Highlights

• Abstinence-induced craving and reward-seeking occur in humans and animals, and they are thought to underlie relapse.

• We speculate that hypothalamic-thalamic-striatal circuitry is a key modulator of abstinence-induce reward-seeking.

• Biological sex affects incubation of craving depending on the reinforcer and the method of abstinence.

1. Introduction

“Semper in absentis felicior aestus amantis” (Propertius, Elegies 2.33b.21)

[“Passion is always greater in absent lovers”] (Kline, A.S. transl., Liber Publications 2001)

The compulsive drive to self-administer drugs of abuse or palatable foods (e.g., fatty or sweet foods) to the point it harms one’s social, psychological, and physical health defines Substance Use Disorder (SUD) and binge eating disorders/obesity. The patterns of behavior and neurobiological mechanisms are similar between the two (Millan et al., 2017), and both are considered chronic, relapsing brain diseases (Volkow et al., 2013). The major impediment to the treatment of these disorders is high rates of relapse after periods of abstinence (O’Brien, 1997). There are numerous external and internal factors that lead to relapse in both humans and animal models. Broadly these factors lead to craving, which is defined as an overwhelmingly strong desire or need to use a drug or eat a palatable food. In humans, the experience of craving is a juggernaut that typically leads to relapse and yet has been nearly impossible to fully understand at a neurobiological level. Clinical and preclinical studies have shown that craving and relapse can be triggered by at least one of the following factors: acute exposure to the reinforcing stimulus (i.e., drug or food), cues or contexts associated with positive or negative reinforcers, or stress (for review, see Venniro et al., 2016). Importantly, stress and cues associated with negative affective states can be independent of the substance disorder or due to abstinence-induced withdrawal symptoms (Chartoff & Carlezon, 2014).

In this review we focus on the effect of abstinence on either opioid- or palatable food-seeking. It is critical to note that “abstinence” is a broad term in the context of this review. First, it can trigger a withdrawal state in the absence of the reinforcer, which is known to contribute to craving and—in animal models—reinstatement of reward-seeking (e.g. negative reinforcement; (Evans & Cahill, 2016; Koob, 2020). Second, abstinence can be as short as a day or as long as months or years. A particularly insidious feature of addictive-like behavior--whether the motivation is to ingest drug or food--is that craving can increase over time with abstinence. Preclinical scientists have dubbed this “incubation of drug craving” (Grimm, 2001), and defined it as a hypothetical motivational process in which there is a time-dependent increase in cue-induced reward-seeking. This has been shown to occur after withdrawal from both opioid and food self-administration in rats and humans (Grimm, 2020; Zhou et al., 2009). Of note, the data from humans include the common caveat that the timing and methods of manipulating abstinence periods are variable, thus adding to the complexity of trying to understand this phenomenon solely in humans.

A number of outstanding, in-depth reviews have covered extinction- and abstinence-based models of relapse to drug/food seeking (see Venniro et al., 2016; Nair et al., 2009; Grimm, 2020). Here we focus on abstinence-induced seeking of opioids and palatable foods, the mechanisms of which both overlap and are distinct from those determined for psychostimulants (Badiani et al. 2011). In that vein, we seize upon a slew of recent studies that have expanded focus outward from the traditional corticolimbic dopamine system to include hypothalamic–thalamic–striatal circuitry (Millan et al., 2017; Otis et al., 2019). Specifically, we describe how the lateral hypothalamus (LH), paraventricular nucleus of the thalamus (PVT), and the nucleus accumbens (NAc) interconnect and contribute to abstinence-induced opioid- and (high fat or sweet) food-seeking. As hypothalamic and midline thalamic regions are integral to the regulation of arousal, energy metabolism, reward and aversion, this broader neural circuit offers many mechanistic answers and avenues for future research.

Evidence suggests that gender and biological sex modulate compulsive, addictive-like behavior, including incubation of craving and relapse (Hallam et al., 2016; McHugh et al., 2018). Since relapse has been identified as one of the key blocks to successful treatment, understanding sex-dependent mechanisms contributing to relapse is an essential component of scientific inquiry. As such, another goal of this review is to describe, when known, studies that reveal sex-dependent differences or comparisons.

2. Clinical relevance of abstinence-induced reward-seeking

Drugs of abuse and highly palatable foods can establish Pavlovian association with external stimuli previously associated with their consumption (Volkow et al., 2013). In humans, drug or food cues elicit a subjective craving state (“urge” or “desire”) to consume or seek the reward. These craving states can lead to relapse, which for both drugs and food, is a major hurdle in the treatment of substance use disorder (O’Brien, 1997) and weight loss in obese people (Nair, 2009). This is particularly important because the subjective craving for both drugs and food increases during abstinence.

2.1. Abstinence-induced food-seeking in humans

Cues such as the sight, smell, and taste of food reliably signal food intake and act as conditioned stimuli that can potentially trigger food-seeking. These food cue responses could increase the probability of overeating (Wardle, 1990). In people who restrain their food intake, food predictive cue presentations can lead to a strong desire to eat and subsequent binging (Sobik et al., 2005). Moreover, people who were exposed to high-fat foods during dieting are more likely to relapse to their unhealthy eating habits (Gorin et al., 2004). Thus, both abstinence from eating and exposure to food cues increases food cravings and increases food-seeking and food intake. Additionally, food deprivation has been shown to increase the physiological responses specifically to food-related cues (Drobes et al., 2001).

It is known that the primary cause of obesity is overeating. Human studies have shown that overweight individuals have stronger brain blood-oxygen-level-dependent (BOLD) imaging signals in response to food cues compared to healthy weight people (Frankort et al., 2012). These BOLD imaging signals have been shown to be strongest in the VTA, PFC, Amy, and the NAc (Martin et al., 2010). Additionally, clinical studies have shown that thalamic nuclei specifically activate in response to reward craving and reward cue presentation (George et al., 2001). However, the resolution in human imaging studies makes it difficult to distinguish the PVT from other thalamic nuclei. Nonetheless, thalamic nuclei closely interact with the mesocorticolimbic reward system and have been implicated in playing a key role in influencing food consumption and food-seeking in preclinical models (see Ferrario et al., 2016; Millan et al., 2017 for review).

There are clear biological sex differences in the regulation of food intake and body weight in humans (Woods et al., 2003). These differences have been reported to be driven by several factors such as: leptin and insulin sensitivity, gonadal hormones, sex chromosome-associated genes that influence energy homeostasis, fat distribution and appetite. Thus, understanding sex-specific mechanisms of the aforementioned factors is important to understand the development of obesity and overeating in both men and women. In women, caloric intake fluctuates across the menstrual cycle (Buffenstein et al., 1995). These changes in caloric intake are mediated by natural fluctuations of ovarian hormones throughout the cycle. For example, fluctuations in estradiol negatively predict shifts in food intake, progesterone shows a positive correlation, and the combination of both estradiol and progesterone mediates a periovulatory drop in eating (Roney and Simmons, 2017). A handful of studies have also shown that ovarian hormones play a key role in modulating the activity of the mesolimbic reward system nuclei (NAc, Amy and LH) in response to the presentation of food cues (Frank et al., 2010; Alonso-Alonso et al., 2011). Although obese or overweight men and women both exhibit increased responsivity to high-calorie food when compared to their lean counterparts, women tend to be more responsive. In addition, women show greater activation than men in cortical regions when food cues are presented (Killgore et al., 2010).

2.2. Abstinence-induced opioid-seeking in humans

Opioid use disorder (OUD) can be segmented into pathological use of illicit opioids (e.g., heroin) or prescription opioids (e.g., oxycodone). Patterns of OUDs have shifted recently, with an increase in concurrent prescription opioid- and heroin-use in people with OUD (Cicero et al., 2015). Although it has been difficult to obtain clear, direct evidence that opioid craving is heightened during periods of abstinence in humans, there are increasing studies examining stress, affect, impulsivity, and how triggers of these states change/increase over opioid abstinence periods. For example, it has been shown that although response inhibition is improved over time in heroin abstainers, that effect can be reduced by exposure to drug-related cues, which may increase the risk of relapse, and is a major impediment to treatment (Su et al., 2020). Increased opioid craving is positively correlated with stronger BOLD fMRI signals within mesocorticolimbic and other limbic regions of the brain, including cortex, dorsal and ventral striatum, thalamus, and hippocampus (Li et al., 2012). As discussed above, common triggers for craving and seeking include drug-paired cues, drug-paired contexts, and stress. Of note, the most effective trigger for opioid-seeking in dependent individuals is often exposure to the drug itself (McHugh et al., 2014).

Gender differences in OUD exist at multiple levels of the addiction cycle (McHugh et al., 2013). Not only do women report increased functional impairment and higher likelihood of misusing opioids to cope with negative affect and pain compared to men, women report significantly more craving (Back et al., 2011). These were not associated with medication dose or pretreatment sensitivity towards OUD. Such behavioral consequences span sensitivity to opioid-reward and negative affective state, and sensitivity to stress hormones in neural circuits that mediate opioid withdrawal-induced negative affective states (Chartoff and McHugh, 2016).

3. Preclinical models of abstinence-induced reward-seeking

In this section we will focus on abstinence-based relapse models. There are three phases to this model: training, abstinence and relapse testing. During training, animals self-administer either drugs or palatable food (high fat or sweet foods/sucrose) over several days. In each day’s self-administration session, responses on an active manipulandum are paired with a cue (auditory or visual stimulus), while simultaneously leading to reward (see Venniro et al., 2016 and Grimm, 2020 for reviews). Following training, rats undergo abstinence, which can be either forced or voluntary and importantly results in increased reward-seeking behaviors. The majority of studies utilize forced abstinence, in which animals are kept in their home cages and no longer have access to the reward. For relapse testing, rats are placed back in the operant chambers using conditions in which responding on the manipulandum results in cue presentation but no reward delivery (Counotte et al., 2014; Grimm, 2020). Depending on the length of abstinence, the original reinforcer, biological sex, and many other factors, cue-triggered responding (i.e., reward-seeking) is enhanced relative to early abstinence. This phenomenon of potentiated drug-seeking has been termed “incubation of craving” (Grimm et al., 2001; Grimm, 2020).

3.1. Abstinence-induced sucrose- and high-fat diet-seeking in rodent models

Incubation of food craving (e.g. sucrose, high-fat diets and saccharin) has been described in rodent models. The majority of studies examining incubation of food craving use a protocol slightly modified from that used with drugs of abuse. For example, studies by Grimm et al. use a palatable food self-administration training schedule similar to that used for drugs of abuse that incorporates a food-paired cue. Animals then undergo forced abstinence for a variable number of days. Unlike abstinence-induced drug-seeking tests; however, incubation of food craving is measured by ultimately allowing rats access to the active manipulandum (previously paired with palatable food) for several hours prior to re-introduction of cues. Broadly, the longer the “abstinence” period, the greater the response on the active manipulandum when cues are introduced. This is interpreted as incubation of craving (Grimm, 2020). Specifically, it has been found that after 15, 21 and 30 days of abstinence (Grimm, 2020; Li and Frantz, 2010; Counotte et al., 2014) from sucrose self-administration, rats have a time-dependent increase in cue-induced sucrose-seeking. Additionally, incubation of craving for high-fat/high-sugar diets has also been shown. For example, it was found that cues previously paired to high-fat food and standard chow pellets increase reward-seeking after 30 days of forced abstinence (Darling et al., 2016; McCue et al., 2019). Interestingly, no incubation of craving was found for chocolate pellets in rats (Noye Tuplin et al., 2018). Although saccharin does not have any caloric value, it is highly palatable (sweet) and saccharin-associated cues can trigger seeking behaviors after 30 days of abstinence in rats (Aoyama et al., 2014).

To date, only one study has directly compared sex differences in incubation of sucrose craving (Madangopal et al., 2019). Although both males and females showed incubation of sucrose craving, this study did not find overt sex differences in sucrose-seeking after abstinence (1, 21, 60, 120 and 200 days). Other studies have examined the role of ovarian hormones on stress-induced food-seeking but no effects of ovarian hormones were observed (Calu et al., 2014; Pickens et al., 2011). Nevertheless, these findings do not exclude the possibility of ovarian hormones and the estrous cycle having a role in incubation of craving as it has been observed in humans and for other types of rewards (e.g. cocaine: Nicolas et al., 2019). Additionally, ovarian hormones and the estrous cycle have been previously shown to play a role in modulating the motivational responses to food cues (in the absence of reward delivery) in female rats (Alonso-Caraballo and Ferrario, 2019).

