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Objective: The behavioral and diagnostic heterogeneity within human opioid use disorder (OUD) diagnosis is not readily captured in current animal models, limiting translational relevance of the mechanistic research that is conducted in experimental animals. We hypothesize that a non-linear clustering of OUD-like behavioral traits will capture population heterogeneity and yield subpopulations of OUD vulnerable rats with distinct behavioral and neurocircuit profiles. Methods: Over 900 male and female heterogeneous stock rats, a line capturing genetic and behavioral heterogeneity present in humans, were assessed for several measures of heroin use and rewarded and non-rewarded seeking behaviors. Using a non-linear stochastic block model clustering analysis, rats were assigned to OUD vulnerable, intermediate and resilient clusters. Additional behavioral tests and circuit analyses using c-fos protein activation were conducted on the vulnerable and resilient subpopulations. Results: OUD vulnerable rats exhibited greater heroin taking and seeking behaviors relative to those in the intermediate and resilient clusters. Akin to human OUD diagnosis, further vulnerable rat sub-clustering revealed subpopulations with different combinations of behavioral traits, including sex differences. Lastly, heroin cue-induced neuronal patterns of circuit activation differed between resilient and vulnerable phenotypes. Behavioral sex differences were recapitulated in patterns of circuitry activation, including males preferentially engaging extended amygdala stress circuitry, and females cortico-striatal drug cue-seeking circuitry. Conclusion: Using a non-linear clustering approach in rats, we captured behavioral diagnostic heterogeneity reflective of human OUD diagnosis. OUD vulnerability and resiliency were associated with distinct neuronal activation patterns, posing this approach as a translational tool in assessing neurobiological mechanisms underpinning OUD.
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The increased prevalence of opioid use disorder (OUD) makes it imperative to disentangle the biological mechanisms contributing to individual differences in OUD vulnerability. OUD shows strong heritability, however genetic variants contributing toward vulnerability remain poorly defined. We performed a genome-wide association study using over 850 male and female heterogeneous stock (HS) rats to identify genes underlying behaviors associated with OUD such as nociception, as well as heroin-taking, extinction and seeking behaviors. By using an animal model of OUD, we were able to identify genetic variants associated with distinct OUD behaviors while maintaining a uniform environment, an experimental design not easily achieved in humans. Furthermore, we used a novel non-linear network-based clustering approach to characterize rats based on OUD vulnerability to assess genetic variants associated with OUD susceptibility. Our findings confirm the heritability of several OUD-like behaviors, including OUD susceptibility. Additionally, several genetic variants associated with nociceptive threshold prior to heroin experience, heroin consumption, escalation of intake, and motivation to obtain heroin were identified. Tom1 , a microglial component, was implicated for nociception. Several genes involved in dopaminergic signaling, neuroplasticity and substance use disorders, including Brwd1 , Pcp4, Phb1l2 and Mmp15 were implicated for the heroin traits. Additionally, an OUD vulnerable phenotype was associated with genetic variants for consumption and break point, suggesting a specific genetic contribution for OUD-like traits contributing to vulnerability. Together, these findings identify novel genetic markers related to the susceptibility to OUD-relevant behaviors in HS rats.
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Introduction: It is well known that chronic opioid use disorder is associated with alterations in gastrointestinal (GI) function that include constipation, reduced motility, and increased bacterial translocation due to compromised gut barrier function. These signs of disrupted GI function can be associated with alterations in the gut microbiome. However, it is not known if long-access opioid self-administration has effects on the gut microbiome. Methods: We used 16S rRNA gene sequencing to investigate the gut microbiome in three independent cohorts (N=40 for each) of NIH heterogeneous stock rats before onset of long-access heroin self-administration (i.e., naïve status), at the end of a 15-day period of self-administration, and after post-extinction reinstatement. Measures of microbial α- and ß-diversity were evaluated for all phases. High-dimensional class comparisons were carried out with MaAsLin2. PICRUSt2 was used for predicting functional pathways impacted by heroin based on marker gene sequences. Results: Community α-diversity was not altered by heroin at any of the three phases by comparison to saline-yoked controls. Analyses of ß-diversity showed that the heroin and saline-yoked groups clustered significantly apart from each other using the Bray-Curtis (community structure) index. Heroin caused significant alterations at the ASV level at the self-administration and extinction phases. At the phylum level, the relative abundance of Firmicutes was increased at the self-administration phase. Deferribacteres was decreased in heroin whereas Patescibacteria was increased in heroin at the extinction phase. Potential biomarkers for heroin emerged from the MaAsLin2 analysis. Bacterial metabolomic pathways relating to degradation of carboxylic acids, nucleotides, nucleosides, carbohydrates, and glycogen were increased by heroin while pathways relating to biosynthesis of vitamins, propionic acid, fatty acids, and lipids were decreased. Discussion: These findings support the view that long access heroin self-administration significantly alters the structure of the gut microbiome by comparison to saline-yoked controls. Inferred metabolic pathway alterations suggest the development of a microbial imbalance favoring gut inflammation and energy expenditure. Potential microbial biomarkers and related functional pathways likely invoked by heroin self-administration could be targets for therapeutic intervention.
