Environmental enrichment selectively counteracts brain metabolic activity during cocaine abstinence

Environmental enrichment selectively counteracts brain metabolic activity during cocaine abstinence | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article Environmental enrichment selectively counteracts brain metabolic activity during cocaine abstinence Marcello Solinas, Pauline Belujon, Virginie Lardeux, Emilie Dugast, and 6 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6984132/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Environmental enrichment (EE) is a promising strategy to promote recovery from addiction, but its neurobiological mechanisms remain poorly understood. In this study we used 18-fluorodeoxyglucose (FDG) microPET imaging to investigate how exposure to EE during abstinence affects brain neuroadaptations induced by voluntary intake of cocaine at different time of abstinence. After establishing escalated cocaine intake, male rats were housed in enriched or standard environments for four weeks and their brain metabolic activity was assessed after one and four weeks of abstinence. Cocaine self-administration produced widespread decreases in cortical metabolic activity, particularly in regions involved in executive function (orbitofrontal cortex, anterior cingulate), interoception (insula) and motivation (nucleus accumbens), while increasing activity in emotional circuits (ventral hippocampus) and the mesencephalon. EE selectively normalized these alterations restoring nucleus accumbens and orbitofrontal cortex activity. These findings reveal circuit-specific effects of environmental enrichment on cocaine-induced brain adaptations and suggest that effective addiction treatment requires both early interventions targeting reward circuits and sustained environmental stimulation to restore executive function, potentially reducing both immediate craving intensity and long-term relapse vulnerability. Health sciences/Diseases/Psychiatric disorders/Addiction Health sciences/Risk factors Biological sciences/Neuroscience/Diseases of the nervous system/Addiction Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Introduction One of the hallmarks of addiction and one of its most troubling aspects is the long-lasting risk of relapse even after protracted periods of abstinence ( 1 – 3 ). Over the past decades, neuroimaging studies in humans and mechanistic studies in animals have demonstrated that drugs of abuse produce persistent plastic changes in the brain that are thought to underlie the chronic nature of addiction ( 4 , 5 ). Based on clinical and brain-imaging data in humans, Goldstein and Volkow proposed the Impaired Response Inhibition and Salience Attribution (I-RISA) model that postulates that addiction results from dysregulation in the coordinated activity of different brain circuits ( 6 , 7 ). In this model, reduced activity in prefrontal and frontal cortical areas leads to deficits in executive function and response inhibition, while increased reactivity in limbic regions leads to heightened responsivity to drug-related cues and stress ( 6 , 7 ). In particular, studies in humans with cocaine addiction have shown hypoactivity in the anterior cingulate and orbitofrontal cortex associated with decision-making deficits ( 8 , 9 ), and hyperactivity in regions like the amygdala associated with increased stress reactivity ( 10 ). Animal studies have been essential in understanding the mechanisms underlying these brain changes ( 4 , 11 , 12 ). In the last decades, brain imaging studies have provided important insights into the changes induced by acute and chronic intake of drugs ( 13 – 20 ) and the neuroadaptations that persist after discontinuation of drug use which could be associated with incubation of craving and long term risks of relapse ( 21 – 26 ). In particular, using microPET imaging in rats, we have shown that extended access to cocaine self-administration produces persistent changes in brain metabolic activity that strikingly parallel those described in humans ( 24 ). Cocaine-exposed rats show decreased activity in cortical regions involved in executive function and decision-making, and increased activity in regions involved in motivation, memory, stress and emotional processing ( 24 ). Despite our growing understanding of cocaine-induced brain changes, developing effective treatments for addiction remains challenging. Among the novel strategies to help recovering from addiction, environmental enrichment (EE) has emerged as a promising therapeutic approach ( 27 – 30 ). EE involves housing animals in stimulating environments that provide opportunities for social interaction, physical activity, and cognitive engagement ( 29 ). When implemented during periods of drug abstinence, EE consistently reduces drug seeking and taking behaviors in animal models of addiction ( 29 , 31 ). The therapeutic benefits of EE likely stem from its ability to engage natural reward systems and promote