Importantly, either ovarian or testicular hormones can underlie sex-specific effects on reward-seeking. For example, abstinence from a junk-food diet enhanced NAcC glutamatergic transmission in males but not females (Alonso-Caraballo et al., 2020). Additionally, a role for testicular hormones in modulating the mesocorticolimbic reward system has also been described (Tobiansky et al., 2018). Overall, females show greater impulsive choice for food reward compared to males, and testicular hormones act to reduce impulsive choice in a food-seeking paradigm in males (Hernandez et al., 2020). As such, research studies examining the role of both ovarian and testicular gonadal hormones in reward-seeking (in general and during abstinence) is essential.

3.2. Abstinence-induced opioid-seeking in rodent models

Incubation of opioid craving has been described in animal models (Zhou et al., 2009), using procedures that incorporate forced or voluntary abstinence from drugs (Reiner et al., 2019). At the most basic level, forced abstinence involves removing the ability of the subject to obtain drug. The temporal effect of abstinence length on opioid-seeking behavior is consistent with an inverted U-shaped curve; there is typically a peak in opioid-seeking observed between 6–25 days post-abstinence, depending on conditions including the opioid itself, and whether context, cue, or stress is used to trigger reinstatement (Shalev et al., 2001).

In addition, there have been several recent studies examining incubation of opioid craving and relapse after voluntary abstinence in both males and females, including (Reiner et al., 2020; Venniro et al., 2019), and no sex differences have been observed in behavior. Interestingly, Venniro et al. (2019) showed that incubation of craving was observed in both sexes only after forced, but not voluntary, abstinence using their procedures (Venniro et al., 2017). Together, these findings suggest that the behavioral expression of incubation of opioid craving are similar in males and females, although it remains to be determined if the neurobiological mechanisms are the same. This is critical information for understanding the behavior itself and for considering translational studies aimed at mitigating incubation of craving. Interestingly, a recent study demonstrated that exogenously administered estradiol to freely cycling female rats nominally improved extinction of heroin-seeking, whereas a combination of estradiol and progesterone had a stronger impact (Vazquez et al., 2020). Although these findings point towards an important role of circulating gonadal hormones in regulating opioid-seeking, the study was under-powered to determine if estrous cycle stage was associated with levels of heroin seeking. As noted with Hernandez et al., 2020)above, it is also likely that male gonadal hormones play a role in incubation of craving. Regardless, these findings are consistent with a role for gonadal hormones in opioid seeking and further study is warranted.

4. Neural circuit-based mechanisms for abstinence-induced reward-seeking

There are multiple regions in the brain that mediate different aspects of reward-processing, motivation state, salience, and reward-seeking, with the mesocorticostriatal pathway being one of the most studied. Recent studies however have brought into sharp focus the effects of subcortical nuclei, such as the PVT and LH, and its interaction with the NAc. The LH in particular has emerged as a key brain region in mediating reward-seeking. The LH is the origin of brain-wide orexinergic projections (Peyron et al., 1998), which are involved in both arousal states and reward-related behavior. Accumulating evidence implicates LH orexin projections in mediating relapse to both drugs of abuse and natural rewards.

The PVT is considered part of hypothalamic–thalamic–striatal circuitry that integrates information related to motivation, reward, and energy balance control (Millan, 2017; Kelley et al., 2005). The major projections of the PVT are to the NAc, amygdala (Amy), and bed nucleus of the stria terminalis (BNST). These regions have been extensively studied in relation to motivated behavior, reward, aversion, and fear/anxiety (Kirouac, 2015). As such, the PVT is a lynchpin for coordinating responses to positive and negative affective states that lead to drug or food seeking and relapse to unregulated intake. The vast majority of PVT neurons project to the NAc (Dong et al., 2017), forming excitatory synaptic contact with NAc medium spiny neurons (MSNs). There is a distinct topography in which the anterior PVT (aPVT) projects preferentially to the dorsal NAc shell (NAcSh), whereas the posterior PVT (pPVT) projects preferentially to the ventromedial NAcSh and NAc core (NAcC) (Dong et al., 2017). This anatomical distinction is important, although not fully understood, because the dorsal and ventral subregions of the NAcSh can have opposing effects on motivated behavior (Al-Hasani et al., 2015; Marchant et al., 2009; Millan et al., 2017). The pPVT also projects strongly to the central nucleus of the amygdala (CeA), where it is understood to regulate the expression of fear responses (Do-Monte et al., 2015; Penzo et al., 2015). In this section we will focus on aspects of the hypothalamic–thalamic–striatal circuitry neuronal projections that mediate reward-seeking (see Fig.1).

Figure 1. Hypothalamic–thalamic–corticostriatal circuitry involved in food- and opioid-seeking.

An important consideration for the study of PVT projections and their functions is the high degree of collateralization in PVT projections. As an example, it was found through retrograde labeling that 50% of PVT neurons projecting to the dorsolateral BNST and the CeA also projected to the NAcSh (Dong, 2017). This extensive collateralization of PVT axons suggests this region coordinates a complex network of cortical and extended amygdala regions important for motivated behavior, reward-seeking, and emotional valence. To date, relatively little is known about how PVT collateralization translates to behavior, resulting in a gap in knowledge.

4.1. Neural circuit-based mechanisms for abstinence-induced food-seeking

Food cues elicit strong activation responses in the brain’s reward system areas. In this section, we will discuss potential neural circuit-based mechanisms underlying incubation of food craving. Our main focus will be on circuits composed of connections among the PVT, LH, Amy, and NAc, as well as projection-specific manipulations and their behavioral outcomes (Fig. 1 and Table 1). Both the PVT and the NAc are strategically positioned as major interfaces between cortical, hypothalamic and mesolimbic structures. In addition, both PVT and NAc play key roles in the control of feeding behaviors, motivation, reward and learning (for an extensive review on the PVT see Millan et al., 2017).

TABLE 1:

Projection-specific role in food-seeking behaviors.

Abbreviations: Amy, Amygdala; BLA; Basolateral Amygdala; LH, Lateral Hypothalamus; NAc, Nucleus Accumbens; PVT, Paraventricular Nucleus of the Thalamus; PFC, Prefrontal Cortex; vmPFC, Ventro-medial PFC.

While the role of the PVT on abstinence-induced food-seeking or incubation of craving has not yet been meticulously examined, there are several studies that clearly describe the role of the PVT in modulating food cue motivated behaviors. For example, ibotenic acid lesions of the PVT increase motivational responses towards food-predictive cues in goal-tracker rats (Haight et al., 2015). These results suggest that the PVT plays a critical role in attenuating the attribution of incentive salience to reward-predictive cues. In contrast, PVT pharmacological inactivation with muscimol (GABA_A agonist; Do-Monte et al., 2017) increases cue-induced sucrose-seeking (only when expected reward is omitted; a frustrative condition). In addition, specific intra-PVT activation of glucagon-like peptide receptor leads to decreased cue-induced sucrose-seeking (Ong et al., 2017), whereas specific stimulation of glucose transporter 2 (GLUT2) in the PVT^GLUT2-NAc increases motivation to obtain sucrose in an operant conditioning task (Labouébe et al., 2016). Furthermore, photostimulation of aPVT projections to NAcSh decreases sucrose-seeking (Do-Monte et al., 2017), whereas aPVT-NAcSh photoinhibition increases sucrose-seeking (when expected reward was omitted). Additionally, facilitation of pPVT orexin-A transmission facilitated neuronal responses in the NAcC and photostimulation of pPVT to NAcC pathway led to increased cue-induced sucrose-seeking (Meffre et al., 2019). Both photostimulation and photoinhibition of aPVT projections to the central nucleus of the amygdala (CeA) inhibits sucrose-seeking in rats (Do-Monte et al., 2017). Moreover, PFC-PVT-NAc connections are involved in formation of cue-reward associations (Otis et al., 2017; 2019), whereas LH-PVT-NAc connections are involved in reward consumption and initiation of feeding behaviors (Otis et al., 2019). It has also been found that photoinhibition of BLA to NAc projections decreases sucrose-seeking (Stuber et al., 2011). When taken together, the wide range of methods, anatomical subregions, behavioral outcomes, and complexity of the systems makes it impossible to ascribe a single function to the PVT in the context of palatable food-seeking. However, the combination of findings described suggests the PVT integrates stimulus valence, affective and arousal states, and metabolic balance to stimulate or suppress food-seeking. This has important ramifications for treatment of compulsive food-related disorders.

Unfortunately, little is known about the role of the estrous cycle or male/female gonadal hormones in the modulation of these pathways. However, sex-specific effects of abstinence from a junk-food diet has been found in obesity-susceptible rats. In this case, NAcC glutamatergic transmission was enhanced in males but not females (Alonso-Caraballo et al., 2020). Given that gonadal hormones play an important role in food intake and motivational responses to food cues, it will be important to extend this line of research to studies of incubation of craving for food reinforcers.

4.2. Neural circuit-based mechanisms for abstinence-induced opioid-seeking

Chemogenetic and optogenetic studies of the PVT-NAc and NAc-LH pathways play key roles in mediating various aspects of addictive-like behaviors such as withdrawal-associated aversion, retrieval, and relapse to opioid-seeking behavior (Keyes et al., 2020; Zhu et al., 2016). Glutamatergic PVT neurons project onto both dopaminergic D1 and D2 receptor-expressing MSNs in the NAcSh. However, repeated opioid exposure increases synaptic potentiation specifically at PVT-D2-MSNs synapses (Zhu et al., 2016). D2-MSNs that project onto neighboring D1-MSNs enhances the feed-forward inhibition of these D1-MSNs that in turn precipitates opioid-relapse. Direct activation of these D1 NAcSh–LH neurons prevents relapse (Keyes et al., 2020). The role of the PVT in addiction is an area of active exploration. The PVT has connections—some being reciprocal—with multiple reward/aversion-related regions of the brain, and numerous studies implicate these pathways in addictive-like behavior (Zhou & Zhu, 2019). LH orexinergic neurons play a major role in motivated behavior, including cocaine- and opioid-seeking in rodent models of addictive-like behavior (James et al., 2017). Studies pairing opioid self-administration and reinstatement models, as well as behavioral economics with systemic inhibition of orexin receptors have shown that the orexinergic system regulates motivation towards drug-seeking (James et al., 2017). However, the projection-specific role of orexinergic neurons in mediating motivation towards opioid-seeking remains to be explored. Studies from natural reward and LH–PVT projections (Meffre et al., 2019) lend credence to this. In addition, projections to the NAc from the ventro-medial prefrontal cortex (vmPFC) mediate opioid-seeking (Bossert et al., 2012). Context-induced heroin-reinstatement is mediated through interaction of vmPFC glutamatergic projections and postsynaptic dopaminergic D1 signaling.

5. Conclusions

This review summarizes a relatively nascent collection of studies suggesting a role for hypothalamic–thalamic–striatal circuitry in both opioid- and palatable food-seeking during abstinence—a strong preclinical model of relapse. Specifically, connections among the LH, PVT, NAc, and AMY serve to integrate internal and external signals related to affective state, arousal, metabolic balance, and motivation to drive relapse-related behaviors. The similarities between opioid- and food- seeking behavior and neural mechanisms are striking and often contrary to literature on psychostimulants (Badiani et al., 2011).

Few studies have as yet directly compared behaviors and neural mechanisms of incubation of craving in both males and females. On balance, the studies that have been reported find similar behavioral responses: both males and females show an increase in reward-seeking with abstinence. This is critical information, as the development of effective treatments depends on how both males and females would respond. It also highlights the importance of digging deeper to parse out the molecular, cellular, and circuit-based mechanisms for behavior. As discussed, gonadal hormones regulate reward-seeking and reward-based behavior, and since estrogen and testosterone are obviously different hormones, it is likely that different mechanisms underlie behavioral responses. This is a rich area for discovery.

Through this review, our goal was to describe what is known thus far and then identify some key gaps in knowledge that we and others can address in future preclinical and clinical studies. For example, one gap is identifying the neural circuits and modulators that connect affective state to motivated behavior. Specifically, does cue-triggered reward-seeking during abstinence arise through positive or negative mood states, and what brain region(s) with hypothalamic–thalamic–striatal circuitry is necessary? Another gap in knowledge is detailed understanding of how neurotransmitters (e.g. glutamate, GABA) and neuromodulators (e.g. dopamine, orexin, dynorphin) interact within this circuitry to affect specific aspects of motivated behavior related to opioid- and palatable food-seeking. There are numerous other questions, but a final gap in knowledge raised in this review is a general lack of understanding of the impact of biological sex on the functioning of this fundamental circuit.

