Alcohol use disorder (AUD) is the most prevalent substance use disorder, with 3 million deaths per year that are attributable to alcohol (Grant et al., 2004, 2015; Hunt et al., 2020). In the United States alone, it is the third leading cause of preventable death, and according to the 2019 National Survey on Drug Use and Health¹, in the U.S., 14.5 million people aged 12 or older (5.3% of this age group) had AUD. This includes 9 million men (6.8% of men in this age group) and 5.5 million women (3.9% of women in this age group). Treatments for AUD include behavioral, psychosocial, and pharmacological approaches, with the goal of reducing drinking and achieving and maintaining long-term abstinence from alcohol use (Witkiewitz et al., 2019). Only three pharmacological compounds have been approved by the United States Food and Drug Administration (FDA) for the treatment of AUD: disulfiram (acetaldehyde dehydrogenase inhibitor), acamprosate (which may act as an N-methyl-D-aspartate receptor agonist), and naltrexone (non-specific opioid receptor antagonist; Rosner et al., 2010; Maisel et al., 2013; Witkiewitz et al., 2019). These medications are effective in a subset of AUD patients, and the development and application of these compounds have helped improve our understanding of AUD pharmacotherapeutics generally, but alternative pharmacological avenues must continue to be explored and tested to treat a greater proportion of AUD patients.

The orexin (Orx; also known as hypocretin) hypothalamic neuropeptide system has arisen as a system of interest to develop AUD-specific therapeutics (Moorman, 2018). Orexin has long been known to be involved in regulating a wide range of physiological processes, including arousal, feeding, energy expenditure, and stress-related behavior (de Lecea et al., 1998; Sakurai et al., 1998; Sutcliffe and de Lecea, 2000; Mieda and Yanagisawa, 2002; Berridge et al., 2010; Teske et al., 2010). More recently, the Orx system has been implicated in highly motivated behavior, in addition to negative aspects of behavior, which together may underlie problematic alcohol use (Lawrence, 2010; Mahler et al., 2012; Brown and Lawrence, 2013; Barson and Leibowitz, 2016; Walker and Lawrence, 2017; Moorman, 2018; Matzeu and Martin-Fardon, 2021). Although there is a dearth of clinical research, some evidence suggests dysregulation of the Orx system in patients with AUD. For example, individuals who were diagnosed with AUD exhibited high blood levels of Orx during early withdrawal (Bayerlein et al., 2011; Ziolkowski et al., 2016). Importantly, these elevated Orx levels correlated with stress-related symptomatology during that same time period of withdrawal (von der Goltz et al., 2011).

Most of our understanding of the Orx system and its specific contribution to alcohol dependence has come from preclinical studies. Orexin was shown to promote excessive alcohol drinking when injected in the nucleus accumbens core but not shell (Schneider et al., 2007; Brown et al., 2013), ventral tegmental area but not substantia nigra (Srinivasan et al., 2012), and the anterior but not posterior paraventricular thalamus (Barson et al., 2015). Orexin cells are strongly recruited during alcohol-seeking behavior. For example, in rodent models of AUD, an increase in the activation of Orx neurons was observed during the reinstatement of alcohol seeking that was induced by alcohol-associated discriminative cues (Dayas et al., 2008), discrete cues (Moorman et al., 2016), alcohol-associated contexts (Hamlin et al., 2007; Millan et al., 2010), and stress (yohimbine; Kastman et al., 2016).

The involvement of the Orx system in alcohol drinking and seeking has been confirmed by pharmacological manipulations of Orx receptor 1 (OrxR1) and OrxR2. The systemic blockade of OrxR1 decreased alcohol drinking in a two-bottle choice paradigm in high-alcohol-preferring Sprague Dawley rats (Moorman and Aston-Jones, 2009), decreased alcohol self-administration under an operant fixed-ratio schedule (Lawrence et al., 2006; Richards et al., 2008; Moorman et al., 2017), and decreased alcohol self-administration under a progressive-ratio schedule (Jupp et al., 2011). The systemic administration of an OrxR1 antagonist decreased alcohol-seeking behavior that was induced by alcohol-related stimuli (Lawrence et al., 2006; Martin-Fardon and Weiss, 2014; Moorman et al., 2017) and decreased the stress (yohimbine)-induced reinstatement of alcohol seeking (Richards et al., 2008). The peripheral administration of an OrxR2 antagonist reduced alcohol but not saccharin self-administration in rats (Shoblock et al., 2011). The systemic administration of a dual orexin receptor antagonist (DORA) decreased breakpoints and reduced alcohol consumption under a progressive ratio schedule in alcohol-preferring rats (Anderson et al., 2014).