adaptive neuroplasticity ( 29 ). While a few studies have investigated neurobiological mechanisms associated with the beneficial effects of EE on relapse ( 30 , 32 – 34 ), the mechanisms by which environmental enrichment produces its therapeutic effects on the brain and their dynamics remain poorly understood. In this study, we used FDG microPET imaging to investigate how exposure to environmental enrichment during abstinence affects cocaine-induced changes in basal brain metabolic activity. We first allowed male rats to self-administer cocaine under extended access conditions that lead to escalation of intake ( 35 ) and then housed them in either enriched or standard environments during four weeks of abstinence. By comparing brain activity patterns across groups and time points, we aimed to better understand how environmental enrichment influences the brain changes associated with cocaine addiction and abstinence. Materials and Methods Subjects and experimental design Adult (11–12 weeks of age) male Sprague-Dawley rats (Janvier, France), experimentally naive at the start of the study, were housed in a temperature- and humidity-controlled room and maintained on a 12-h light/dark cycle (light on at 7.00 AM). All experiments were conducted in accordance with European Union directives (2010/63/EU) for the care of laboratory animals approved by the local ethics committee (COMETHEA, CEEA Val de Loire, #2015033014555999). Housing Conditions On arrival, rats were housed two per cage for about 1 week before intrajugular catheterization surgery. Rats were anaesthetized using isoflurane (5% induction, 2.5% surgery) and implanted with catheters into the right jugular vein. Animals were allowed to recover for 5–7 days before the self-administration sessions began. After surgery, rats were housed individually during the entire period of active self-administration. At the end of the last self-administration session, rats were pseudo-randomly divided into two groups, one housed in standard environments (SE) and the other in EE conditions assuring that similar levels of drug self-administration were observed in the two groups. SE and EE conditions were the same as previously used ( 36 – 38 ). For SE, rats were housed in groups of three in cages sized 60 × 38 × 20 cm. For EE, rats were housed in groups of three in cages sized 80 × 50 × 100 cm. Each EE cage contained a house, a running wheel, three floors connected by ramps or tunnels and four toys that were changed once per week. Rats were housed in the animal facility in SE or EE housing conditions during a 4-week period of abstinence. Four experimental groups were obtained: NaiveSE, NaiveEE, CocSE and CocEE. The timeline of the experiment, including the different experimental groups, is illustrated in Fig. 1 A. Cocaine self-administration apparatus and procedure Catheter implantation . Rats were prepared for cocaine self-administration by surgical catheterization of the right jugular vein. Briefly, rats were anesthetized with isoflurane (5% induction, 2.5% maintenance) in O2 and administered with the nonsteroidal anti-inflammatory ketoprofen (2.5mg/kg, s.c.). Animals were placed on a heating pad from the induction of the anesthesia to the end of the surgery. A handmade silastic catheter was inserted into the jugular vein and the distal end was led to the back between the scapulae. Rats were allowed to recover for 7 days and flushed daily with 0.1ml sterile saline (0.9%), gentamicin (20mg/ml) and heparin (100 UI/mL) in sterile saline to help protect against infection and catheter occlusion. Apparatus. Experiments were conducted in MedAssociates operant-conditioning chambers, equipped with retractable levers as operanda, a cue-light above the active lever, a house light and controlled by MedAssociates interfaces and MED-PC IV software ( www.medassociates.com ). Cocaine self-administration procedure . Rats were allowed to self-administer cocaine (Cooper, France; 6g/L in saline solution 0.9%; 0.75mg/kg/infusion) for 6h/day for 25 sessions, using a Fixed Ratio 1 (FR1) schedule of reinforcement. A single press on the active lever resulted in one intravenous (i.v.) cocaine infusion with the concomitant activation of the light that remained on for 5s and then pulsed for 5s, followed by a 5s time-out. Inactive lever presses were recorded but did not produce any consequences. Naive control animals were age-matched rats that were house similarly to cocaine rats but did not undergo surgery and self-administration procedures. At the end of the last self-administration session, all rats were transferred from the animal facility at the University of Poitiers to the animal facility at University of Tours by an authorized transporter, placed in the SE or EE previously described and underwent abstinence for 4 weeks (EE: N = 8 and SE: N = 8). Metabolic imaging using FDG was performed in the same rats after 6–8 days (1 week) and 27–29 days (4 weeks) of abstinence. Naive rats of the same age and