TABLE 2:

Projection-specific role in opioid-seeking behaviors.

Abbreviations: Amy, Amygdala; BLA; Basolateral Amygdala; LH, Lateral Hypothalamus; NAc, Nucleus Accumbens; PVT, Paraventricular Nucleus of the Thalamus; PFC, Prefrontal Cortex; vmPFC, Ventro-medial PFC.

Abbreviations:

Amy: amygdala

BLA: basolateral amygdala

BOLD: blood-oxygen-level-dependent imaging

CeA: central amygdala

GLUT2: glucose transporter 2

LH: lateral hypothalamus

MSNs: medium spiny neurons

NAc: nucleus accumbens

NacC: NAc core

NAcSh: NAc shell

PVT: paraventricular nucleus of the thalamus

aPVT: anterior PVT

pPVT: posterior PVT

PFC: prefrontal cortex

OUD: opioid use disorder

SUD: substance use disorder

vmPFC: ventro medial PFC

Highlights

Strong desires for rewards and seeking them can increase after a period of not having them. This is seen in both humans and animals and is believed to cause relapse. Brain pathways involving the hypothalamus, thalamus, and striatum are thought to be important in controlling reward-seeking after abstinence. The impact of biological sex on the increase in craving over time depends on the type of reward and how abstinence is managed.

Introduction

Substance Use Disorder (SUD) and binge eating disorders, including obesity, involve a strong, uncontrolled urge to consume drugs or highly appealing foods, such as fatty or sweet items. This consumption continues to a degree that it negatively impacts social, psychological, and physical well-being. The brain mechanisms for these disorders are similar, and both are considered chronic, relapsing brain diseases. A significant challenge in treating these disorders is the high rate of relapse following periods of abstinence. Many external and internal factors lead to relapse in both humans and animal models. These factors generally lead to craving, defined as an overwhelmingly strong desire or need to use a drug or eat a palatable food. In humans, craving is an overwhelming feeling that often leads to relapse, but its brain mechanisms have been difficult to fully grasp. Studies in humans and animals show that craving and relapse can be triggered by immediate exposure to the drug or food, cues or situations linked to these rewards, or stress. Stress and cues linked to negative emotional states can arise independently or from withdrawal symptoms during abstinence.

This review focuses on the effect of abstinence on opioid- or palatable food-seeking. "Abstinence" is a broad term in this context. First, it can trigger a withdrawal state when the reward is absent, which contributes to craving and, in animal models, to the return of reward-seeking. Second, abstinence periods can range from a day to months or years. A particularly concerning feature of addiction-like behavior, whether the motivation is to ingest drugs or food, is that craving can increase over time with abstinence. Scientists have named this "incubation of drug craving," defining it as a process where cue-induced reward-seeking increases over time during abstinence. This phenomenon has been observed after withdrawal from both opioid and food self-administration in rats and humans. Human data often comes with the challenge that the timing and methods of abstinence periods vary, which complicates understanding this phenomenon in people alone.

Several comprehensive reviews have discussed extinction- and abstinence-based models of relapse to drug/food seeking. This review examines abstinence-induced seeking of opioids and palatable foods. The brain mechanisms for these behaviors share similarities but also differ from those for psychostimulants. Recent studies have broadened the focus beyond the traditional corticolimbic dopamine system to include the hypothalamic–thalamic–striatal circuitry. Specifically, the review describes how the lateral hypothalamus (LH), paraventricular nucleus of the thalamus (PVT), and the nucleus accumbens (NAc) connect and contribute to the seeking of opioids and high-fat or sweet foods during abstinence. Since the hypothalamus and midline thalamus are vital for controlling arousal, energy use, reward, and aversion, this wider brain circuit offers many insights and directions for future research.

Evidence suggests that biological sex influences compulsive, addiction-like behaviors, including the incubation of craving and relapse. Because relapse is a major obstacle to successful treatment, understanding how sex-specific mechanisms contribute to it is an essential part of scientific research. Therefore, another goal of this review is to describe studies that reveal sex-dependent differences or comparisons where known.

Clinical Relevance of Abstinence-Induced Reward-Seeking

Drugs of abuse and highly appealing foods can create learned associations with external cues previously linked to their use. In humans, drug or food cues trigger a strong desire or urge to consume or seek the reward. These craving states can lead to relapse, which for both drugs and food, is a major hurdle in the treatment of substance use disorder and in weight loss for individuals with obesity. This is particularly important because the subjective craving for both drugs and food increases during abstinence.

Abstinence-Induced Food-Seeking in Humans

Cues such as the sight, smell, and taste of food reliably signal food intake and act as conditioned stimuli that can potentially trigger food-seeking. These food cue responses could increase the probability of overeating. In people who limit their food intake, exposure to cues that predict food can lead to a strong desire to eat and subsequent binging. Moreover, individuals on diets who are exposed to high-fat foods are more likely to return to unhealthy eating habits. Thus, both abstinence from eating and exposure to food cues increase food cravings and lead to more food-seeking and food intake. Additionally, lack of food increases physical responses specifically to food-related cues.

The primary cause of obesity is known to be overeating. Human studies have shown that overweight individuals exhibit stronger brain activity signals (measured by BOLD imaging) in response to food cues compared to individuals of healthy weight. This brain activity is strongest in areas like the VTA, PFC, amygdala (Amy), and nucleus accumbens (NAc). Additionally, clinical studies have shown that specific thalamic regions activate in response to reward craving and when reward cues are present. However, the detail level in human imaging studies makes it hard to tell the PVT apart from other thalamic areas. Nevertheless, thalamic regions work closely with the brain's reward system and are thought to be key in affecting food consumption and seeking, as shown in animal studies.

There are clear biological sex differences in how food intake and body weight are regulated in humans. These differences are reported to be driven by factors such as sensitivity to leptin and insulin, sex hormones, and genes linked to sex chromosomes that affect energy balance, fat distribution, and appetite. Therefore, understanding sex-specific mechanisms of these factors is important for understanding the development of obesity and overeating in both men and women. In women, food intake changes during the menstrual cycle. These changes are influenced by natural fluctuations of ovarian hormones throughout the cycle. For example, changes in estradiol predict reduced food intake, progesterone shows a positive link, and both hormones together lead to a drop in eating around ovulation. A few studies have also shown that ovarian hormones are important in adjusting the activity of reward system areas (NAc, Amy, and LH) when food cues are present. Although both overweight men and women show increased responses to high-calorie food compared to leaner individuals, women tend to be more responsive. Women also show greater brain activity in cortical regions than men when food cues are presented.

Abstinence-Induced Opioid-Seeking in Humans

Opioid Use Disorder (OUD) includes the harmful use of illegal opioids like heroin or prescription opioids such as oxycodone. Recently, patterns of OUD have changed, with more individuals using both prescription opioids and heroin at the same time. While it has been challenging to find clear evidence that opioid craving increases during abstinence in humans, there are growing studies examining stress, emotion, impulsivity, and how triggers for these states change or increase during opioid abstinence periods. For example, while the ability to stop a response improves over time for individuals abstaining from heroin, this improvement can be lessened by exposure to drug-related cues. This may increase relapse risk and is a significant barrier to treatment. Higher opioid craving is linked to stronger brain activity signals (BOLD fMRI) in reward and emotional brain regions, including the cortex, striatum, thalamus, and hippocampus. Common triggers for craving and seeking include drug-paired cues, drug-paired contexts, and stress. The most powerful trigger for opioid-seeking in dependent individuals is often exposure to the drug itself.

Biological sex differences in OUD are present at various stages of the addiction cycle. Women report greater difficulties in daily life and are more likely to misuse opioids to manage negative emotions and pain than men. Women also report significantly more craving. These differences were not associated with medication dose or initial sensitivity to OUD. These behavioral effects include differences in sensitivity to opioid reward, negative emotional states, and stress hormones within brain circuits that control negative feelings during opioid withdrawal.

Preclinical Models of Abstinence-Induced Reward-Seeking

This section focuses on abstinence-based relapse models. This model involves three main stages: training, abstinence, and relapse testing. During training, animals self-administer either drugs or appealing food (high-fat or sweet foods/sucrose) over several days. In each daily self-administration session, responses on an active lever or button are linked to a cue (sound or light) and lead to a reward. Following training, rats undergo abstinence, which can be either forced or voluntary and leads to increased reward-seeking behaviors. Most studies use forced abstinence, where animals are kept in their home cages without access to the reward. For relapse testing, rats return to the experimental chambers. Under these conditions, pressing the lever or button results in a cue but no reward. The amount of reward-seeking triggered by cues is increased compared to early abstinence, depending on factors such as the length of abstinence, the original reward, and biological sex. This heightened drug-seeking behavior is called "incubation of craving."

Abstinence-Induced Sucrose- and High-Fat Diet-Seeking in Rodent Models

The phenomenon of increased food craving over time (for rewards like sucrose, high-fat diets, and saccharin) has been observed in rodent studies. Most studies on the incubation of food craving use a method slightly adapted from those used for drugs of abuse. For example, studies use a palatable food self-administration training schedule similar to that used for drugs of abuse that incorporates a food-paired cue. Animals then experience forced abstinence for a varying number of days. Unlike abstinence-induced drug-seeking tests, incubation of food craving is measured by first allowing rats access to the active lever (previously linked to palatable food) for several hours before cues are reintroduced. Generally, longer abstinence periods lead to greater responses on the active lever when cues are introduced, which is seen as incubation of craving. Specifically, it has been found that after 15, 21, and 30 days of abstinence from sucrose self-administration, rats show an increase in cue-induced sucrose-seeking that grows with time. Incubation of craving has also been shown for high-fat/high-sugar diets. For example, cues previously linked to high-fat food and standard chow pellets increase reward-seeking after 30 days of forced abstinence. Interestingly, no increase in craving was found for chocolate pellets in rats. Saccharin, though it has no calories, is very sweet, and cues linked to it can trigger seeking behaviors in rats after 30 days of abstinence.

To date, only one study has directly compared sex differences in the incubation of sucrose craving. Although both males and females showed incubation of sucrose craving, this study did not find obvious sex differences in sucrose-seeking after abstinence (1, 21, 60, 120, and 200 days). Other studies have looked at how ovarian hormones affect stress-induced food-seeking, but no effects were observed. However, these results do not rule out a role for ovarian hormones and the estrous cycle in the incubation of craving, as seen in humans and for other types of rewards (e.g., cocaine). Additionally, ovarian hormones and the estrous cycle have been shown to adjust motivational responses to food cues (without reward delivery) in female rats.

Sex-specific effects on reward-seeking can be influenced by either ovarian or testicular hormones. For example, abstinence from a junk-food diet increased glutamate signaling in the NAc core of males, but not females. Testicular hormones have also been shown to play a role in regulating the mesocorticolimbic reward system. Overall, females show more impulsive choices for food rewards than males, and testicular hormones help reduce impulsive choice in males during food-seeking tasks. As such, research studying the role of both ovarian and testicular hormones in reward-seeking (generally and during abstinence) is crucial.

Abstinence-Induced Opioid-Seeking in Rodent Models

The incubation of opioid craving has been observed in animal models, using methods that involve either forced or voluntary abstinence from drugs. At the most basic level, forced abstinence means preventing the subject from accessing the drug. The effect of abstinence length on opioid-seeking behavior follows an inverted U-shaped pattern, usually peaking between 6 and 25 days after abstinence. This depends on factors such as the specific opioid and whether reinstatement is triggered by context, cues, or stress.

Several recent studies have investigated the incubation of opioid craving and relapse after voluntary abstinence in both males and females, finding no sex differences in behavior. Interestingly, one study showed that incubation of craving occurred in both sexes only after forced abstinence, not voluntary abstinence, using specific procedures. Overall, these findings suggest that males and females show similar behavioral patterns in the incubation of opioid craving, though the underlying brain mechanisms may differ and require further investigation. A recent study demonstrated that giving estradiol to female rats during their natural cycle slightly improved the reduction of heroin-seeking, while a combination of estradiol and progesterone had a greater effect. Although these findings point towards an important role for circulating sex hormones in regulating opioid-seeking, the study lacked enough statistical power to determine if the estrous cycle stage was linked to levels of heroin seeking. It is also likely that male sex hormones play a role in the incubation of craving. Regardless, these findings support the idea that sex hormones play a role in opioid seeking, and further research is needed.