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RATIONALE: The ongoing rise in opioid use disorder (OUD) has made it imperative to better model the individual variation within the human population that contributes to OUD vulnerability. Using animal models that capture such variation can be a useful tool. Individual variation in novelty-induced locomotion is predictive of substance use disorder (SUD) propensity. In this model, rats are characterized as high-responders (HR) or low-responders (LR) using a median split based on distance travelled during a locomotor test, and HR rats are generally found to exhibit a more SUD vulnerable behavioral phenotype. OBJECTIVES: The HR/LR model has commonly been used to assess behaviors in male rats using psychostimulants, with limited knowledge of the predictive efficacy of this model in females or the use of an opioid as the reward. In the current study, we assessed several behaviors across the different phases of drug addiction (heroin taking, refraining, and seeking) in over 500 male and female heterogeneous stock rats run at two geographically separate locations. Rats were characterized as HRs or LRs within each sex for analysis. RESULTS: Overall, females exhibit a more OUD vulnerable phenotype relative to males. Additionally, the HR/LR model was predictive of OUD-like behaviors in male, but not female rats. Furthermore, phenotypes did not differ in anxiety-related behaviors, reacquisition of heroin-taking, or punished heroin-taking behavior in either sex. CONCLUSIONS: These results emphasize the importance of assessing females in models of individual variation in SUD and highlight limitations in using the HR/LR model to assess OUD propensity.
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Conducta Exploratoria , Dependencia de Heroína , Humanos , Femenino , Ratas , Animales , Masculino , Analgésicos Opioides/farmacología , Actividad Motora , Heroína/farmacologíaRESUMEN
RATIONALE: Relapse often occurs when individuals are exposed to stimuli or cues previously associated with the drug-taking experience. The ability of drug cues to trigger relapse is believed to be a consequence of incentive salience attribution, a process by which the incentive value of reward is transferred to the reward-paired cue. Sign-tracker (ST) rats that attribute enhanced incentive value to reward cues are more prone to relapse compared to goal-tracker (GT) rats that primarily attribute predictive value to such cues. OBJECTIVES: The neurobiological mechanisms underlying this individual variation in relapse propensity remains largely unexplored. The paraventricular nucleus of the thalamus (PVT) has been identified as a critical node in the regulation of cue-elicited behaviors in STs and GTs, including cue-induced reinstatement of drug-seeking behavior. Here we used a chemogenetic approach to assess whether "top-down" cortical input from the prelimbic cortex (PrL) to the PVT plays a role in mediating individual differences in relapse propensity. RESULTS: Chemogenetic inhibition of the PrL-PVT pathway selectively decreased cue-induced reinstatement of drug-seeking behavior in STs, without affecting behavior in GTs. In contrast, cocaine-primed drug-seeking behavior was not affected in either phenotype. Furthermore, when rats were characterized based on a different behavioral phenotype-locomotor response to novelty-inhibition of the PrL-PVT pathway had no effect on either cue- or drug-induced reinstatement. CONCLUSIONS: These results highlight an important role for the PrL-PVT pathway in vulnerability to relapse that is consequent to individual differences in the propensity to attribute incentive salience to discrete reward cues.