with analogous housing conditions but that did not undergo self-administration were used as controls (EE: N = 7 and SE: N = 8). Brain Imaging Local uptake of FDG reflects cerebral metabolic rates of glucose utilization and allows the investigation of regional brain metabolic status ( 39 ). Metabolic imaging using FDG was performed under basal conditions using the same procedure as previously described ( 24 ). Briefly, rats were habituated to the PET experimental procedures for 4 days before each scan and fasted overnight before each scan. The day of brain-imaging acquisition, awake rats were injected with FDG (18.5 MBq/100 g i.p.; Cyclopharma, Tours), and placed in the habituation cage for 45 min. This timing was chosen based on previous studies that used similar approaches to investigate brain activity associated with behavioral performance ( 40 ) activity before anesthesia. Then, they were anesthetized using isoflurane 4% (Baxter, Maurepas, France), placed on a heating pad (Minerve, Esternay, France) and centered in the field of view of the Explore VISTA-CT microPET camera (GE Healthcare, Velizy, France). CT-scan was performed for attenuation correction of PET images and a list-mode PET acquisition of 30 min started 60 min after FDG injection. After data reconstruction using a 2-D OSEM algorithm, all images were co-registered and normalized for tissue activity in the whole brain. Quantitative results were expressed as mean ± SD and were presented on Z-score maps (for more details, refer to ( 41 )). Analyses focused on brain areas known to be key nodes in addiction and that exhibited changes in FDG uptake during cocaine abstinence: the cingulate (Cg), orbitofrontal (OFC), prelimbic/infralimbic (PrL/IL), insular and motor cortices, in addition to the dorsal striatum (DStr), nucleus accumbens (NAc), substantia nigra/ventral tegmental area (SN/VTA), amygdala (Amyg), and hippocampus (Hipp) ( 24 , 42 ). Statistical analysis For self-administration, data were analyzed by two-way repeated measures ANOVA with time designated as a within-subject factor and future environment exposure (Coc SE or Coc EE) as a between-subject factor. For micro-PET data, a voxel-based analysis was used to assess the differences in cerebral FDG uptake between the averaged brains of cocaine vs control rats at each stage of abstinence for rats housed in EE vs SE. The regions of interest were derived from Schiffer’s templates ( 43 ) using PMOD v3.2 software (PMOD Technologies Ltd, Switzerland) and applied to Z-score and effect size maps to obtain the Z-score and d values in these areas ( 41 , 44 ). Statistical analyses focused on inter-group comparisons investigating differences in metabolic activity between (i) cocaine and naive rats housed in each environment at each time point, and (ii) EE vs SE in cocaine and naive rats at each time point using a two-tail unpaired Student t-test. Cohen d values were calculated to evaluate effect size of the differences. Notably, the effect of time of abstinence was not included as a factor in statistical analysis to avoid excessive comparisons which could inflate alpha errors. However, data in naive SE rats demonstrate that apart from a reduction in the activity in the dorsal striatum, brain activity was not significantly affected by testing at different times (Fig. S1 ), suggesting that time per se and repeated testing produce limited effects on brain metabolic activity. Differences were considered significant when p < 0.01 for signals of at least 50 contiguous voxels, with effect sizes showing Cohen d values of at least 0.80. Results Cocaine self-administration Prior to being assigned to different housing conditions, all rats showed similar patterns of cocaine self-administration (Fig. 1 B, C). Statistical analysis revealed no significant difference between future SE and future EE rats in the number of active lever presses (two-way ANOVA: time, F (24,360) = 5.81, p < 0.05 ; group SE/EE, F (1,14) = 0.1150, p = 0.7395 ; interaction, F (24,336) = 1.006, p = 0.4582) or in cocaine intake (two-way ANOVA: time, F (24,336) = 5.777,p < 0.05 ; group SE/EE, F (1,14) = 0.1147, p = 0.7399 ; interaction, F (24,336) = 0.7573, p = 0.7896). Effects of Environmental Enrichment on Brain Activity First, we investigated the effects of EE in naive rats (NaiveEE vs NaiveSE). One week of EE produced specific changes in brain metabolic activity (Supplementary Table 1 and Fig. S2A). Decreased activity was observed in the anterior cingulate cortex (ACC) (Cohen d = 1.14, p = 0.0149), insula (Ins) (Cohen d = 1.24, p = 0.0019), motor cortex (Mot) (Cohen d = 1.13, p = 0.0018), hippocampus (Hipp) (Cohen d = 1.11, p = 0.0104), and substantia nigra/ventral tegmental area (SN/VTA) (Cohen d = 1.22, p = 0.0002). Increased activity was found in prelimbic/infralimbic (PrL/IL) (Cohen d = 1.16, p = 0.0047), dorsomedial striatum (DMStr) (Cohen d = 1.21, p = 0.0011), dorsolateral striatum (DLStr) (Cohen d = 1.19, p = 0.0021), and posterior hippocampus (Hipp) (dorsal: Cohen d = 1.15, p = 0.0043; ventral: Cohen d = 1.11, p = 0.0067). After four weeks (Supplementary Table 1 and Fig. S2B), most changes normalized except for persistent alterations in motor cortex (increased, Cohen d = 1.09, p = 0.0081), insula (decreased, Cohen d = 1.10, p = 0.0220), ventral posterior hippocampus (increased, Cohen d = 1.16, p = 0.0011), and SN/VTA (decreased, Cohen d = 1.09, p = 0.0085). Effects of Cocaine on Brain Metabolic Activity During Abstinence Then, we verified that we could replicate our findings of cocaine-induced changes in metabolic activity ( 24 ). Consistent with our previous findings, in rats housed in standard environments (CocSE vs NaiveSE), one week of abstinence was characterized by widespread decreases in metabolic activity (Supplementary Table 2 and Fig. 2 A), particularly in cortical regions including the ACC (Cohen d = 1.14, p = 0.0018), motor cortex (Cohen d = 1.22, p < 0.0001), OFC (Cohen d = 1.26, p < 0.0001), PrL/IL (Cohen d = 1.12, p = 0.0011), and insula (Cohen d = 1.17, p = 0.0017). The NAc also showed decreased activity (Cohen d = 1.22, p = 0.002). In contrast, increased activity was observed in the dorsomedial striatum (Cohen d = 1.00, p = 0.0242), dorsal posterior hippocampus (Cohen d = 1.05, p = 0.0155), ventral posterior hippocampus (Cohen d = 1.05, p = 0.0038), and SN/VTA (Cohen d = 1.04, p = 0.0396). After four weeks of abstinence (Fig. 2 B), while some brain regions showed recovery (PrL/IL, NAc, and the motor cortex), persistent decreases were observed in the ACC (Cohen d = 1.00, p = 0.0362), OFC (Cohen d = 1.21, p = 0.0002), insula (Cohen d = 1.17, p = 0.0018), and dorsolateral striatum (Cohen d = 1.15, p = 0.012). The ventral posterior hippocampus (Cohen d = 1.14, p = 0.0008) and SN/VTA (Cohen d = 1.00, p = 0.0325) maintained increased activity. Finally, the anterior hippocampus showed switch from decrease to increase in metabolic activity (Cohen d = 0.98, p = 0.0074). Effects of Environmental Enrichment on Cocaine-Induced Brain Changes When cocaine-exposed rats were housed in enriched environments (CocEE vs NaiveEE), changes in metabolic activity were found only in a few regions (Supplementary Table 3 and Fig. 3 ). Indeed, after one week (Fig. 3 A), changes were limited to decreases in the ACC (Cohen d = 1.04, p = 0.0160), PrL/IL (Cohen d = 1.12, p = 0.0104), amygdala (Cohen d = 1.16, p = 0.0035), and anterior hippocampus (Cohen d = 1.13, p = 0.0071), with increases in NAc (Cohen d = 1.12, p = 0.0006) and posterior hippocampus (dorsal: Cohen d = 1.12, p = 0.0032; ventral: Cohen d = 1.13, p = 0.0138). After four weeks (Fig. 3 B), only significant increases in ventral posterior hippocampus (Cohen d = 1.11, p = 0.0176) and significant decreases in the ACC (Cohen d = 1.15, p = 0.0049) and in the motor cortex (Cohen d = 1.19, p = 0.0004). Direct comparison between cocaine-experienced rats in EE versus SE (CocEE vs CocSE; Supplementary table 4 and Fig. 4 ) revealed that after one week (Fig. 4 A), EE rats showed higher metabolic activity in the ACC (Cohen d = 1.13, p = 0.0042), OFC (Cohen d = 1.13, p = 0.0003), insula (Cohen d = 1.10, p = 0.0017), dorsolateral striatum (Cohen d = 1.06, p = 0.0092), and NAc (Cohen d = 1.11, p = 0.0005), lower activity in the amygdala (Cohen d = 1.15, p = 0.0052), anterior hippocampus (Cohen d = 1.05, p = 0.0288), and SN/VTA (Cohen d = 0.99, p = 0.0485). After four weeks (Fig. 4 B), differences persisted in the OFC (Cohen d = 1.13, p = 0.0028) and dorsolateral striatum (Cohen d = 1.10, p = 0.0039), while the ACC showed decreased activity (Cohen d = 0.97, p = 0.0257) and new differences emerged in ventral posterior hippocampus (Cohen d = 1.03, p = 0.0379). Focus on the metabolic responses to cocaine in the OFC and the NAc as a function of exposure to SE and EE Because the effects of EE appeared particularly robust in the OFC and NAc, we focused our final comparison on these two regions. To better highlight differences between groups, we plotted the normalized FDG uptake values for individual rats. After one week of abstinence CocSE rats exhibited significantly lower uptake in both the OFC (Fig. 5 A) and the NAc (Fig. 5 B) compared with NaiveSE rats (OFC, p < 0.0001; NAc, p < 0.001). After four weeks, this reduction persisted in the OFC (p < 0.001, Fig. 5 C) but was no longer apparent in the NAc (p = 0.0777, Fig. 5 D). Compared to CocSE, exposure to CocEE showed increased normalized FDG uptake in the OFC both at 1 and 4 weeks (Fig. 5 A, 5 C) and in the NAc 1 week (Fig. 5 C). In contrast, EE did not alter NAc uptake at 4 weeks (Fig. 5 D, p = 0.6737). Discussion Our findings reveal that cocaine self-administration produces persistent changes in brain metabolic activity that environmental enrichment can partially counteract. Specifically, cocaine decreased activity in cortical regions involved in executive function and the nucleus accumbens while increasing activity in emotional circuits. Environmental enrichment