Neural Circuit-Based Mechanisms for Abstinence-Induced Reward-Seeking

Many brain regions control different aspects of reward processing, motivation, importance, and reward-seeking. The mesocorticostriatal pathway is one of the most studied of these. Recent research has highlighted the roles of subcortical brain areas, such as the PVT and LH, and how they interact with the NAc. The LH, in particular, has been identified as a key brain region for controlling reward-seeking. The LH is the source of orexin projections throughout the brain, which are involved in both alertness and behaviors related to reward. Growing evidence suggests that LH orexin projections play a role in relapse to both drugs of abuse and natural rewards.

The PVT is part of a brain circuit (hypothalamic–thalamic–striatal) that combines information about motivation, reward, and energy balance. The PVT primarily projects to the NAc, amygdala (Amy), and bed nucleus of the stria terminalis (BNST). These areas have been widely studied for their roles in motivated behavior, reward, aversion, and fear/anxiety. Therefore, the PVT is a central hub for coordinating responses to positive and negative emotional states that drive drug or food seeking and relapse to uncontrolled intake. Most PVT neurons project to the NAc, forming connections that excite NAc medium spiny neurons (MSNs). There is a specific arrangement where the anterior PVT (aPVT) mainly projects to the dorsal NAc shell (NAcSh), while the posterior PVT (pPVT) mainly projects to the ventromedial NAcSh and NAc core (NAcC). This anatomical difference is important but not fully understood, as the dorsal and ventral parts of the NAcSh can have opposite effects on motivated behavior. The pPVT also projects strongly to the central nucleus of the amygdala (CeA), which is known to regulate fear responses. This section focuses on aspects of the hypothalamic–thalamic–striatal circuitry's neuronal projections that control reward-seeking.

Figure 1. Hypothalamic–thalamic–corticostriatal circuitry involved in food- and opioid-seeking.

An important consideration for studying PVT projections and their functions is the high degree of collateralization in PVT projections. For example, it was found through retrograde labeling that 50% of PVT neurons projecting to the dorsolateral BNST and the CeA also projected to the NAcSh. This extensive branching of PVT axons suggests this region coordinates a complex network of cortical and extended amygdala regions important for motivated behavior, reward-seeking, and emotional value. To date, relatively little is known about how PVT collateralization translates to behavior, resulting in a gap in knowledge.

TABLE 1:

Projection-specific role in food-seeking behaviors.

Abbreviations: Amy, Amygdala; BLA; Basolateral Amygdala; LH, Lateral Hypothalamus; NAc, Nucleus Accumbens; PVT, Paraventricular Nucleus of the Thalamus; PFC, Prefrontal Cortex; vmPFC, Ventro-medial PFC.

Neural Circuit-Based Mechanisms for Abstinence-Induced Food-Seeking

Food cues trigger strong activity in the brain's reward system areas. This section discusses possible brain circuit mechanisms that contribute to the incubation of food craving. The primary focus will be on circuits involving connections between the PVT, LH, Amy, and NAc, and how specific manipulations of these projections affect behavior. Both the PVT and NAc are key connection points between cortical, hypothalamic, and mesolimbic brain structures. Both PVT and NAc play crucial roles in controlling feeding behaviors, motivation, reward, and learning.

While the PVT's role in food-seeking during abstinence or incubation of craving has not been fully explored, several studies show its importance in controlling food cue-motivated behaviors. For example, damage to the PVT (using ibotenic acid) increases motivation toward food-predictive cues in rats that actively seek goals. These findings suggest that the PVT is crucial for reducing the importance or 'draw' of cues that predict rewards. In contrast, blocking PVT activity with muscimol (a GABAA agonist) increases cue-induced sucrose-seeking, especially when the expected reward is not given (a frustrating situation). In addition, specific activation of glucagon-like peptide receptors within the PVT leads to reduced cue-induced sucrose-seeking, whereas specific stimulation of glucose transporter 2 (GLUT2) in the PVT-NAc pathway increases the motivation to obtain sucrose in a task where behavior is learned through consequences.

Furthermore, activating aPVT projections to the NAc shell decreases sucrose-seeking, while inhibiting these same projections increases sucrose-seeking (when the expected reward is withheld). Additionally, increasing orexin-A transmission in the pPVT boosted neuronal responses in the NAc core, and activating the pPVT to NAc core pathway led to increased cue-induced sucrose-seeking. Both activating and inhibiting aPVT projections to the central nucleus of the amygdala (CeA) reduce sucrose-seeking in rats. Moreover, connections between the PFC, PVT, and NAc are involved in forming links between cues and rewards, while LH-PVT-NAc connections are involved in consuming rewards and starting feeding behaviors. It has also been found that inhibiting projections from the BLA to the NAc decreases sucrose-seeking. When taken together, the variety of methods, brain regions, behavioral results, and complex systems involved makes it impossible to assign a single role to the PVT in palatable food-seeking. However, the findings suggest that the PVT combines information about stimulus value, emotional and arousal states, and metabolic balance to either encourage or stop food-seeking. This has significant implications for treating compulsive eating disorders.

Unfortunately, little is known about how the estrous cycle or male/female sex hormones affect these pathways. However, sex-specific effects of abstinence from a junk-food diet have been observed in rats prone to obesity. In these cases, glutamate signaling in the NAc core was increased in males but not females. Since sex hormones are important for food intake and motivational responses to food cues, it is crucial to extend this research to studies of craving incubation for food rewards.

Neural Circuit-Based Mechanisms for Abstinence-Induced Opioid-Seeking

Studies using chemogenetic and optogenetic methods show that PVT-NAc and NAc-LH pathways are crucial for various addiction-like behaviors, including aversion linked to withdrawal, memory retrieval, and relapse to opioid-seeking. PVT neurons that use glutamate project to both D1 and D2 dopamine receptor-expressing MSNs in the NAc shell. However, repeated opioid exposure strengthens connections specifically at PVT-D2-MSN synapses. D2-MSNs that project to nearby D1-MSNs increase the forward inhibition of these D1-MSNs, which then contributes to opioid relapse. Direct activation of these D1 NAc shell–LH neurons prevents relapse. The role of the PVT in addiction is an area of ongoing research. The PVT has connections—some reciprocal—with many brain regions involved in reward and aversion. Many studies suggest these pathways are involved in addiction-like behavior.

LH neurons that use orexin are very important in motivated behavior, including the seeking of cocaine and opioids in animal models of addiction-like behavior. Studies combining opioid self-administration, relapse models, and behavioral economics with general blocking of orexin receptors have shown that the orexin system controls motivation for drug-seeking. However, the specific role of orexin neurons in controlling motivation for opioid-seeking, depending on their projections, still needs to be explored. Research on natural rewards and LH–PVT projections supports this idea. In addition, projections from the ventromedial prefrontal cortex (vmPFC) to the NAc control opioid-seeking. Relapse to heroin use triggered by context involves interactions between vmPFC glutamate projections and D1 dopamine signaling at the receiving neurons.

Conclusions

This review summarizes emerging research that suggests the hypothalamic–thalamic–striatal brain circuit plays a role in seeking both opioids and appealing foods during abstinence, which is a key animal model of relapse. Specifically, connections between the LH, PVT, NAc, and amygdala (Amy) combine internal and external signals related to emotions, alertness, metabolic state, and motivation to drive behaviors associated with relapse. The similarities in behavior and brain mechanisms for seeking opioids and food are notable and often differ from what is known about psychostimulants.

Few studies have directly compared the behaviors and brain mechanisms of craving incubation in both males and females. Overall, reported studies indicate similar behavioral responses: both males and females show increased reward-seeking during abstinence. This is crucial information, as effective treatments must consider how both males and females respond. It also emphasizes the importance of investigating the molecular, cellular, and circuit-level brain mechanisms behind these behaviors. As discussed, sex hormones regulate reward-seeking and reward-based behavior. Since estrogen and testosterone are distinct hormones, it is probable that different mechanisms drive these behavioral responses. This area offers many opportunities for new discoveries.

This review aimed to describe current knowledge and identify key gaps that can be addressed in future animal and human studies. For example, one gap is identifying the brain circuits and regulatory substances that link emotional state to motivated behavior. Specifically, does cue-triggered reward-seeking during abstinence result from positive or negative moods, and which brain region(s) within the hypothalamic–thalamic–striatal circuitry are essential? Another knowledge gap is a detailed understanding of how neurotransmitters (like glutamate, GABA) and neuromodulators (like dopamine, orexin, dynorphin) interact within this circuitry to influence specific aspects of motivated behavior related to seeking opioids and appealing foods. A final knowledge gap highlighted in this review is a general lack of understanding regarding the impact of biological sex on how this fundamental circuit functions.

TABLE 2:

Projection-specific role in opioid-seeking behaviors.

Abbreviations: Amy, Amygdala; BLA; Basolateral Amygdala; LH, Lateral Hypothalamus; NAc, Nucleus Accumbens; PVT, Paraventricular Nucleus of the Thalamus; PFC, Prefrontal Cortex; vmPFC, Ventro-medial PFC.

Abbreviations:

Amy: amygdala BLA: basolateral amygdala BOLD: blood-oxygen-level-dependent imaging CeA: central amygdala GLUT2: glucose transporter 2 LH: lateral hypothalamus MSNs: medium spiny neurons NAc: nucleus accumbens NacC: NAc core NAcSh: NAc shell PVT: paraventricular nucleus of the thalamus aPVT: anterior PVT pPVT: posterior PVT PFC: prefrontal cortex OUD: opioid use disorder SUD: substance use disorder vmPFC: ventro medial PFC

Key Insights

The intense desire for rewards, like drugs or certain foods, along with the drive to seek them out, increases after a period of not having them. This phenomenon is observed in both humans and animals and is believed to contribute to relapse. It is thought that specific brain circuits, involving the hypothalamus, thalamus, and striatum, play a crucial role in regulating this reward-seeking behavior during abstinence. Additionally, biological sex influences how this intense desire for rewards develops over time, depending on the type of reward and the method of abstinence.

Introduction

A strong, compulsive urge to consume drugs or highly appealing foods, often to the detriment of one's health, defines conditions like Substance Use Disorder and binge eating disorders. These conditions share similar behavioral patterns and brain mechanisms, both being considered chronic, relapsing brain diseases. A major challenge in treating these disorders is the high rate of relapse after periods of abstinence. Various factors, both external and internal, can trigger craving, an overwhelming desire for the drug or food, which often leads to relapse. However, understanding the brain's role in craving has been difficult. Research shows that craving and relapse can be set off by exposure to the reward itself, cues linked to the reward, or stress, including stress from withdrawal symptoms during abstinence.

This discussion focuses on how abstinence affects the desire for opioids and palatable foods. Abstinence itself can lead to withdrawal, which contributes to craving. The duration of abstinence can vary from a day to months or even years. A concerning aspect of addictive behavior is that craving can actually increase over time during abstinence, a concept known as "incubation of craving." This refers to a time-dependent increase in reward-seeking triggered by cues, and it has been observed after withdrawal from both opioid and food consumption in rats and humans.

The following sections will explore abstinence-induced seeking of opioids and palatable foods, highlighting both shared and distinct mechanisms. Recent studies have expanded focus beyond the traditional dopamine system to include a brain network involving the hypothalamus, thalamus, and striatum. Specifically, the roles of the lateral hypothalamus (LH), paraventricular nucleus of the thalamus (PVT), and the nucleus accumbens (NAc) and their connections in abstinence-induced reward-seeking will be described. These brain regions are vital for regulating arousal, metabolism, reward, and aversion, offering many insights into the underlying mechanisms.

It is also recognized that gender and biological sex influence compulsive behaviors, including the incubation of craving and relapse. Since relapse is a major barrier to successful treatment, understanding sex-dependent mechanisms is an essential area of research. This review will therefore include findings that reveal differences related to biological sex when available.

Clinical Relevance: Food Seeking in Humans

Drugs and highly palatable foods can become linked with external cues, such as their sight, smell, or taste. In humans, these cues create a subjective feeling of craving or desire, which can lead to relapse. For both drugs and food, relapse is a significant challenge in treatment. This is particularly important because the subjective craving for both drugs and food often increases during abstinence.

Food cues can increase the likelihood of overeating. For individuals who restrict their food intake, exposure to cues predicting food can lead to a strong desire to eat and subsequent binge eating. People exposed to high-fat foods while dieting are also more likely to return to unhealthy eating habits. Thus, both abstaining from eating and being exposed to food cues increase food cravings and the desire to seek and consume food. Additionally, being deprived of food can heighten physiological responses specifically to food-related cues.

Overeating is a primary cause of obesity. Studies in humans show that overweight individuals exhibit stronger brain activity (measured by BOLD imaging) in response to food cues compared to those of healthy weight. This activity is strongest in areas like the VTA, prefrontal cortex, amygdala, and nucleus accumbens. Clinical research also indicates that thalamic nuclei activate in response to reward craving and cue presentation. While human imaging studies often lack the detail to distinguish specific thalamic nuclei, the thalamus is known to interact closely with the brain's reward system and plays a key role in influencing food consumption and seeking in animal models.

