immediately after ghrelin administration and then tested in the plus maze. MRN injections of 8-OH-DPAT were anxiolytic, and when rats were pretreated with 8-OH-DPAT prior to ghrelin, the anxiogenic action of the peptide was attenuated. In contrast, MRN administration of 8-OH-DPAT potentiated the eating-stimulant effect of PVN ghrelin. Overall, our findings demonstrate that ghrelinergic and serotonergic circuits interact in the neural control of eating and anxiety-like behaviors, with 5-HT1A receptor mechanisms potentiating the orexigenic action of ghrelin while inhibiting ghrelin-induced anxiogenesis as measured via the EPM.

Sex differences in the reversal of fluoxetine-induced anorexia following raphe injections of 8-OH-DPAT.

RATIONALE AND OBJECTIVES: Serotonin (5-hydroxytryptamine, 5-HT) is widely distributed in the central nervous system and it is well established that this neurotransmitter plays an inhibitory role in the control of ingestive behavior. Administration of various 5-HT agonists and selective serotonin reuptake inhibitors, including fluoxetine (FLU), suppress food intake under free-feeding and food-restricted conditions. In contrast, activation of somatodendritc 5-HT(1A) receptors in the raphe nuclei reduces forebrain 5-HT bioavailability and agonists of these receptors, including 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT), reliably stimulate eating behavior. METHODS: In the present study male ( n=8 per group) and female ( n=8 per group) rats were pretreated with 8-OH-DPAT in an attempt to reverse FLU's inhibitory effect on feeding. 8-OH-DPAT was injected into either the dorsal or median raphe of male and female rats at doses of 0.1-0.4 nmol. FLU was then injected IP at a dose of 2 mg/kg. Both compounds were administered just prior to the onset of the dark cycle. Food intake was measured 2 h post-injection. Similar effects were also measured in female rats after ovariectomy. RESULTS AND CONCLUSIONS: FLU suppressed food intake comparably in male and female rats. Dorsal and median raphe injections of 8-OH-DPAT dose dependently reversed the anorectic effect of FLU in male rats. Higher doses of 8-OH-DPAT completely antagonized this response. In female rats, however, 8-OH-DPAT pretreatment was largely ineffective except in ovariectomized females where results were similar to male rats. These findings suggest that male and female rats are differentially sensitive to the ability of 5-HT(1A) receptor agonists to antagonize the feeding suppressive action of FLU and imply a role for neuroendocrine mechanisms in enabling the somatodendritic autoreceptor to control serotonergic neurotransmission under these conditions.

Hypothalamic paraventricular 5-hydroxytryptamine: receptor-specific inhibition of NPY-stimulated eating and energy metabolism.

The feeding effects of 5-hydroxytryptamine (5-HT)(1) and 5-HT(2) receptor agonists injected into the hypothalamic paraventricular nucleus (PVN) immediately prior to PVN administration of neuropeptide Y (NPY) were examined. The impact of these same compounds on NPY-induced alterations in energy metabolism was also assessed in an attempt to characterize further the potential interactive relationship of PVN NPY and 5-HT on feeding and whole body calorimetry. Specifically, several experiments examined the effect of various 5-HT receptor agonists on NPY-stimulated eating and alterations in energy substrate utilization [respiratory quotient (RQ)]. This included the 5-HT(1A) receptor agonist 8-OH-DPAT, the 5-HT(1B/1A) agonist RU 24969, the 5-HT(1D) agonist L-694,247, the 5-HT(2A/2C) agonist DOI, the 5-HT(2B) agonist BW 723C86 and the 5-HT(2C) agonist mCPP. In feeding tests conducted at the onset of the dark cycle, drugs were administered 5 min prior to PVN injection of NPY and food intake was measured 2 h postinjection. The metabolic effects of NPY following a similar pretreatment were monitored using an open-circuit calorimeter measuring the volume of oxygen consumed (VO(2)), carbon dioxide produced (VCO(2)) and RQ (VCO(2)/VO(2)). PVN injection of NPY (100 pmol) potentiated feeding and evoked reliable increases in RQ. Only DOI (2.5--5 nmol) pretreatment antagonized NPY-induced eating and blocked the peptide's effect on energy substrate utilization. Direct PVN pretreatment with spiperone (SPRN), a 5-HT(2A) receptor antagonist, and ketanserin (KTSN), a 5-HT(2A/2C) antagonist, but not SDZ SER 082, a 5-HT(2B/2C) antagonist, or the 5-HT(2C) antagonist RS 102221, blocked the effect of DOI in both feeding and metabolic tests providing additional evidence that activation of PVN 5-HT(2A) receptors inhibits NPY's action on feeding and substrate utilization.