normalized activity in regions crucial for decision-making and reward processing, though some cocaine-induced changes remained unaffected. Cocaine addiction has been shown to be associated with long-lasting neuroadaptations in brain structures and functions that are believed to play a role in the persistent risks of relapse ( 5 , 45 ). These changes evolve throughout the course of abstinence: some emerge during the early stages and disappear after prolonged abstinence, others persist and some only manifest after several weeks of abstinence ( 19 , 46 – 48 ). Importantly, whereas the temporary changes may reflect tolerance/withdrawal processes, those persisting after long periods of abstinence could contribute to the phenomenon of incubation of craving ( 49 , 50 ) and may be more relevant to relapse after treatment and detoxification ( 19 , 46 – 48 ). Consistent with our previous work ( 24 ), the changes we observed in cocaine-exposed rats closely parallel those reported in human addiction studies and align with the I-RISA model ( 6 , 7 ). During early abstinence, we found decreased metabolic activity in the OFC, ACC, and insula, accompanied by disrupted activity in the NAc and SN/VTA. Many of these alterations persisted through prolonged abstinence, while new changes emerged in the dorsolateral striatum and hippocampus, consistent with the development of habits and enhanced emotional processing associated with addiction ( 4 , 5 , 51 ). In this study, environmental enrichment produced temporally distinct patterns of brain activity in naive versus cocaine-exposed animals. In naive rats, EE initially triggered widespread adaptations including decreased activity in the ACC, motor cortex, and insula, coupled with increased activity in striatal regions and hippocampus. Most of these changes normalized by four weeks, suggesting they represent acute adaptations to environmental stimulation rather than permanent circuit reorganization. These results are generally in agreement with previous studies using brain imaging approaches to investigate changes in brain activity following exposure to EE in mice ( 52 – 55 ) and in rats ( 56 ) and with the known molecular neuroadaptations induced by EE ( 57 , 58 ). In cocaine-exposed animals, however, EE demonstrated markedly different effects. In particular, EE selectively normalized cocaine-induced disruptions in specific circuits. Interesting effects were seen in the OFC and NAc, where cocaine-induced decreases in metabolic activity were prevented when animals were housed in enriched conditions. These regions are crucial for value-based decision making and reward processing ( 59 , 60 ), suggesting EE may restore the ability to properly evaluate natural rewards and make adaptive choices. Importantly, these therapeutic-like effects showed circuit specificity. While executive and reward circuits showed substantial recovery, other cocaine-induced alterations persisted despite enrichment. Most notably, the increased metabolic activity in the hippocampus remained largely unchanged, suggesting that EE may not affect all addiction-related neural adaptations equally. This selective pattern of effects provides insight into which aspects of addiction may be most amenable to environmental interventions and which may require additional therapeutic approaches. The effects of environmental enrichment reveal distinct phases of therapeutic action. In early abstinence, EE rapidly normalized activity in reward-related circuits, particularly the NAc and in regions involved in interoception such as the insular cortex. This rapid normalization likely reflects immediate effects on reward processing and stress response systems. The quick restoration of NAc function could explain the rapid effects of EE on drug seeking ( 36 ), possibly by enhancing the salience of natural rewards provided by enriched environment. In contrast, whereas changes in cortical regions like the OFC and in the and dorsolateral striatum were sustained from early to late abstinence, changes in the ACC evolved from increased metabolic activity at the beginning of abstinence to a decrease after four weeks of abstinence. This response suggests involvement of sustained neuroadaptive processes, including structural plasticity through dendritic remodeling and synaptogenesis, alterations in gene expression patterns, circuit-level reorganization, and possible restoration of normal neurotransmitter function ( 29 , 57 , 58 ). This temporal pattern suggests that successful treatment requires initial targeting of reward and stress systems while maintaining enrichment long enough for executive function recovery, accounting for different temporal windows of plasticity across circuits. Whereas most research has investigated the neurobiological consequences of drug taking, considerably less research has been dedicated to the investigation of the mechanisms associated with recovery from addiction. Our study addresses this knowledge gap by examining the