Clear differences exist between biological sexes in how food intake and body weight are regulated. These differences are influenced by factors such as hormone sensitivity, gonadal hormones, and sex chromosome-associated genes that affect energy balance and appetite. Understanding these sex-specific mechanisms is crucial for comprehending the development of obesity and overeating in both men and women. In women, caloric intake changes throughout the menstrual cycle, driven by natural fluctuations in ovarian hormones. For example, changes in estradiol negatively predict shifts in food intake, while progesterone shows a positive correlation. Both hormones together contribute to a drop in eating during ovulation. Studies also show that ovarian hormones influence activity in the brain's reward system (nucleus accumbens, amygdala, and lateral hypothalamus) in response to food cues. Although both obese men and women show increased responsiveness to high-calorie food cues compared to leaner individuals, women tend to be more responsive and exhibit greater activation in cortical regions.

Clinical Relevance: Opioid Seeking in Humans

Opioid use disorder (OUD) involves the problematic use of illicit or prescription opioids. There has been a recent increase in people with OUD using both prescription opioids and heroin. While direct evidence that opioid craving significantly increases during abstinence in humans has been difficult to obtain, studies are exploring how stress, mood, impulsivity, and related triggers change over periods of opioid abstinence. For instance, even though the ability to control responses improves over time in people abstaining from heroin, exposure to drug-related cues can lessen this improvement, increasing relapse risk. Increased opioid craving is linked to stronger brain activity signals within reward-related areas of the brain, including the cortex, striatum, thalamus, and hippocampus. Common triggers for craving and seeking include drug-paired cues, drug-paired contexts, and stress. Notably, for dependent individuals, the most effective trigger for opioid-seeking is often exposure to the drug itself.

Gender differences are evident throughout the addiction cycle for OUD. Women report more functional impairment and a higher likelihood of misusing opioids to cope with negative emotions and pain compared to men. Women also report significantly more craving, which is not tied to medication dose or prior sensitivity to OUD. These behavioral consequences extend to sensitivity to opioid rewards, negative emotional states, and stress hormones within brain circuits that mediate negative affective states caused by opioid withdrawal.

Preclinical Models: Food Seeking in Rodents

This section focuses on abstinence-based models of relapse in animals. These models typically involve three phases: training, abstinence, and relapse testing. During training, animals learn to self-administer either drugs or palatable foods (like high-fat or sweet items) over several days. Each self-administration session pairs a response, such as pressing a lever, with a cue (auditory or visual stimulus) that also leads to the reward. After training, rats undergo a period of abstinence, which can be either forced or voluntary, and importantly, results in increased reward-seeking behaviors. Most studies use forced abstinence, where animals are kept in their home cages without access to the reward. For relapse testing, rats are returned to the experimental chambers under conditions where their responses trigger cues but do not deliver the reward. Depending on the length of abstinence, the original reward, biological sex, and other factors, cue-triggered responding (reward-seeking) is enhanced compared to early abstinence. This intensified drug-seeking is termed "incubation of craving."

The incubation of food craving, for substances like sucrose, high-fat diets, and saccharin, has been observed in rodent models. Most studies use a protocol similar to that for drugs of abuse, involving self-administration training with a food-paired cue, followed by varying days of forced abstinence. Unlike drug-seeking tests, however, incubation of food craving is often measured by first allowing rats access to the active device (previously paired with palatable food) for several hours before reintroducing the cues. Generally, longer abstinence periods lead to a greater response on the active device when cues are presented, which is interpreted as incubation of craving. Specifically, rats have shown a time-dependent increase in cue-induced sucrose-seeking after 15, 21, and 30 days of abstinence from sucrose self-administration. Incubation of craving has also been shown for high-fat/high-sugar diets, where cues previously paired with these foods increased reward-seeking after 30 days of forced abstinence. Interestingly, no incubation of craving was found for chocolate pellets. Even saccharin, which has no caloric value but is highly palatable, can trigger seeking behaviors after 30 days of abstinence in rats.

Only one study has directly compared sex differences in the incubation of sucrose craving, finding that both males and females showed incubation, but no obvious sex differences in sucrose-seeking after abstinence periods of various lengths (1 to 200 days). Other research has looked at the role of ovarian hormones in stress-induced food-seeking, but no effects were observed. However, these findings do not rule out the possibility that ovarian hormones and the estrous cycle play a role in incubation of craving, as seen in humans and with other rewards like cocaine. Ovarian hormones and the estrous cycle have also been shown to influence motivational responses to food cues in female rats.

Importantly, both ovarian and testicular hormones can affect sex-specific reward-seeking. For example, abstinence from a junk-food diet increased glutamatergic transmission in a specific brain region (nucleus accumbens core) in males but not females. Testicular hormones have also been shown to modulate the brain's reward system. Overall, females tend to show more impulsive choices for food rewards compared to males, and testicular hormones in males reduce impulsive choices in food-seeking tasks. Therefore, research examining the role of both ovarian and testicular hormones in reward-seeking, especially during abstinence, is essential.

Preclinical Models: Opioid Seeking in Rodents

Incubation of opioid craving has also been documented in animal models using procedures that involve either forced or voluntary abstinence from drugs. Forced abstinence simply means removing the animal's ability to obtain the drug. The length of abstinence typically shows an inverted U-shaped effect on opioid-seeking behavior, with a peak often observed between 6 to 25 days post-abstinence. This timing depends on factors such as the specific opioid, and whether context, cues, or stress are used to trigger a return to drug-seeking.

Several recent studies have investigated the incubation of opioid craving and relapse after voluntary abstinence in both male and female animals, generally finding no sex differences in behavior. Interestingly, one study found that incubation of craving occurred in both sexes only after forced, but not voluntary, abstinence under their specific conditions. These findings suggest that the observable behaviors related to incubation of opioid craving are similar in males and females, though the underlying brain mechanisms might differ. This is crucial for understanding the behavior and for developing treatments to reduce incubation of craving. Furthermore, a recent study indicated that administering estradiol to female rats improved the reduction of heroin-seeking behavior, with a combination of estradiol and progesterone having an even stronger effect. While these results suggest an important role for circulating gonadal hormones in regulating opioid-seeking, the study was not powerful enough to determine if the estrous cycle stage was directly linked to heroin-seeking levels. It is also probable that male gonadal hormones play a role in the incubation of craving. Regardless, these findings support the idea that gonadal hormones influence opioid-seeking, and further research is warranted.

Neural Circuits: Brain Mechanisms for Reward Seeking

Multiple brain regions mediate different aspects of reward processing, motivation, and reward-seeking, with the mesocorticostriatal pathway being a widely studied area. However, recent research has highlighted the significant role of subcortical nuclei, such as the paraventricular nucleus of the thalamus (PVT) and the lateral hypothalamus (LH), and how they interact with the nucleus accumbens (NAc). The LH, in particular, has emerged as a key brain region for mediating reward-seeking, as it is the source of widespread orexinergic projections throughout the brain. These orexin projections are involved in both arousal states and behaviors related to reward. Growing evidence suggests that LH orexin projections contribute to relapse for both drugs of abuse and natural rewards.

The PVT is considered part of a brain network involving the hypothalamus, thalamus, and striatum, which integrates information about motivation, reward, and energy balance. The PVT primarily projects to the NAc, amygdala (Amy), and bed nucleus of the stria terminalis (BNST). These regions are extensively studied for their roles in motivated behavior, reward, aversion, and fear or anxiety. Thus, the PVT is crucial for coordinating responses to positive and negative emotional states that drive drug or food seeking and lead to uncontrolled intake. Most PVT neurons project to the NAc, forming excitatory connections. There is a specific organization where the anterior PVT (aPVT) mainly projects to the dorsal NAc shell (NAcSh), while the posterior PVT (pPVT) projects more to the ventromedial NAcSh and the NAc core (NAcC). This anatomical difference is important, though not fully understood, because the dorsal and ventral parts of the NAcSh can have opposite effects on motivated behavior. The pPVT also sends strong projections to the central nucleus of the amygdala (CeA), where it is known to regulate fear responses. These described brain connections are involved in reward-seeking.

An important aspect of PVT projections is their high degree of collateralization, meaning single neurons send branches to multiple targets. For example, it has been found that many PVT neurons projecting to the dorsolateral BNST and CeA also project to the NAcSh. This extensive branching suggests that the PVT coordinates a complex network of cortical and extended amygdala regions important for motivated behavior, reward-seeking, and emotional valence. Currently, little is known about how this collateralization of PVT axons translates into specific behaviors, representing a gap in understanding.

Neural Circuits: Food Seeking

Food cues trigger strong activation in the brain's reward system. This section will discuss potential brain mechanisms underlying the incubation of food craving, focusing on circuits involving the PVT, LH, Amy, and NAc, as well as how specific pathways influence behavior. Both the PVT and NAc are strategically positioned as major links between cortical, hypothalamic, and mesolimbic structures. They both play key roles in controlling feeding behaviors, motivation, reward, and learning.

While the role of the PVT in abstinence-induced food-seeking or incubation of craving has not been thoroughly examined, several studies clearly describe its role in modulating food cue-motivated behaviors. For example, damage to the PVT increases motivational responses to food-predictive cues in rats, suggesting the PVT helps reduce the importance attributed to reward-predictive cues. Conversely, pharmacological inactivation of the PVT increases cue-induced sucrose-seeking, especially when the expected reward is withheld. Furthermore, specific activation of a glucagon-like peptide receptor within the PVT leads to decreased cue-induced sucrose-seeking, while stimulating a specific glucose transporter in the PVT increases motivation for sucrose.

Direct stimulation of aPVT projections to the NAcSh decreases sucrose-seeking, whereas inhibiting these projections increases sucrose-seeking when the expected reward is omitted. Additionally, enhanced orexin-A transmission from the pPVT boosts neuronal responses in the NAcC, and stimulating the pPVT to NAcC pathway increases cue-induced sucrose-seeking. Both stimulating and inhibiting aPVT projections to the central nucleus of the amygdala (CeA) can inhibit sucrose-seeking. Connections between the prefrontal cortex, PVT, and NAc are involved in forming cue-reward associations, while LH-PVT-NAc connections are involved in reward consumption and starting feeding behaviors. Inhibiting basolateral amygdala to NAc projections also decreases sucrose-seeking. The diverse methods, brain regions, behavioral outcomes, and system complexity make it challenging to assign a single function to the PVT in palatable food-seeking. However, the combined findings suggest the PVT integrates information about stimulus value, emotional and arousal states, and metabolic balance to either stimulate or suppress food-seeking, which has important implications for treating compulsive food-related disorders.

Unfortunately, little is known about the role of the estrous cycle or male/female gonadal hormones in modulating these pathways. However, sex-specific effects of abstinence from a junk-food diet have been found in obesity-prone rats. In these animals, glutamatergic transmission in the NAcC was enhanced in males but not females. Given that gonadal hormones significantly influence food intake and motivational responses to food cues, it will be important to extend this research to studies of incubation of craving for food rewards.

Neural Circuits: Opioid Seeking

Studies using advanced genetic and light-based techniques have shown that pathways between the PVT and NAc, and between the NAc and LH, are crucial for mediating various aspects of addictive behaviors, such as withdrawal-related aversion, memory retrieval, and relapse to opioid-seeking. Glutamatergic neurons from the PVT project to both D1 and D2 receptor-expressing neurons in the NAcSh. However, repeated opioid exposure specifically increases synaptic strength at connections between the PVT and D2-type neurons. These D2-type neurons project to neighboring D1-type neurons, enhancing an inhibitory feedback that contributes to opioid relapse. Activating these D1 NAcSh–LH neurons can prevent relapse. The role of the PVT in addiction is an active area of research, as it connects, often reciprocally, with multiple brain regions involved in reward and aversion, with numerous studies linking these pathways to addictive behaviors.

LH orexinergic neurons play a major role in motivated behavior, including cocaine and opioid-seeking in animal models of addiction. Studies using opioid self-administration and relapse models, combined with systemic inhibition of orexin receptors, have shown that the orexinergic system regulates motivation for drug-seeking. However, the specific role of these orexinergic neurons in mediating motivation for opioid-seeking, based on their projection targets, still needs exploration. Studies on natural rewards and LH–PVT projections support this idea. Additionally, projections from the ventromedial prefrontal cortex (vmPFC) to the NAc mediate opioid-seeking. Heroin relapse triggered by context is mediated by the interaction of glutamatergic projections from the vmPFC and postsynaptic D1 dopamine signaling.