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Señales (Psicología) , Comportamiento de Búsqueda de Drogas , Animales , Masculino , Motivación , Ratas , Ratas Sprague-Dawley , Recurrencia , Recompensa , TálamoRESUMEN
Opioid use disorder is a psychological condition that affects over 200,000 people per year in the U.S., causing the Centers for Disease Control and Prevention to label the crisis as a rapidly spreading public health epidemic. The behavioral relationship between opioid exposure and development of opioid use disorder (OUD) varies greatly between individuals, implying existence of sup-populations with varying degrees of opioid vulnerability. However, effective pre-clinical identification of these sub-populations remains challenging due to the complex multivariate measurements employed in animal models of OUD. In this study, we propose a novel non-linear network-based data analysis workflow that employs seven behavioral traits to identify opioid use sub-populations and assesses contributions of behavioral variables to opioid vulnerability and resiliency. Through this analysis workflow we determined how behavioral variables across heroin taking, refraining and seeking interact with one another to identify potentially heroin resilient and vulnerable behavioral sub-populations. Data were collected from over 400 heterogeneous stock rats in two geographically distinct locations. Rats underwent heroin self-administration training, followed by a progressive ratio and heroin-primed reinstatement test. Next, rats underwent extinction training and a cue-induced reinstatement test. To enter the analysis workflow, we integrated data from different cohorts of rats and removed possible batch effects. We then constructed a rat-rat similarity network based on their behavioral patterns and implemented community detection on this similarity network using a Bayesian degree-corrected stochastic block model to uncover sub-populations of rats with differing levels of opioid vulnerability. We identified three statistically distinct clusters corresponding to distinct behavioral sub-populations, vulnerable, resilient and intermediate for heroin use, refraining and seeking. We implement this analysis workflow as an open source R package, named mlsbm.
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RATIONALE: Prior research suggests that the neural pathway from the lateral hypothalamic area (LHA) to the paraventricular nucleus of the thalamus (PVT) mediates the attribution of incentive salience to Pavlovian reward cues. However, a causal role for the LHA and the neurotransmitters involved have not been demonstrated in this regard. OBJECTIVES: To examine (1) the role of LHA in the acquisition of Pavlovian conditioned approach (PavCA) behaviors, and (2) the role of PVT orexin 1 receptors (OX1r) and orexin 2 receptors (OX2r) in the expression of PavCA behaviors and conditioned reinforcement. METHODS: Rats received excitotoxic lesions of the LHA prior to Pavlovian training. A separate cohort of rats characterized as sign-trackers (STs) or goal-trackers (GTs) received the OX1r antagonist SB-334867, or the OX2r antagonist TCS-OX2-29, into the PVT, to assess their effects on the expression of PavCA behavior and on the conditioned reinforcing properties of a Pavlovian reward cue. RESULTS: LHA lesions attenuated the development of sign-tracking behavior. Administration of either the OX1r or OX2r antagonist into the PVT reduced sign-tracking behavior in STs. Further, OX2r antagonism reduced the conditioned reinforcing properties of a Pavlovian reward cue in STs. CONCLUSIONS: The LHA is necessary for the development of sign-tracking behavior; and blockade of orexin signaling in the PVT attenuates the expression of sign-tracking behavior and the conditioned reinforcing properties of a Pavlovian reward cue. Together, these data suggest that LHA orexin inputs to the PVT are a key component of the circuitry that encodes the incentive motivational value of reward cues.