neurobiological mechanisms underlying one specific pathway to recovery—environmental enrichment. Brain imaging studies in humans and rats have demonstrated that long-term abstinence from cocaine is associated with heightened prefrontal cortical activity compared to early abstinence periods ( 19 ), which corresponds with our findings of progressive normalization of prefrontal function in EE animals. Furthermore, as discussed by Engeln & Ahmed ( 61 ), recovery can involve both the reversal of drug-induced changes and the introduction of new compensatory mechanisms that compete with drug-related circuits. These time-dependent effects of EE align with these observations, demonstrating that EE not only counteracts some drug-induced adaptations but potentially creates additional circuit-level changes that support recovery. The neuroadaptive changes we observed during EE-induced recovery likely represent just one pathway among many that can lead to remission. While our study shows that EE can counteract drug-induced neuroadaptations, particularly in the nucleus accumbens and prefrontal regions, it is important to recognize that other interventions targeting different neural mechanisms may achieve similar outcomes. Several aspects should be considered when interpreting our results. First, our measures reflect metabolic activity under basal conditions, and therefore we cannot determine if and how environmental enrichment affects the reactivity of different brain regions to specific stimuli such as drug-related cues or stressors. Future studies using different behavioral challenges before imaging could provide insights into how environmental enrichment modifies brain responses to addiction-relevant triggers. Second, while PET imaging provides a valuable whole-brain perspective, its spatial resolution (approximately 1mm) limits our ability to distinguish small adjacent structures or subregions. For example, we analyzed the SN/VTA as a single region despite their distinct roles and could not differentiate between different amygdala nuclei or striatal subregions. More precise techniques like high-resolution fMRI or region-specific molecular analyses could complement our findings by providing finer anatomical detail. Third, to minimize the number of animals used, we performed repeated scans in the same rats. While this longitudinal approach has advantages for tracking changes over time, we cannot completely rule out potential effects of repeated manipulations such as fasting and anesthesia on brain activity. However, analysis of control rats at different time points showed limited effects of repeated testing (Fig. S1 ). Fourth, we focused only on male rats in this study. Significant differences exist in self-administration of drugs in male and female rats ( 62 , 63 ). On the other hand, the beneficial effects of EE on drug-related effects appears to be similar between males and females ( 64 , 65 ). Future studies will be needed to determine whether the neurobiological mechanisms underlying the anti-craving effects of EE are similar in male and female rats. Fifth, the control group used in this study was naive i.e. did not undergo surgery and was not exposed to self-administration cages. While we cannot completely rule out the possibility that these manipulations contributed to differences in metabolic activity in cocaine vs control rats, the contribution of these experimental differences is likely limited compared to exposure to cocaine. Moreover, it should be noted that the main comparison of this work was between rats exposed to SE or EE during abstinence after a similar history of cocaine self-administration. Finally, while environmental enrichment in laboratory settings is well-defined, translation to human conditions is complex. Our enriched environment combines social, physical, and cognitive stimulation in ways that may not directly parallel human conventional environmental interventions. However, an accumulating body of literature suggests that specific aspects of enrichment such as physical activity ( 66 , 67 ), cognitive training ( 68 , 69 ) and social support ( 70 ) can be effective in the treatment of addiction. Importantly, a clinical trial is in progress to investigate the effects of combination of several of these stimulations on relapse to severe alcohol use disorder ( 71 ). In conclusion, this study reveals that environmental enrichment can normalize or counteract cocaine-induced brain metabolic changes through distinct temporal mechanisms. Our findings provide a neurobiological framework explaining how environmental interventions can help recovery from addiction: immediate effects may reduce craving intensity by normalizing nucleus accumbens activity, while sustained environmental stimulation gradually repairs prefrontal cortical function, potentially improving decision-making and impulse control. Our results suggest that effective addiction treatment should combine early intensive intervention targeting