Conclusions

This overview summarizes a growing body of research that points to a significant role for a specific brain network, involving the hypothalamus, thalamus, and striatum, in both opioid and palatable food-seeking during abstinence. This is a strong model for studying relapse. Specifically, the connections between the lateral hypothalamus (LH), paraventricular nucleus of the thalamus (PVT), nucleus accumbens (NAc), and amygdala (Amy) integrate internal and external signals related to emotional state, arousal, metabolic balance, and motivation, ultimately driving behaviors associated with relapse. The similarities in both behavior and brain mechanisms between opioid and food seeking are notable and often differ from findings related to psychostimulants.

Few studies have directly compared behaviors and brain mechanisms of incubation of craving in both males and females. Generally, reported studies show similar behavioral responses: both males and females exhibit increased reward-seeking with abstinence. This information is vital because the development of effective treatments depends on understanding how both sexes would respond. It also underscores the importance of further investigation into the molecular, cellular, and circuit-level mechanisms underlying these behaviors. As discussed, gonadal hormones regulate reward-seeking, and since estrogen and testosterone are distinct hormones, it is likely that different mechanisms underpin the behavioral responses in males and females. This represents a fertile area for scientific discovery.

The goal of this review was to outline current knowledge and identify key gaps that researchers can address in future studies. For instance, one gap is identifying the specific brain circuits and modulators that link emotional state to motivated behavior. Specifically, does cue-triggered reward-seeking during abstinence stem from positive or negative mood states, and which regions within the hypothalamic–thalamic–striatal circuitry are essential for this? Another gap is a detailed understanding of how neurotransmitters (like glutamate, GABA) and neuromodulators (like dopamine, orexin, dynorphin) interact within this circuitry to affect specific aspects of motivated behavior related to opioid and palatable food seeking. Many other questions remain, but a final gap highlighted is the general lack of understanding of how biological sex impacts the functioning of this fundamental brain circuit.

Highlights

• Strong urges and reward-seeking behaviors after stopping drug or food use happen in both people and animals. These are believed to be major causes of relapse.

• Specific brain pathways involving the hypothalamus, thalamus, and striatum are thought to be key in controlling reward-seeking after a period of not using.

• A person's biological sex affects how these urges grow over time, depending on the type of reward and how long they have stopped using it.

Introduction

A powerful, uncontrolled desire to use drugs or highly appealing foods, even when it harms one's social life, mental health, and physical well-being, is a defining characteristic of Substance Use Disorder (SUD) and binge eating disorders. Both conditions share similar behaviors and brain processes and are considered long-term, relapsing brain diseases. A major challenge in treating these disorders is the high rate of relapse after periods of not using. Many factors, both external and internal, contribute to relapse in people and in animal studies. Generally, these factors lead to craving, which is an extremely strong desire or need to use a drug or eat a specific food. In people, the experience of craving often leads to relapse, but it has been hard to fully understand how it works in the brain. Studies show that craving and relapse can be triggered by direct exposure to the rewarding substance, by sights or situations linked to the reward, or by stress. It is important to note that stress and cues linked to negative feelings can arise independently or from withdrawal symptoms caused by stopping the substance.

This review focuses on how stopping drug or food use affects the desire to seek opioids or appealing foods. The term "abstinence" here covers a wide range. First, it can lead to withdrawal, which is known to cause craving and, in animal models, a return to reward-seeking. Second, the period of not using can last anywhere from a day to months or even years. A particularly tricky aspect of addictive behaviors is that craving can actually increase over time when a person stops using. Scientists call this "incubation of drug craving," meaning a time-dependent increase in the desire for a reward when reminded of it by cues. This has been observed after stopping both opioid and food self-administration in rats and people. However, human studies are complicated because the timing and methods of stopping are often varied.

This discussion highlights recent studies that have expanded beyond the usual brain areas to include pathways involving the hypothalamus, thalamus, and striatum. Specifically, it describes how the lateral hypothalamus (LH), paraventricular nucleus of the thalamus (PVT), and the nucleus accumbens (NAc) are connected and how they contribute to seeking opioids and high-fat or sweet foods after not using them. Since the hypothalamus and middle thalamus are crucial for controlling alertness, energy use, reward, and aversion, studying this wider brain network offers many explanations and new directions for future research.

Evidence suggests that a person's sex influences compulsive, addictive behaviors, including the growth of craving and relapse. Since relapse is a key obstacle to successful treatment, understanding how sex-specific differences affect relapse is essential. Therefore, another goal of this review is to describe studies that show sex-dependent differences when they are known.

Clinical Relevance of Abstinence-Induced Reward-Seeking

Drugs and highly appealing foods can create learned connections with things that were present during their use. In people, sights or situations related to drugs or food trigger a feeling of craving or desire to consume the reward. These craving states can lead to relapse, which is a major barrier in treating substance use disorder and in helping people with obesity lose weight. This is particularly important because the subjective craving for both drugs and food increases during periods of not using them.

Abstinence-Induced Food-Seeking in Humans

Cues like the sight, smell, and taste of food naturally signal eating and act as triggers that can potentially cause a person to seek food. These responses to food cues can increase the likelihood of overeating. For people who try to limit their food intake, seeing food-related cues can lead to a strong desire to eat and then to binge eating. Also, people on a diet who are exposed to high-fat foods are more likely to return to unhealthy eating habits. This means that both not eating and being exposed to food cues increase food cravings and the desire to seek and eat food. Additionally, going without food has been shown to increase physical responses specifically to food-related cues.

It is known that overeating is the main cause of obesity. Studies in people show that overweight individuals have stronger brain activity when responding to food cues compared to people of healthy weight. This stronger brain activity is most noticeable in the VTA, PFC, Amygdala, and NAc. Additionally, studies have shown that parts of the thalamus specifically become active in response to craving and when presented with reward cues. However, human brain imaging studies often cannot clearly distinguish the PVT from other thalamic areas. Nevertheless, parts of the thalamus work closely with the brain's reward system and are believed to play a key role in influencing food consumption and food-seeking in animal models.

There are clear biological sex differences in how food intake and body weight are regulated in people. These differences are caused by several factors, such as sensitivity to hormones like leptin and insulin, sex hormones, and genes on sex chromosomes that affect energy balance, fat distribution, and appetite. Therefore, understanding these sex-specific mechanisms is important for understanding how obesity and overeating develop in both men and women. In women, the amount of food eaten changes during the menstrual cycle. These changes are controlled by natural shifts in ovarian hormones throughout the cycle. For example, changes in estrogen levels tend to predict decreases in food intake, progesterone shows a positive link, and the combination of both estrogen and progesterone causes a drop in eating around ovulation. Some studies have also shown that ovarian hormones play a key role in adjusting the activity of brain reward centers in response to food cues. Although both obese or overweight men and women show increased reactivity to high-calorie food compared to leaner individuals, women tend to be more reactive. Additionally, women show greater activity than men in brain areas involved in thinking when food cues are presented.

Abstinence-Induced Opioid-Seeking in Humans

Opioid Use Disorder (OUD) can involve the problematic use of illegal opioids (like heroin) or prescription opioids (like oxycodone). Patterns of OUD have changed recently, with more people with OUD using both prescription opioids and heroin. While it has been hard to get clear, direct proof that opioid craving increases during periods of not using in people, a growing number of studies are looking at stress, mood, impulsivity, and how these triggers change or increase during opioid abstinence. For instance, it has been shown that even though the ability to control responses improves over time in people who have stopped using heroin, this improvement can be reduced by exposure to drug-related cues. This may increase the risk of relapse and is a major obstacle to treatment. Increased opioid craving is linked to stronger brain activity signals within reward-related and other emotional areas of the brain, including the cortex, striatum, thalamus, and hippocampus. As mentioned before, common triggers for craving and seeking include cues, environments, and stress linked to the drug. Notably, the most effective trigger for opioid-seeking in dependent individuals is often exposure to the drug itself.

Differences in OUD exist for males and females at various stages of the addiction cycle. Women not only report more problems in daily life and a higher chance of misusing opioids to cope with negative feelings and pain compared to men, but they also report significantly more craving. These differences were not linked to medication dose or previous sensitivity to OUD. These behavioral effects include sensitivity to opioid reward and negative emotional states, as well as sensitivity to stress hormones in brain circuits that control negative feelings during opioid withdrawal.

Preclinical Models of Abstinence-Induced Reward-Seeking

This section focuses on animal models that study relapse after a period of not using. This model has three phases: training, abstinence, and relapse testing. During training, animals learn to self-administer either drugs or appealing food (high-fat or sweet foods/sucrose) over several days. In each training session, pressing a specific lever is linked to a cue (sound or light) and also leads to getting the reward. After training, rats undergo abstinence, which can be forced or voluntary, and importantly, leads to increased reward-seeking behaviors. Most studies use forced abstinence, where animals are kept in their home cages without access to the reward. For relapse testing, rats are put back into the testing chambers under conditions where pressing the lever results in the cue appearing but no reward is given. Depending on how long the abstinence lasts, the original reward, the animal's sex, and many other factors, the cue-triggered seeking of the reward is stronger compared to early abstinence. This effect of increased drug-seeking has been called "incubation of craving."

Abstinence-Induced Sucrose- and High-Fat Diet-Seeking in Rodent Models

The increase in food craving (for example, for sucrose, high-fat diets, and saccharin) has been observed in rodent models. Most studies looking at this "incubation" of food craving use a method slightly changed from what is used for drugs. For instance, studies use a palatable food self-administration training schedule similar to that for drugs, which includes a food-linked cue. Animals then go through forced abstinence for a varying number of days. However, unlike drug-seeking tests after abstinence, the increase in food craving is measured by eventually allowing rats access to the lever (previously linked with palatable food) for several hours before cues are reintroduced. Generally, the longer the period of not using, the stronger the response on the lever when cues are presented. This is seen as an increase in craving. Specifically, studies have found that after 15, 21, and 30 days of stopping sucrose self-administration, rats show a time-dependent increase in cue-induced sucrose-seeking. Additionally, an increase in craving for high-fat/high-sugar diets has also been shown. For example, cues previously linked to high-fat food and standard chow pellets increase reward-seeking after 30 days of forced abstinence. Interestingly, no increase in craving was found for chocolate pellets in rats. Although saccharin has no calories, it is very sweet, and cues linked to saccharin can trigger seeking behaviors after 30 days of abstinence in rats.

So far, only one study has directly compared sex differences in the increase of sucrose craving. While both males and females showed this increase, the study did not find clear sex differences in sucrose-seeking after abstinence (at 1, 21, 60, 120, and 200 days). Other studies have looked at the role of ovarian hormones on stress-induced food-seeking, but no effects of ovarian hormones were observed. However, these findings do not rule out the possibility that ovarian hormones and the estrous cycle play a role in the increase of craving, as seen in humans and for other types of rewards. Additionally, ovarian hormones and the estrous cycle have been shown to affect how female rats respond to food cues (without reward delivery).

Importantly, either ovarian or testicular hormones can cause sex-specific effects on reward-seeking. For example, stopping a junk-food diet increased certain brain signals in the NAc in males but not females. Additionally, testicular hormones have been described as playing a role in adjusting the brain's reward system. Overall, females tend to make more impulsive choices for food rewards compared to males, and testicular hormones help reduce impulsive choice in males in a food-seeking situation. Therefore, research studying the role of both ovarian and testicular hormones in reward-seeking (generally and during abstinence) is essential.

Abstinence-Induced Opioid-Seeking in Rodent Models

The increase in opioid craving has been observed in animal models, using methods that involve forced or voluntary periods of not using drugs. At its simplest, forced abstinence means taking away the animal's ability to get the drug. The effect of how long abstinence lasts on opioid-seeking behavior typically follows a pattern: opioid-seeking usually peaks between 6–25 days after stopping, depending on factors like the specific opioid, and whether the environment, a cue, or stress is used to trigger a return to seeking.

In addition, several recent studies have looked at the increase in opioid craving and relapse after voluntary abstinence in both male and female animals, and no sex differences in behavior have been found. Interestingly, one study showed that this increase in craving was observed in both sexes only after forced, but not voluntary, abstinence, using their methods. Together, these findings suggest that the behavioral expression of increased opioid craving is similar in males and females, though it is still unknown if the brain mechanisms are the same. This is crucial information for understanding the behavior itself and for considering how these findings might apply to human treatments aimed at reducing this increase in craving. Interestingly, a recent study showed that estrogen given to female rats improved their ability to stop heroin-seeking, while a combination of estrogen and progesterone had a stronger effect. While these findings suggest an important role for circulating sex hormones in controlling opioid-seeking, the study did not have enough power to determine if the stage of the estrous cycle was linked to levels of heroin seeking. It is also likely that male sex hormones play a role in the increase of craving. Regardless, these findings are consistent with sex hormones having a role in opioid seeking, and further study is needed.