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Señales (Psicología) , Área Hipotalámica Lateral/fisiología , Núcleos Talámicos de la Línea Media/fisiología , Motivación/fisiología , Receptores de Orexina/fisiología , Recompensa , Animales , Benzoxazoles/administración & dosificación , Conducta de Elección/efectos de los fármacos , Conducta de Elección/fisiología , Condicionamiento Clásico/efectos de los fármacos , Condicionamiento Clásico/fisiología , Área Hipotalámica Lateral/efectos de los fármacos , Isoquinolinas/administración & dosificación , Masculino , Núcleos Talámicos de la Línea Media/efectos de los fármacos , Motivación/efectos de los fármacos , Naftiridinas/administración & dosificación , Antagonistas de los Receptores de Orexina/administración & dosificación , Piridinas/administración & dosificación , Ratas , Ratas Sprague-Dawley , Urea/administración & dosificación , Urea/análogos & derivadosRESUMEN
Drug addiction is a neuropsychiatric disorder with grave personal consequences that has an extraordinary global economic impact. Despite decades of research, the options available to treat addiction are often ineffective because our rudimentary understanding of drug-induced pathology in brain circuits and synaptic physiology inhibits the rational design of successful therapies. This understanding will arise first from animal models of addiction where experimentation at the level of circuits and molecular biology is possible. We will review the most common preclinical models of addictive behavior and discuss the advantages and disadvantages of each. This includes non-contingent models in which animals are passively exposed to rewarding substances, as well as widely used contingent models such as drug self-administration and relapse. For the latter, we elaborate on the different ways of mimicking craving and relapse, which include using acute stress, drug administration or exposure to cues and contexts previously paired with drug self-administration. We further describe paradigms where drug-taking is challenged by alternative rewards, such as appetitive foods or social interaction. In an attempt to better model the individual vulnerability to drug abuse that characterizes human addiction, the field has also established preclinical paradigms in which drug-induced behaviors are ranked by various criteria of drug use in the presence of negative consequences. Separation of more vulnerable animals according to these criteria, along with other innate predispositions including goal- or sign-tracking, sensation-seeking behavior or impulsivity, has established individual genetic susceptibilities to developing drug addiction and relapse vulnerability. We further examine current models of behavioral addictions such as gambling, a disorder included in the DSM-5, and exercise, mentioned in the DSM-5 but not included yet due to insufficient peer-reviewed evidence. Finally, after reviewing the face validity of the aforementioned models, we consider the most common standardized tests used by pharmaceutical companies to assess the addictive potential of a drug during clinical trials.
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Cues in the environment can elicit complex emotional states, and thereby maladaptive behavior, as a function of their ascribed value. Here we capture individual variation in the propensity to attribute motivational value to reward-cues using the sign-tracker/goal-tracker animal model. Goal-trackers attribute predictive value to reward-cues, and sign-trackers attribute both predictive and incentive value. Using chemogenetics and microdialysis, we show that, in sign-trackers, stimulation of the neuronal pathway from the prelimbic cortex (PrL) to the paraventricular nucleus of the thalamus (PVT) decreases the incentive value of a reward-cue. In contrast, in goal-trackers, inhibition of the PrL-PVT pathway increases both the incentive value and dopamine levels in the nucleus accumbens shell. The PrL-PVT pathway, therefore, exerts top-down control over the dopamine-dependent process of incentive salience attribution. These results highlight PrL-PVT pathway as a potential target for treating psychopathologies associated with the attribution of excessive incentive value to reward-cues, including addiction.
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Conducta Animal , Señales (Psicología) , Sistema Límbico/fisiología , Vías Nerviosas/fisiología , Animales , Motivación , Ratas , RecompensaRESUMEN
RATIONALE: The paraventricular nucleus of the thalamus (PVT) has been shown to mediate cue-motivated behaviors, such as sign- and goal-tracking, as well as reinstatement of drug-seeking behavior. However, the role of the PVT in mediating individual variation in cue-induced drug-seeking behavior remains unknown. OBJECTIVES: This study aimed to determine if inactivation of the PVT differentially mediates cue-induced drug-seeking behavior in sign-trackers and goal-trackers. METHODS: Rats were characterized as sign-trackers (STs) or goal-trackers (GTs) based on their Pavlovian conditioned approach behavior. Rats were then exposed to 15 days of cocaine self-administration, followed by a 2-week forced abstinence period and then extinction training. Rats then underwent tests for cue-induced reinstatement and general locomotor activity, prior to which they received an infusion of either saline (control) or baclofen/muscimol (B/M) to inactivate the PVT. RESULTS: Relative to control animals of the same phenotype, GTs show a robust increase in cue-induced drug-seeking behavior following PVT inactivation, whereas the behavior of STs was not affected. PVT inactivation did not affect locomotor activity in either phenotype. CONCLUSION: In GTs, the PVT appears to inhibit the expression of drug-seeking, presumably by attenuating the incentive value of the drug cue. Thus, inactivation of the PVT releases this inhibition in GTs, resulting in an increase in cue-induced drug-seeking behavior. PVT inactivation did not affect cue-induced drug-seeking behavior in STs, suggesting that the role of the PVT in encoding the incentive motivational value of drug cues differs between STs and GTs.