Two general observations can be made based on the literature with regard to the relationship between the Orx system and alcohol dependence: (i) exposure to alcohol strongly recruits the Orx system, and (ii) blocking Orx receptors (OrxR1, OrxR2, and both) decreases alcohol use. Recent interest has been seen in repurposing the FDA-approved DORA suvorexant (SUV), marketed by Merck as Belsomra^®, for the treatment of insomnia (James et al., 2017; Campbell et al., 2020b). In addition to reducing drug craving (primarily via OrxR1), SUV may have the additional benefit of indirectly reducing relapse risk by normalizing sleep disturbances (primarily via OrxR2) that are commonly observed in AUD patients (Koob and Colrain, 2020).

One of the features of AUD is the significant increase of alcohol consumption (heavy drinking) to relieve or avoid withdrawal symptoms (Koob and Colrain, 2020). Moreover, knowing that stress is a major factor that contributes to the chronic relapsing and compulsive nature of substance use disorder, including AUD (Stephens and Wand, 2012), the present study’s objectives were to test the effect of SUV on two aspects of AUD: (i) the increase in alcohol consumption in alcohol-dependent rats and (ii) the stress-induced reinstatement of alcohol seeking.

The present study assessed the ability of SUV, an FDA-approved DORA for the treatment of insomnia, to be repurposed to reduce alcohol self-administration in animals with a history of alcohol dependence. This study also tested SUV’s ability to decrease the stress-induced reinstatement of alcohol-seeking behavior in alcohol-dependent rats. Alcohol-dependent rats exhibited an increase in alcohol self-administration (i.e., escalation) during alcohol dependence that was induced by chronic intermittent alcohol vapor exposure, which is consistent with several previous studies (O’Dell et al., 2004; Vendruscolo and Roberts, 2014; Matzeu et al., 2018a; Matzeu and Martin-Fardon, 2020). In agreement with previous reports, exposure to intermittent footshock stress reliably reinstated previously extinguished alcohol-seeking behavior (Le et al., 1999; Martin-Fardon et al., 2000; Matzeu and Martin-Fardon, 2020; Flores-Ramirez et al., 2022). Notably, SUV significantly reduced alcohol self-administration only in alcohol-dependent rats. Furthermore, SUV selectively blocked the stress-induced reinstatement of alcohol-seeking behavior in alcohol-dependent rats. These results support the hypothesis that both OrxR1 and OrxR2 play significant roles in drug self-administration and the reinstatement of drug-seeking behavior (Plaza-Zabala et al., 2012, 2013; Uslaner et al., 2014), further implicating the Orx system in maladaptive motivation, reflected by an increase in the motivation to drink and seek alcohol during dependence (Moorman and Aston-Jones, 2009; Moorman et al., 2017). Altogether, the results highlight the significance of targeting the Orx system for the treatment of substance use disorders, and SUV could be repurposed for the treatment of AUD.

Alcohol-dependent rats in the present study exhibited an increase in alcohol self-administration and greater somatic withdrawal signs during weeks 4–6 of intermittent alcohol vapor exposure. These results mirror findings that rats with a history of alcohol dependence exhibit an increase in alcohol self-administration (i.e., escalation) and somatic and motivational signs of withdrawal that are characteristic of dependence, which are observable at 6–8 h of abstinence from alcohol vapor (Roberts et al., 1996; O’Dell et al., 2004; Vendruscolo and Roberts, 2014; Matzeu et al., 2018a; Matzeu and Martin-Fardon, 2020). The present results add to the growing body of literature that shows that intermittent exposure to alcohol vapor elicits behavioral and neurobiological signs of dependence. One explanation is that these changes in neurobehavioral systems, especially those that underlie response inhibition and reward- and stress-related behavior, contribute to hyperkatifeia and thus negative reinforcement, whereby dependent subjects seek relief from negative symptoms that are exacerbated during alcohol withdrawal, further motivating them to seek and take alcohol, particularly during times of heightened stress (Koob, 2014).