Activation of urocortin 1 and ghrelin signaling in the basolateral amygdala induces anxiogenesis.

Prior anatomical and functional studies have demonstrated the importance of the basolateral region of the amygdala in the regulation of anxiogenic and anxiolytic behaviors. In the present report we investigated the anxiety-inducing effects of the corticotropin-releasing hormone-related peptide urocortin 1 (Ucn1) and the gut-brain peptide ghrelin. Both peptides were injected directly into the basolateral amygdala of male Sprague-Dawley rats and performance in the elevated plus maze was assessed. Ghrelin was administered at doses of 3-300 pmol and Ucn1 at doses of 0.01-1.0 pmol. Separate groups of rats were pretreated with Ucn1 before ghrelin treatment. In all experiments each test was performed as a single trial per animal. Results indicated that both ghrelin and Ucn1 elicited an increase in anxiogenic behavior. Moreover, Ucn1 pretreament potentiated the anxiogenic action of ghrelin. Overall these findings provide support for an integrated role of ghrelin and urocortin signaling within the basolateral amygdala in the expression of anxiogenesis.

Ghrelin alters the stimulatory effect of cocaine on ethanol intake following mesolimbic or systemic administration.

Emerging evidence suggests that ghrelinergic and dopaminergic signaling interact in the neural control of motivation and ethanol reward. To further investigate a possible interaction between these two neurochemical systems, we examined the impact of ghrelin, cocaine and combined injections of ghrelin with cocaine, on voluntary ethanol intake. Male Sprague-Dawley rats were habituated to an 8% ethanol solution until intakes stabilized. Rats were then injected with ghrelin (2.5-10 nmol IP), cocaine (0.625-10 mg/kg IP) or ghrelin paired with cocaine. We also examined the impact of direct ghrelin (30-300 pmol) injections into the ventral tegmental area (VTA) co-administered with systemic cocaine. Ethanol consumption was measured at 2 and 6 h postinjection. While ghrelin and cocaine reliably increased ethanol intake, peripheral administration of the peptide elicited a dose-dependent differential effect on cocaine-induced intake. Pretreatment with ghrelin potentiated the effect of cocaine on ethanol intake at a low dose of 2.5 nmol, whereas 10 nmol suppressed cocaine-induced ethanol intake. This same 10 nmol dose was found to induce anxiogenic behavior as measured using an elevated plus maze paradigm. Finally, when injected directly into the VTA, ghrelin (300 pmol) potentiated the effect of systemic cocaine on ethanol intake. Combined subthreshold dosing of VTA ghrelin with a subthreshold dose of cocaine also evoked reliable increases in intake compared to vehicle. Overall, our data suggest that low doses of ghrelin elicit a stimulatory effect on cocaine-induced ethanol consummatory behavior and provide further support for an interactive role of dopaminergic and ghrelinergic transmission in ethanol reward.

Microinjection of Ghrelin into the Ventral Tegmental Area Potentiates Cocaine-Induced Conditioned Place Preference.

Prior work has shown that systemic cocaine pretreatment augments cocaine conditioned place preference (CPP) in rats. In contrast, ghrelin receptor antagonism attenuates cocaine and amphetamine-induced CPP. In order to further investigate ghrelin's role in dopamine-mediated reward, the present report examined whether pretreament with ghrelin, administered directly into the ventral tegmental area (VTA) of the midbrain, would potentiate the rewarding properties of cocaine as measured by CPP. Adult male Sprague-Dawley rats were given access to either side of the CPP chamber in order to determine initial side preferences. The rats were then restricted to either their non-preferred or preferred side over the course of conditioning which lasted for a total of 16 consecutive days. This was followed by a final test day to then reassess preference. On days where rats were confined to their non-preferred side, ghrelin (30 - 300 pmol) and cocaine (0.625 - 10 mg/kg IP) were administered immediately prior to the conditioning trial. On alternate days rats were treated with vehicle and placed into what was initially determined to be their preferred side. CPP was calculated as the difference in percentage of total time spent in the treatment-paired compartment during the post-conditioning session and the pre-conditioning session. Our results indicated that both cocaine and ghrelin elicited CPP and that ghrelin pre-treatment potentiated the effect of cocaine on place preference. Overall, these findings provide additional support for the argument that ghrelin signaling within the VTA enhances the rewarding effects of psychostimulant compounds.