Neural Circuit-Based Mechanisms for Abstinence-Induced Reward-Seeking

Multiple brain regions control different aspects of reward processing, motivation, importance, and reward-seeking. The pathway connecting the midbrain, cortex, and striatum is one of the most studied. However, recent studies have strongly highlighted the effects of brain areas below the cortex, such as the PVT and LH, and how they interact with the NAc. The LH, in particular, has emerged as a key brain region in controlling reward-seeking. The LH is the source of widespread brain signals involving orexin, which are involved in both alertness and reward-related behavior. A growing body of evidence suggests that LH orexin pathways are involved in the return to seeking both drugs and natural rewards.

The PVT is considered part of a brain network that connects the hypothalamus, thalamus, and striatum. This network combines information about motivation, reward, and energy balance control. The main connections from the PVT go to the NAc, amygdala (Amy), and bed nucleus of the stria terminalis (BNST). These areas have been extensively studied for their role in motivated behavior, reward, aversion, and fear/anxiety. As such, the PVT is a key connection point for coordinating responses to positive and negative emotional states that lead to drug or food seeking and a return to uncontrolled intake. Most PVT neurons connect to the NAc, forming activating connections with NAc neurons. There is a clear layout where the front part of the PVT (aPVT) mainly connects to the top part of the NAc shell (NAcSh), while the back part of the PVT (pPVT) mainly connects to the bottom-middle of the NAcSh and the NAc core (NAcC). This anatomical difference is important, though not fully understood, because the top and bottom parts of the NAcSh can have opposite effects on motivated behavior. The pPVT also strongly connects to the central nucleus of the amygdala (CeA), where it helps control fear responses. This section will focus on parts of the hypothalamic–thalamic–striatal brain pathways that control reward-seeking.

The PVT sends signals that branch out widely. For example, it was found that half of PVT neurons connecting to the dorsolateral BNST and the CeA also connected to the NAcSh. This extensive branching of PVT connections suggests this region coordinates a complex network of cortical and extended amygdala regions important for motivated behavior, reward-seeking, and emotional value. So far, relatively little is known about how this branching of PVT connections translates into specific behaviors, creating a gap in our understanding.

Neural Circuit-Based Mechanisms for Abstinence-Induced Food-Seeking

Food cues cause strong activity in the brain's reward system. This section will discuss possible brain mechanisms behind the increase in food craving during abstinence. The main focus will be on circuits made up of connections between the PVT, LH, Amy, and NAc, as well as how specific changes to these connections affect behavior. Both the PVT and the NAc are well-positioned as major points where signals from the cortex, hypothalamus, and midbrain structures come together. Additionally, both PVT and NAc play key roles in controlling eating behaviors, motivation, reward, and learning.

While the role of the PVT in food-seeking after stopping or in the increase of craving has not been thoroughly studied, several studies clearly describe its role in adjusting behaviors motivated by food cues. For example, damage to the PVT increases motivational responses to cues that predict food in certain rats. These results suggest that the PVT plays a critical role in reducing the importance given to cues that predict a reward. In contrast, turning off the PVT with a specific drug increases cue-induced sucrose-seeking (but only when the expected reward is not given, a frustrating situation). Additionally, activating a specific receptor within the PVT leads to decreased cue-induced sucrose-seeking, while stimulating a specific glucose transporter in the PVT increases motivation to get sucrose in a learning task. Furthermore, using light to stimulate aPVT connections to the NAcSh decreases sucrose-seeking, while stopping aPVT connections with light increases sucrose-seeking (when the expected reward was not given). Also, increasing a specific brain chemical in the pPVT made NAcC neurons more responsive, and using light to stimulate the pPVT to NAcC pathway led to increased cue-induced sucrose-seeking. Both stimulating and stopping aPVT connections to the central nucleus of the amygdala (CeA) with light inhibits sucrose-seeking in rats. Moreover, connections between the PFC, PVT, and NAc are involved in forming links between cues and rewards, while LH-PVT-NAc connections are involved in consuming rewards and starting to eat. It has also been found that stopping certain connections to the NAc with light decreases sucrose-seeking. When all these findings are considered, the wide range of methods, brain areas, behavioral outcomes, and complex systems involved make it impossible to say the PVT has just one function in the context of seeking appealing food. However, the combination of findings suggests the PVT combines the emotional value of a stimulus, emotional and alertness states, and metabolic balance to either encourage or stop food-seeking. This has important implications for treating compulsive eating disorders.

Unfortunately, little is known about the role of the menstrual cycle or male/female sex hormones in adjusting these pathways. However, sex-specific effects of stopping a junk-food diet have been found in rats prone to obesity. In this case, a certain type of brain signaling in the NAc was stronger in males but not females. Given that sex hormones play an important role in food intake and motivational responses to food cues, it will be important to expand this research to studies of increasing craving for food rewards.

Neural Circuit-Based Mechanisms for Abstinence-Induced Opioid-Seeking

Studies using chemical and light methods on the PVT-NAc and NAc-LH pathways show they play key roles in various aspects of addictive behaviors, such as aversion linked to withdrawal, recalling past drug use, and returning to opioid-seeking behavior. Specific PVT neurons send signals to different types of NAc neurons. However, repeated opioid use strengthens these connections specifically at one type of NAc neuron. These strengthened NAc neurons then block signals from other NAc neurons, which in turn leads to opioid relapse. Directly activating a specific type of NAc neuron prevents relapse. The role of the PVT in addiction is an active area of research. The PVT has connections—some going both ways—with multiple brain regions involved in reward and aversion, and many studies link these pathways to addictive behaviors. LH neurons involving orexin play a major role in motivated behavior, including seeking cocaine and opioids in animal models of addictive behavior. Studies combining opioid self-administration and relapse models, as well as behavioral economics with general blocking of orexin receptors, have shown that the orexin system controls motivation towards seeking drugs. However, the specific role of different orexin pathways in controlling motivation towards opioid-seeking still needs to be explored. Studies on natural rewards and LH–PVT connections support this idea. Additionally, connections to the NAc from the ventro-medial prefrontal cortex (vmPFC) control opioid-seeking. Heroin relapse triggered by an environment is controlled by the interaction of vmPFC signals and specific dopamine signals.

Conclusions

This review summarizes a relatively new collection of studies suggesting that brain pathways involving the hypothalamus, thalamus, and striatum play a role in both opioid- and appealing food-seeking during abstinence—a strong animal model of relapse. Specifically, connections among the LH, PVT, NAc, and Amy work together to combine internal and external signals related to emotional state, alertness, energy balance, and motivation to drive behaviors linked to relapse. The similarities between opioid- and food-seeking behavior and their brain mechanisms are striking and often differ from what is seen with stimulant drugs.

Few studies have directly compared the behaviors and brain mechanisms of increasing craving in both males and females. Overall, the studies that have been reported find similar behavioral responses: both males and females show an increase in reward-seeking when they stop using. This is crucial information, as the development of effective treatments depends on how both males and females would respond. It also emphasizes the importance of looking deeper to understand the molecular, cellular, and circuit-based mechanisms for behavior. As discussed, sex hormones regulate reward-seeking and reward-based behavior, and since estrogen and testosterone are clearly different hormones, it is likely that different mechanisms underlie the behavioral responses. This is a promising area for new discoveries.

Through this review, the goal was to describe what is currently known and then to identify some key gaps in knowledge that researchers can address in future animal and human studies. For example, one gap is identifying the brain circuits and chemical messengers that link emotional state to motivated behavior. Specifically, does cue-triggered reward-seeking during abstinence arise from positive or negative mood states, and which brain region(s) within the hypothalamus-thalamus-striatum network are necessary? Another gap in knowledge is a detailed understanding of how neurotransmitters (like glutamate, GABA) and neuromodulators (like dopamine, orexin, dynorphin) interact within this network to affect specific aspects of motivated behavior related to opioid- and appealing food-seeking. There are many other questions, but a final gap in knowledge highlighted in this review is a general lack of understanding of the impact of a person's biological sex on how this fundamental brain circuit functions.

Abbreviations:

• Amy: amygdala

• BLA: basolateral amygdala

• BOLD: brain blood flow imaging signals

• CeA: central amygdala

• GLUT2: glucose transporter 2

• LH: lateral hypothalamus

• MSNs: medium spiny neurons

• NAc: nucleus accumbens

• NacC: NAc core

• NAcSh: NAc shell

• PVT: paraventricular nucleus of the thalamus

• aPVT: anterior PVT

• pPVT: posterior PVT

• PFC: prefrontal cortex

• OUD: opioid use disorder

• SUD: substance use disorder

• vmPFC: ventro medial PFC

Main Ideas

When people and animals stop using drugs or eating certain foods, they often strongly desire them again. This strong desire can lead them back to using or eating those things. It seems that a specific brain system helps control this strong desire. Also, whether someone is male or female can change how this strong desire grows over time, depending on what they are trying to stop using and how they stop.

Introduction

A strong, uncontrollable urge to use drugs or eat certain foods can hurt a person's health, mind, and relationships. This is known as substance use disorder or binge eating. These problems are like long-lasting brain illnesses that often come back. A big challenge in treating these issues is that many people go back to old habits after trying to stop. Many things can cause this, both from inside and outside a person. These things often lead to a strong desire, called craving, for the drug or food. This craving usually makes people go back to using or eating. Scientists are still trying to fully understand how craving works in the brain. Studies show that craving can be caused by being around the drug or food again, seeing things linked to it, or feeling stressed.

This paper looks at how stopping opioids or tasty foods affects the urge to seek them out. "Abstinence" means different things here. It can mean going through withdrawal when the drug or food is not available, which makes cravings stronger. Abstinence can also last for a short time, like a day, or a very long time, like months or years. A tricky part of addiction is that the craving can grow stronger the longer someone stays away from the drug or food. Scientists call this "incubation of craving." It means that over time, seeing something linked to the drug or food makes the desire to get it even stronger. This has been seen in both rats and people who stop using opioids or eating certain foods.

Many other papers have explored why people return to drugs or food. This paper, however, focuses on the urge to seek opioids and tasty foods after stopping. The ways these urges work can be similar and different from urges for other drugs. Recent studies have looked at a wider network of brain parts, not just the usual ones. This network includes the lateral hypothalamus (LH), the paraventricular nucleus of the thalamus (PVT), and the nucleus accumbens (NAc). These brain areas work together and play a key role in making someone seek opioids and foods high in fat or sugar after stopping. These brain parts are important for how awake someone feels, how the body uses energy, and how it handles rewards or things a person dislikes. Looking at this wider brain network can help answer many questions and guide future research.

Studies show that being male or female changes how strong and addictive behaviors appear, including how cravings grow and how often people go back to old habits. Since returning to old habits is a main reason why treatments fail, it is very important to understand how being male or female affects this process. So, another goal of this paper is to share what is known about these differences between males and females.

Clinical Relevance of Abstinence-Induced Reward-Seeking

People can learn to link drugs or tasty foods with things they see or hear around them. For example, seeing a certain place can make someone think of a drug. In people, these signals for drugs or food create a strong desire or urge to use them again. These urges can make people go back to using drugs or overeating, which is a big problem for treatments aimed at stopping drug use or losing weight. This is especially true because the desire for both drugs and food gets stronger when someone tries to stop.

Abstinence-Induced Food-Seeking in Humans

Seeing, smelling, or tasting food can signal that it is time to eat, which might cause someone to look for food. These signals can make someone eat too much. For people trying to limit what they eat, seeing or smelling food can cause a strong urge to eat a lot, known as binging. Also, people on a diet who are around high-fat foods are more likely to go back to unhealthy eating. So, both stopping eating and being around food signals make cravings stronger and lead to more food seeking and eating. Not eating enough also makes the body react more strongly to food signals.

Eating too much is the main cause of being very overweight. Studies of people show that those who are overweight have stronger brain activity when they see food signals, compared to people with a healthy weight. This activity is strongest in certain brain areas like the VTA, PFC, Amy, and NAc. Other studies show that parts of the brain called thalamic nuclei become active when someone craves a reward or sees a signal for it. However, current brain scans cannot clearly tell the difference between the PVT and other thalamic nuclei. Still, these thalamic nuclei work closely with the brain's reward system and are thought to be very important in how animals eat and look for food.