reward and stress systems with sustained environmental support to allow executive function recovery. This multi-phase approach mirrors clinical observations that successful recovery requires both acute crisis management and long-term lifestyle changes. By understanding these neuroadaptive processes, we can design more targeted interventions that address the circuit-specific and temporally distinct aspects of addiction recovery, ultimately improving treatment outcomes for those struggling with substance use disorders. Declarations Acknowledgments We thank Miriam Melis and Celine Nicolas for helpful comments on a previous version of this manuscript. This study has benefited from the facilities and expertise of PREBIOS platform (Université de Poitiers). Funding This work was supported by the Centre National pour la Recherche Scientifique, the Institut National de la Santé et de la Recherche Médicale, the University of Poitiers, the Nouvelle Aquitaine CPER 2015-2020 / FEDER 2014-2020 program “Habisan”, the Fondation pour la Recherche Medicale (FRM, DPA20140629806 grant, PI: M. Solinas) and the Institut National du Cancer (Inca, CAD-V124-023-2024-105, PI: M. Solinas). Conflict of Interest Section: The authors declare no conflict of interest. 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1","display":"","copyAsset":false,"role":"figure","size":35265,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eGeneral experimental design and levels of self-administration in rats that were housed in SE or EE during abstinence. \u003c/strong\u003eA) Cocaine rats were allowed to self-administer cocaine for 25 X 6h sessions and then pseudo-randomly assigned to SE or EE conditions assuring similar levels of cocaine intake. 18FDG scan were performed after 1 week and 4 weeks of abstinence. Naive control animals were age-matched rats that were house similarly to cocaine rats but did not undergo surgery and self-administration procedures. B) Number of cocaine injections/sessions in future SE and future EE animals. C) Cocaine Intake in future SE and future EE animals. Notice that cocaine intake did not differ between the two groups.\u003c/p\u003e","description":"","filename":"fig1.png","url":"https://assets-eu.researchsquare.com/files/rs-6984132/v1/e364d07973905a4543ccab93.png"},{"id":98752901,"identity":"463c911c-2580-4225-894c-712b1004feb8","added_by":"auto","created_at":"2025-12-22 09:19:19","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":159480,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eEffects of cocaine self-administration on brain metabolic activity after one (A) and four (B) weeks of housing in SE. \u003c/strong\u003eChanges in metabolic activity in rats that self-administered cocaine (CocSE) compared to naive rats (NaiveSE) housed in standard environments during abstinence presented on representative coronal plates of the Paxinos and Watson atlas (upper panel), and on coronal images of z-score maps fused with an MRI template (lower panel). Increases in FDG uptake from dark red to yellow, decreases in FDG uptake from black to light blue; Student’s two-tailed t-test; p \u0026lt; 0.01. ACC, anterior cingulate cortex; DLStr, dorsolateral striatum; DMStr, dorsomedial striatum; Hipp, hippocampus; Ins, insula; Mot, motor cortex; NAc, nucleus accumbens; OFC, orbitofrontal cortex; PrL/IL, prelimbic/infralimbic cortices; SN/VTA: substantia nigra/ventral tegmental area.\u003c/p\u003e","description":"","filename":"fig2.png","url":"https://assets-eu.researchsquare.com/files/rs-6984132/v1/9fbe8a0ae6584ddf235ed412.png"},{"id":98779388,"identity":"656bac07-7449-4e52-aae7-229e32ede309","added_by":"auto","created_at":"2025-12-22 12:30:19","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":149108,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eEffects of cocaine self-administration on brain metabolic activity after one (A) and four (B) weeks of housing in EE. \u003c/strong\u003eChanges in metabolic activity in rats that self-administered cocaine (CocEE) compared to naive rats (NaiveEE) and were housed in enriched environments during abstinence presented on representative coronal plates of the Paxinos and Watson atlas (upper panel), and on coronal images of z-score maps fused with an MRI template (lower panel). Increases in FDG uptake from dark red to yellow, decreases in FDG uptake from black to light blue; Student’s two-tailed t-test; p \u0026lt; 0.01. ACC, anterior cingulate cortex; Amyg, amygdala; Hipp, hippocampus; Mot, motor cortex; NAc, nucleus accumbens; PrL/IL, prelimbic/infralimbic cortices.