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A fixed dose combination of bupropion (BPP) and naltrexone (NTX), Contrave®, is an FDA approved pharmacotherapy for the treatment of obesity. A recent study found that combining BPP with low-dose NTX reduced alcohol drinking in alcohol-preferring male rats. To explore potential pharmacological effects of the BPP + NTX combination on alcohol drinking, both male and female C57Bl/6J mice were tested on one-week drinking-in-the dark (DID) and three-week intermittent access (IA) models. Neuronal proopiomelanocortin (POMC) enhancer knockout (nPE -/- ) mice with hypothalamic-specific deficiency of POMC, and its bioactive peptides melanocyte stimulating hormone and beta-endorphin, were used as a genetic control for the effects of the BPP + NTX. A single administration of BPP + NTX (10 mg/kg + 1 mg/kg) decreased alcohol intake after DID in C57Bl/6J males, but not females. Also in C57Bl/6J males, BPP + NTX reduced intake of the caloric reinforcer sucrose, but not the non-caloric reinforcer saccharin. In contrast, BPP + NTX had no effect on alcohol DID in nPE -/- males. Pretreatment with the selective melanocortin 4 receptor (MC4R) antagonist HS014 reversed the anti-dipsogenic effect of BPP + NTX on alcohol DID in C57Bl/6J males. In the 3-week chronic IA model, single or repeated administrations for four days of BPP + NTX reduced alcohol intake and preference in C57Bl/6J males only. The behavioral measures observed in C57Bl/6J mice provide clear evidence that BPP + NTX profoundly reduced alcohol drinking in males, but the doses tested were not effective in females. Furthermore, our results suggest a hypothalamic POMC/MC4R-dependent mechanism for the observed BPP + NTX effects on alcohol drinking in male mice., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Corticotropin-releasing factor (CRF), a neuropeptide released during stress, has been reported to modulate startle behavior, including reducing the threshold for acoustic startle responding and reducing prepulse inhibition (PPI). The central mechanisms mediating CRF system regulation of startle and PPI are still unclear. Some antipsychotic drugs attenuate CRF-induced deficits in PPI in rats and mice. Here we tested the hypothesis that indirect activation of DA(1)-receptors (D(1)) and DA(2)-receptors (D(2)) contributes to the effects of CRF on PPI. We compared the effect of central administration of h/r-CRF (0.2-0.6 nmol) on PPI in mice with either a D(1) or D(2) receptor null mutation (knockout, KO) or in mice pretreated with D(1) or D(2) receptor antagonists SCH23390 (1 mg/kg) or haloperidol (1 mg/kg). D(1) and D(2) KO mice exhibited no significant differences in their sensitivity to CRF-induced disruptions of PPI. Similarly, neither SCH23390 nor haloperidol pretreatment altered the CRF-induced disruption in PPI, although both increased PPI at baseline. CRF-induced increases in startle also remained unchanged by any of the DA receptor manipulations. These results indicate that neither D(1)- nor D(2)-receptor activation is necessary for CRF to exert its effects on acoustic startle and PPI in mice.

Endogenous opioids within the central nervous system are postulated to mediate hedonic aspects of feeding behavior. To identify the relevant endogenous opioid receptor ligands, mice lacking one or two of the opioid peptide families beta-endorphin, enkephalins or dynorphins were tested for sucrose preference in a two-bottle free-choice drinking paradigm under drug-naïve conditions and following treatment with an opioid antagonist (1 mg/kg naloxone i.p.) or saline. Basal sucrose consumption was unaltered in all of the knockout genotypes compared to their congenic wild-type C57BL/6 littermates during 0.5 and 6 h access to a bottle containing 2, 4, 8, or 16% sucrose and a second bottle containing water. Moreover, all mutant genotypes and wildtype mice exhibited a similar compensatory decrease in overnight food intake following the extra caloric load from 6 h sucrose access. Although these basal responses to sucrose were unaffected by the knockout genotypes, naloxone reduced sucrose consumption by 50% compared to saline treatment during the first 0.5 h in wild-type and beta-endorphin knockout mice, but had no effect in enkephalin knockouts, beta-endorphin and enkephalin double knockouts, or dynorphin knockouts. These data suggest that naloxone reduces sucrose consumption in wild-type mice by blocking endogenous enkephalin and/or dynorphin signaling, but not beta-endorphin. Dynorphin knockouts in the current study had bar-pressing responses for a palatable food reinforcer in an operant procedure under free-feeding conditions similar to wild-type mice while we found in a previous study that beta-endorphin and enkephalin knockout mice had reduced motivation to respond [Hayward MD, Pintar JE, Low MJ. Selective reward deficit in mice lacking beta-endorphin and enkephalin. J Neurosci 2002;22:8251-8258.]. We conclude from these studies directly comparing three strains of opioid peptide knockout mice that enkephalin and dynorphin can modulate sucrose preference but are not necessary to support sucrose consumption. However, dynorphin was not necessary to support wildtype levels of operant responding suggesting that only enkephalin and beta-endorphin modulate conditioned food reinforcement.