The present results showed that 5, 10, and 20 mg/kg SUV significantly and selectively decreased alcohol self-administration in rats with a history of alcohol dependence, without affecting alcohol intake in non-dependent rats. These results are consistent with previous reports that manipulations of the Orx system, via the pharmacological blockade of OrxR1 and OrxR2, influence alcohol consumption in rodents (for review, see Kim et al., 2012). Indeed, systemic administration of the OrxR1 antagonist SB334867 decreased voluntary home-cage alcohol consumption (Moorman and Aston-Jones, 2009; Anderson et al., 2014) and reduced operant self-administration (Lawrence et al., 2006; Richards et al., 2008; Jupp et al., 2011). Similarly, the OrxR2 antagonist LSN2424100 decreased home-cage alcohol consumption in alcohol-preferring (P) rats and alcohol-preferring C57BL/6J mice (Anderson et al., 2014). Furthermore, the DORA almorexant (ACT-078573) reduced alcohol self-administration under both fixed- and progressive-ratio schedules of reinforcement and reduced home-cage alcohol drinking in rats and mice (Srinivasan et al., 2012; Anderson et al., 2014). The present findings, together with the extant literature, provide further evidence that the Orx system is a prominent player in the increase in alcohol consumption under conditions of alcohol dependence. The data further suggest that the Orx system is dysregulated as a function of alcohol exposure, demonstrated by the selective effect of SUV to decrease alcohol self-administration only in alcohol-dependent rats.

Intermittent footshock stress induced alcohol-seeking behavior in non-dependent and dependent rats under the vehicle condition (i.e., 0 mg/kg SUV), at a similar magnitude, even though alcohol dependent rats displayed greater active lever responses during dependence induction (X¯ = 81 ± 5 vs. X¯ = 46 ± 4, respectively at week 6). Importantly, 5 mg/kg SUV prevented the stress-induced reinstatement of alcohol-seeking behavior in dependent rats only. In agreement with the present findings, previous studies showed that peripheral (Martin-Fardon and Weiss, 2014) and central (intra-prelimbic cortex) injections of the OrxR1 antagonist SB334867 decreased reinstatement that was induced by alcohol-related stimuli (Martin-Fardon and Weiss, 2014; Brown et al., 2016), and that systemic SB334867 administration also reduced yohimbine (stress) induced reinstatement (Richards et al., 2008). More recently, an injection of the DORA TCS1102 in the posterior paraventricular nucleus of the thalamus prevented the footshock stress-induced reinstatement of alcohol-seeking behavior in alcohol-dependent rats only (Matzeu and Martin-Fardon, 2020). Altogether, these findings indicate that the Orx system plays a pivotal role in motivational aspects of alcohol-seeking behavior, and the antagonism of its target receptors may be an effective treatment to reverse Orx dysregulation that is induced by exposure to alcohol. Dysregulation of the Orx system by chronic alcohol is consistent with clinical observations in individuals with AUD who present high plasma Orx levels during acute withdrawal (Bayerlein et al., 2011; Ziolkowski et al., 2016) that were correlated with exacerbations in affect- and stress-related symptomatology (von der Goltz et al., 2011).

An interesting finding was that SUV administration selectively decreased alcohol taking and seeking in alcohol-dependent rats. A recurring theme in the expanding Orx literature is that the pharmacological blockade of OrxRs is more effective in subjects with high motivation for alcohol seeking or when alcohol drinking is exacerbated (i.e., alcohol preference or dependence induction). For example, the administration of an OrxR1 antagonist decreased alcohol self-administration and reinstatement behavior in rats that were trained to exhibit high motivation for alcohol (Moorman and Aston-Jones, 2009; Moorman et al., 2017) and in rats that were bred for high alcohol preference (Lawrence et al., 2006; Dhaher et al., 2010; Anderson et al., 2014). OrxR1 blockade selectively decreased escalated alcohol drinking in dependent but not non-dependent mice (Lopez et al., 2016), and blockade of both OrxR1 and OrxR2 decreased alcohol drinking in dependent rats in agreement with the present study (Aldridge et al., 2022). Ultimately, two recent studies from our group (Matzeu and Martin-Fardon, 2020; Flores-Ramirez et al., 2022) also found that the effects of a DORA on alcohol-seeking behavior are more robust in dependent animals than in non-dependent animals. A possible explanation for this phenomenon is that the unique contribution of Orx transmission to motivational aspects of alcohol taking and stress-induced alcohol seeking does not play a significant role until anti-reward systems are sufficiently engaged or recruited. Thus, in animals that are highly motivated to consume alcohol (e.g., alcohol-dependent animals), Orx transmission is potentially compromised and promotes the incentive for alcohol drinking and seeking through negative reinforcement mechanisms. Collectively, these findings may have significant clinical implications. Treatment with SUV may be beneficial for decreasing alcohol craving and relapse in individuals who have been diagnosed with AUD.