There are clear differences in how men and women control how much they eat and their body weight. These differences come from things like how sensitive their bodies are to certain hormones, other hormones related to being male or female, and genes that affect energy use, where fat is stored, and appetite. So, it is important to know how these things work differently in men and women to understand why some people become very overweight or eat too much. For women, how much they eat changes during their monthly cycle. This is because of natural changes in hormones. For example, changes in certain hormones can make women eat less or more at different times. Some studies also show that hormones in women are important for how certain brain areas react to food signals. While both very overweight men and women react more to high-calorie food when compared to people of a healthy weight, women often react even more. Women also show more brain activity than men in certain brain areas when food signals are shown.

Abstinence-Induced Opioid-Seeking in Humans

Opioid use disorder means someone uses opioids in a harmful way, whether it is illegal drugs like heroin or prescription drugs like oxycodone. Recently, more people with opioid use disorder are using both prescription opioids and heroin. It has been hard to prove directly that craving for opioids gets stronger when people stop using them. But more studies are looking at stress, feelings, and impulsive actions, and how these change when someone stops using opioids. For example, people who stop using heroin get better at controlling their reactions over time, but this can be undone by seeing things related to the drug. This can make them more likely to start using again and makes treatment harder. A stronger craving for opioids is linked to more activity in certain parts of the brain. The most common things that cause craving are seeing things or being in places linked to the drug, or feeling stressed. For people who depend on opioids, often the strongest cause for seeking the drug is being around the drug itself.

Differences exist between men and women in how opioid use disorder affects them. Women often say they have more problems in their daily lives and are more likely to use opioids to deal with bad feelings or pain, compared to men. Women also report much stronger cravings. These differences were not linked to how much medicine they took or how sensitive they were to the disorder before treatment. These different behaviors include how sensitive people are to the good feelings from opioids, to bad feelings, and to stress hormones in the brain areas that handle bad feelings when stopping opioids.

Preclinical Models of Abstinence-Induced Reward-Seeking

This part looks at how scientists study going back to old habits after stopping drugs or food. This process has three steps: training, stopping, and testing for going back. In training, animals learn to give themselves drugs or tasty foods like high-fat or sweet foods over many days. During each training session, if the animal presses a lever, it gets a signal (like a sound or light) and a reward. After training, the rats stop getting the reward. This can be either forced, meaning they are kept away from the reward, or voluntary. Stopping the reward often leads to more seeking behavior. Most studies use forced stopping, where animals stay in their cages and cannot get the reward. For the "going back" test, rats are put back in the same room, and pressing the lever brings back the signal, but no reward. Depending on how long they stopped, what the reward was, and if the animal is male or female, the signals make them seek the reward more strongly than they did early on. This growing strength of drug or food seeking is called "incubation of craving."

Abstinence-Induced Sucrose- and High-Fat Diet-Seeking in Rodent Models

Scientists have seen "incubation of food craving" for things like sugar, high-fat foods, and fake sugar in animals like rats. Most studies on food craving use a method similar to studies on drug craving. For example, rats are trained to give themselves tasty food, and a signal is linked to the food. Then, animals are forced to stop eating those foods for different numbers of days. When testing for food craving, the rats are allowed to use the lever that used to give them food for a few hours before the food signals are brought back. In general, the longer the animals have stopped eating the food, the more they react to the lever when the signals return. This shows that the craving has grown, or "incubated." Studies have shown that after 15, 21, or 30 days of stopping sugar, rats look for sugar more and more when they see the signals. Cravings for high-fat and high-sugar diets also grow over time. For example, after 30 days of stopping high-fat foods, rats sought out rewards more when they saw related signals. However, rats did not show more craving for chocolate pellets. Even though fake sugar has no calories, it is very sweet, and signals linked to it can make rats seek it out after 30 days of stopping.

So far, only one study has directly looked at how sugar craving grows differently in male and female rats. Both male and female rats showed increased sugar craving, but the study did not find clear differences between the sexes in how much they sought sugar after stopping for various lengths of time. Other studies have looked at how female hormones affect food seeking caused by stress, but no effects were seen. Still, this does not mean that female hormones and the female cycle do not play a role in how cravings grow, as this has been seen in humans and for other rewards. In addition, it has been shown that female hormones and the female cycle affect how female rats respond to food signals when no food is given.

It is important that hormones from either females or males can cause different effects on how they seek rewards. For instance, when male rats that are likely to become overweight stop eating junk food, a chemical signal in a brain area called the NAcC became stronger in males, but not in females. Also, male hormones have been shown to affect the brain's reward system. Generally, female rats are more impulsive when choosing food rewards than males. Male hormones help reduce this impulsive choice in male rats when they are looking for food. Because of this, studies that look at how both male and female hormones affect seeking rewards, both generally and when someone is trying to stop, are very important.

Abstinence-Induced Opioid-Seeking in Rodent Models

Scientists have also found that opioid craving grows stronger in animal studies. This involves either forcing animals to stop using drugs or letting them stop on their own. Simply put, forced stopping means taking away the animal's way to get the drug. How long an animal stops using opioids changes how much they seek the drug later. The urge to seek opioids usually peaks between 6 to 25 days after stopping. This peak depends on the type of opioid and whether signals, places, or stress cause the animal to start seeking again.

In addition, there have been several recent studies examining how opioid craving grows and how often male and female animals go back to using after stopping on their own. These studies found no differences between males and females in their behavior. One study found that cravings grew in both sexes only after they were forced to stop, not when they stopped on their own. These results suggest that how male and female animals show growing opioid cravings is similar, even if the brain's inner workings are different. This is important for understanding the behavior and for finding ways to reduce cravings. Another study showed that giving female rats a hormone called estradiol helped them stop seeking heroin, and a mix of estradiol and another hormone, progesterone, helped even more. While these findings suggest that hormones play a role in seeking opioids, more study is needed to know if the female cycle affects this. It is also likely that male hormones play a part in growing cravings. So, more research is needed on how hormones affect opioid seeking.

Neural Circuit-Based Mechanisms for Abstinence-Induced Reward-Seeking

Many parts of the brain help with how people feel rewards, how motivated they are, and how they seek rewards. One common brain pathway studied is the mesocorticostriatal pathway. But recent studies have focused on how other brain areas, like the PVT and LH, work together with the NAc. The LH has become known as a very important brain area for seeking rewards. The LH sends signals throughout the brain that are involved in how awake someone feels and behaviors linked to rewards. More and more evidence shows that these LH signals play a key role in making someone go back to using drugs or seeking natural rewards.

The PVT is part of a brain network that brings together information about motivation, rewards, and how the body balances energy. The PVT mainly sends signals to the NAc, amygdala (Amy), and another area called the BNST. These areas have been studied a lot for their role in motivated behavior, rewards, dislikes, and fear. So, the PVT is like a central hub that helps control how the brain reacts to good or bad feelings, which can lead to seeking drugs or food and losing control over eating. Most PVT cells send signals to the NAc, making strong connections. Different parts of the PVT send signals to specific parts of the NAc. This matters because different parts of the NAc can have opposite effects on motivated behavior. The back part of the PVT also sends strong signals to the central amygdala, which helps control fear responses. This section will focus on how these brain signals help control seeking rewards.

Figure 1. Brain Connections for Food and Opioid Seeking.

This figure shows the brain pathways involved in seeking food and opioids.

When studying the PVT and what it does, it is important to know that its signals spread out widely. For example, half of the PVT cells that send signals to the BNST and CeA also send signals to the NAcSh. This wide spread of PVT signals suggests that this brain area helps to manage a complex network of other brain regions important for motivated behavior, seeking rewards, and feelings. So far, scientists do not fully understand how this wide spreading of PVT signals changes behavior, which means more research is needed.

Neural Circuit-Based Mechanisms for Abstinence-Induced Food-Seeking

Signals related to food cause strong activity in the brain's reward system. This section will talk about how different brain pathways might cause food cravings to grow. The main focus is on the connections between the PVT, LH, Amy, and NAc, and how changing these specific connections affects behavior. The PVT and NAc are important meeting points between different brain parts. Both the PVT and NAc play key roles in controlling how people eat, how motivated they are, how they feel rewards, and how they learn.

TABLE 1: How Specific Brain Connections Affect Food-Seeking Behaviors.

This table lists different brain connections and their roles in food-seeking behavior.

While how the PVT affects food seeking after stopping, or how cravings grow, has not been fully studied, many papers show its role in motivated food behaviors. For example, damaging the PVT in rats makes them more motivated by food signals. This suggests the PVT helps reduce how important food signals seem. On the other hand, turning off the PVT with a certain drug makes rats seek sugar more when they expect a reward but do not get it. Also, activating a specific protein in the PVT leads to less sugar seeking, while stimulating another protein in a PVT-NAc pathway makes rats more motivated to get sugar. Sending light signals to parts of the PVT that connect to the NAcSh can either decrease or increase sugar seeking, depending on how the light is used. Stimulating signals from the back part of the PVT to the NAcC also leads to more sugar seeking. Activating or turning off PVT connections to another brain area, the CeA, stops sugar seeking in rats. Other connections are involved in linking signals to rewards, or in eating and starting to eat. Also, turning off certain connections from the BLA to the NAc reduces sugar seeking. All these studies use different ways to look at many brain parts and their effects, making it hard to say the PVT does just one thing for food seeking. But overall, these findings suggest the PVT brings together information about how good something is, feelings, how awake someone feels, and body energy to either start or stop food seeking. This is very important for treating problems where people cannot stop eating.

Sadly, not much is known about how the female cycle or male and female hormones affect these brain pathways. However, when male rats that are likely to become overweight stop eating junk food, changes were seen in a brain area called the NAcC in males, but not in females. Since hormones play a big role in how much people eat and how they react to food signals, it will be important to study how they affect the growth of cravings for food rewards.

Neural Circuit-Based Mechanisms for Abstinence-Induced Opioid-Seeking

Studies using special methods to control brain cells show that connections between the PVT and NAc, and the NAc and LH, are very important in behaviors related to addiction. These include disliking withdrawal, remembering drug use, and going back to seeking opioids. Brain cells in the PVT send signals to specific cells in the NAcSh. But using opioids repeatedly makes these connections stronger for certain NAcSh cells. These stronger connections can then make other NAcSh cells less active, which pushes someone towards going back to opioids. Directly activating certain NAcSh-LH brain cells can stop this return to using opioids. Scientists are actively studying how the PVT affects addiction. The PVT is connected to many brain areas linked to rewards and dislikes, and many studies show these pathways are involved in addictive behaviors. Special brain cells in the LH play a big role in motivated behavior, including seeking cocaine and opioids in animal models of addiction. Studies have shown that a system called the orexinergic system controls the drive to seek drugs. However, how specific orexinergic cells cause the drive to seek opioids still needs more study. Connections between the vmPFC and NAc also play a part in seeking opioids. Going back to heroin use, caused by being in a certain place, happens through the interaction of vmPFC signals and another brain signal.

Conclusions

This paper has brought together new studies that suggest a brain network, including the hypothalamus, thalamus, and striatum, is involved in seeking both opioids and tasty foods after stopping. This is a good way to study how people go back to old habits. Specifically, the connections between the LH, PVT, NAc, and AMY bring together inner and outer signals about feelings, how awake someone feels, how the body uses energy, and motivation. These signals then drive behaviors that lead to going back to old habits. It is surprising how similar the behaviors and brain processes are for seeking opioids and food, and this is often different from what is seen with other stimulant drugs.

Not many studies have directly compared how cravings grow and how the brain works for both males and females. But most studies so far show similar behaviors: both males and females seek rewards more after stopping. This is very important because effective treatments need to work for everyone. It also shows why it is important to look deeper into the small parts of the brain and how they work together for behavior. As discussed, hormones in males and females affect seeking rewards. Since male and female hormones are different, it is likely that different things happen in the brain to cause these behaviors. This is a promising area for new discoveries.

In this paper, the aim was to explain what is known and to point out what scientists still need to learn for future studies. For instance, one thing not fully understood is how feelings connect to motivated behavior. Specifically, when someone stops drugs or food, does seeing a signal for it cause them to seek rewards because they feel good or bad? And which parts of the brain network are needed for this? Another thing not fully understood is how brain chemicals work together in this network to affect seeking opioids and tasty foods. There are many other questions, but one last thing not well understood is how being male or female affects this basic brain network.

TABLE 2: How Specific Brain Connections Affect Opioid-Seeking Behaviors.

This table lists different brain connections and their roles in opioid-seeking behavior.