To test whether endogenous opioid peptides are necessary for the rewarding effects of ethanol, we examined operant oral self-administration of ethanol in mice congenic to the C57BL/6J strain but lacking expression of beta-endorphin, enkephalin or both peptides. The influences of prandial state, schedule interval and type, and ethanol concentration were all examined. Food-restricted subjects were tested in postprandial and preprandial states and subsequently at normal body weight when feeding ad libitum (ad lib). Operant studies were conducted using fixed ratio (FR) intervals of 2 and 8 as well as a progressive ratio (PR) interval of 2. The main significant effect relevant to our hypothesis was increased responding by female mice lacking beta-endorphin under ad lib feeding conditions and only for lower ethanol concentrations (3% and 6%). Importantly, all subjects including those lacking both beta-endorphin and enkephalins learned to self-administer ethanol similarly to wild-type mice and maintained responding for ethanol under a variety of procedural variables. Consequently, the two opioid peptides believed to be the endogenous ligands for the micro-opioid receptor (MOR) were not necessary to shape or perpetuate ethanol-reinforced operant responding. These results suggest that the rewarding effects of ethanol do not require beta-endorphin or enkephalin signaling.

Alcohol-induced activation of the opioid system may contribute to the reinforcing properties of alcohol. This study investigated whether elimination of beta-endorphin (BE) synthesis via site-directed mutagenesis in embryonic stem cells would alter alcohol intake in mice. Both BE-deficient and wildtype (WT) mice generated from the targeted stem cells were backcrossed for nine generations onto a C57BL/6 background, and were maintained with ad libitum food and water. Mice had access to alcohol (10% v/v) under the following conditions: 24 h, scheduled access for 2 h/day, following acute (1 or 2 days) or chronic (5 weeks) alcohol deprivation, and scheduled access following six doses of naltrexone (0.125-16.0 mg/kg BW, ip) or saline treatment. Alcohol intake was similar in BE-deficient and WT mice given chronic access to alcohol, but greater in BE-deficient compared with WT mice during the first 10 days of scheduled access to alcohol, but not after more extensive experience with scheduled access. BE-deficient, but not WT mice, increased alcohol intake following 2 days, but not 1 day or 5 weeks, of deprivation. Naltrexone reduced alcohol drinking both in BE-deficient and WT mice, suggesting that drinking is mediated, in part, by activation of opioid receptors in both genotypes.

Pharmacological blockade studies have supported a role of the dopamine system in ethanol reward for many years, but receptor subtype specificity has been difficult to establish. Recently, genetically engineered mice lacking functional dopamine D2 receptors have been shown to drink less ethanol in a two-bottle choice task. To determine whether reduced ethanol intake reflects a reduction in ethanol reward, D2 receptor-deficient [knockout (KO)] mice were compared to heterozygous (HET) and wild-type (WT; C57BL/6xDBA/2 F2 hybrid) mice in a place conditioning task. Under conditions that produced reliable place preference in both WT and HET mice, KO mice showed no evidence of place conditioning, suggesting that D2 receptor gene inactivation reduced ethanol reward or the ability to learn about ethanol reward. Consistent with previous findings, this mutation also produced a gene dose-related reduction in basal activity levels. Moreover, KO and HET mice showed enhancement of ethanol-stimulated activity relative to WT mice. However, differences in basal and ethanol-stimulated activity did not explain the differences in place conditioning. Overall, this study strongly supports the conclusion that dopamine D2 receptors normally influence ethanol reward in mice.