Although Orx is exclusively produced in the hypothalamus (HYP), including the lateral HYP, dorsomedial HYP, and perifornical area (Baldo et al., 2003; DiLeone et al., 2003; Winsky-Sommerer et al., 2004), Orx neurons project throughout the brain, densely innervating an array of brain regions that are involved in arousal, motivation, and responsivity to stress-related stimuli (Peyron et al., 1998; Baldo et al., 2003; Grafe and Bhatnagar, 2018). Although systemic approaches to understand how the Orx system influences motivational processes that underlie compulsive alcohol taking and seeking have been successful, it is important to also uncover unique contributions of discreet brain regions. Previous research showed that OrxR1 blockade in the ventral tegmental area (VTA) decreased the cue-induced reinstatement of alcohol seeking (Brown et al., 2016), and that intra-VTA DORA administration decreased alcohol self-administration (Srinivasan et al., 2012). Furthermore, targeted OrxR1 blockade in the medial prefrontal cortex decreased the cue-induced reinstatement of responding for alcohol, and the blockade of OrxR1 in the nucleus accumbens shell decreased alcohol self-administration (Lei et al., 2016), and decreased alcohol-seeking when the blockade occurred in the lateral HYP (Campbell et al., 2020a). The direct administration of a OrxR2 antagonist in the central nucleus of the amygdala decreased alcohol intake in mice (Olney et al., 2017), whereas an infusion of the DORA TCS1102 in the posterior paraventricular nucleus of the thalamus prevented the stress-induced reinstatement of alcohol-seeking behavior (Matzeu and Martin-Fardon, 2020). These findings suggest that Orx system activity throughout a wide system of brain regions is key in mediating behaviors that are related to alcohol taking and seeking. The present study administered SUV only systemically, but the brain regions mentioned above likely play a significant role, which requires further testing to delineate the exact anatomical and network bases of the behavioral effects of SUV.

One limitation of the present study was that female rats were not included in our experimental design, which limits generalizability of the results. The literature shows well-established sex differences in alcohol intake and preference in two-bottle choice tests (Li and Lumeng, 1984; Blanchard et al., 1993; Walker et al., 2008), differences in BALs after self-administration and somatic withdrawal signs during intermittent alcohol vapor exposure (Matzeu et al., 2018b), and differences in reactivity to rewarding and aversive properties of alcohol (Torres et al., 2014). Furthermore, orexins have also been found to mediate sex-dependent effects in stress responsivity (Grafe et al., 2017). Future studies should elucidate possible sex-specific effects of SUV on alcohol taking and seeking. Another possible limitation of this study is the relatively small number of rats used for testing SUV on the stress induced-reinstatement of alcohol-seeking behavior (n = 4/dose). Although the probability exists that a substantially larger cohort of rats might yield a different result, previous research from our laboratory argues against this possibility. In fact, it was shown that TCS1102, another DORA is more efficacious at reducing stress-induced reinstatement of alcohol-seeking behavior in rats with a history of alcohol dependence (Matzeu and Martin-Fardon, 2020; Flores-Ramirez et al., 2022), strongly supporting the beneficial effects of targeting both OrxR1 and OrxR2 to prevent stress-induced craving and relapse in individuals suffering form AUD. Individuals who are prescribed SUV for the treatment of insomnia are taking the medication before bedtime and are advised not to consume alcohol because of their possible additive effects, which may result in increased risk of somnolence (Sun et al., 2015). Therefore, an important concern will be the time during the day when SUV is administered to be the most efficacious at treating AUD and minimize daytime somnolence. In this context, several clinical trials with SUV have been initiated (i.e., ClinicalTrials.gov Identifier: NCT04229095 and NCT03897062) and further clinical research is warranted to determine the safest approach to administer SUV to patients suffering of AUD.

In summary, the present findings demonstrate that the administration of SUV, a currently FDA-approved treatment for insomnia, selectively decreased alcohol self-administration and the stress-induced reinstatement of alcohol-seeking behavior in animals with a history of dependence. The present results highlight the significance of targeting the Orx system for the treatment of substance use disorders and suggest that repurposing SUV could be a good alternative for the treatment of AUD to prevent compulsive-like drinking and stress-induced craving and relapse.

The original contributions presented in this study are included in the article/supplementary materials, further inquiries can be directed to the corresponding author.

The animal study was reviewed and approved by Institutional Animal Care and Use Committee of The Scripps Research Institute.

RM-F conceived the study. RM-F and BM designed the research. FF-R, JI, GP, and AM performed the experimental procedures, analyzed the results, and created the figures. All authors co-wrote the manuscript, contributed to data interpretation, and manuscript editing and approved the submitted version.