\u003c/p\u003e","description":"","filename":"fig3.png","url":"https://assets-eu.researchsquare.com/files/rs-6984132/v1/1c5f8f2efafc3285a2367e98.png"},{"id":98752903,"identity":"e4366fed-24bf-47f8-bbcd-772a4f4277eb","added_by":"auto","created_at":"2025-12-22 09:19:19","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":154951,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eEffects of housing in EE and SE for one (A) and four (B) weeks on cocaine-induced metabolic changes. \u003c/strong\u003eChanges in metabolic activity in rats that self-administered cocaine and were housed either in EE (CocEE) or SE (CocSE) during abstinence presented on representative coronal plates of the Paxinos and Watson atlas (upper panel), and on coronal images of z-score maps fused with an MRI template (lower panel). Increases in FDG uptake from dark red to yellow, decreases in FDG uptake from black to light blue; Student’s two-tailed t-test; p \u0026lt; 0.01. ACC, anterior cingulate cortex; Amyg, Amygdala; DLStr, dorsolateral striatum; Hipp, hippocampus; Ins, insula; NAc, nucleus accumbens; OFC, orbitofrontal cortex; SN/VTA: substantia nigra/ventral tegmental area.\u003c/p\u003e","description":"","filename":"fig4.png","url":"https://assets-eu.researchsquare.com/files/rs-6984132/v1/e24322a15b42f26e9dc391e5.png"},{"id":98777995,"identity":"4c2ddf8a-6bf1-48b1-85cd-503a0ad3aaf3","added_by":"auto","created_at":"2025-12-22 12:28:46","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":19525,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eIndividual normalized uptake values of FDG in the OF and NAc relative to its average cerebral uptake\u003c/strong\u003e. \u003cstrong\u003eA.\u003c/strong\u003e OFC, 1 week of abstinence. Left: cocaine vs naive in SE. Right: cocaine SE vs cocaine EE. \u003cstrong\u003eB. \u003c/strong\u003eNAc, 1 week of abstinence. Left: cocaine vs naive in SE. Right: cocaine SE vs cocaine EE. \u003cstrong\u003eC. \u003c/strong\u003eOFC, 4 weeks of abstinence. Left: cocaine vs naive in SE. Right: cocaine SE vs cocaine EE. \u003cstrong\u003eD.\u003c/strong\u003e NAc, 4 weeks of abstinence. Left : cocaine vs naive in SE. Right : cocaine SE vs cocaine EE. Notably, the normalized uptake values of FDG were calculated for voxels that met criteria described in the methods (p\u0026lt;0.01 for signals of at least 50 contiguous voxels, with effect size showing Cohen \u003cem\u003ed\u003c/em\u003e values of at least 0.80) and depended on specific comparisons. * p\u0026lt;0.05, ** p\u0026lt;0.01, *** p\u0026lt;0.001, **** p\u0026lt;0.0001.\u003c/p\u003e","description":"","filename":"fig5.png","url":"https://assets-eu.researchsquare.com/files/rs-6984132/v1/1aa17949e6c70cf0de9ba8fb.png"},{"id":103507972,"identity":"7d6f33ad-dd94-43d5-9ef6-33358a826f76","added_by":"auto","created_at":"2026-02-26 13:46:43","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1397295,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6984132/v1/a7595fc0-19a7-49f1-bc63-3cd874e2d0a9.pdf"},{"id":98777885,"identity":"8501c2c5-3b93-41f3-975c-7b96afe60ba8","added_by":"auto","created_at":"2025-12-22 12:28:37","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":2438363,"visible":true,"origin":"","legend":"Supplementary Figures and Tables","description":"","filename":"PBEEfDGPETNPPSupplementaryRevision.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6984132/v1/3eb6342e7f82711eeae306c9.pdf"}],"financialInterests":"There is \u003cb\u003eNO\u003c/b\u003e conflict of interest to disclose.","formattedTitle":"Environmental enrichment selectively counteracts brain metabolic activity during cocaine abstinence","fulltext":[{"header":"Introduction","content":"\u003cp\u003eOne of the hallmarks of addiction and one of its most troubling aspects is the long-lasting risk of relapse even after protracted periods of abstinence (\u003cspan additionalcitationids=\"CR2\" citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e). Over the past decades, neuroimaging studies in humans and mechanistic studies in animals have demonstrated that drugs of abuse produce persistent plastic changes in the brain that are thought to underlie the chronic nature of addiction (\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e). Based on clinical and brain-imaging data in humans, Goldstein and Volkow proposed the Impaired Response Inhibition and Salience Attribution (I-RISA) model that postulates that addiction results from dysregulation in the coordinated activity of different brain circuits (\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e). In this model, reduced activity in prefrontal and frontal cortical areas leads to deficits in executive function and response inhibition, while increased reactivity in limbic regions leads to heightened responsivity to drug-related cues and stress (\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e). In particular, studies in humans with cocaine addiction have shown hypoactivity in the anterior cingulate and orbitofrontal cortex associated with decision-making deficits (\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e), and hyperactivity in regions like the amygdala associated with increased stress reactivity (\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eAnimal studies have been essential in understanding the mechanisms underlying these brain changes (\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e, \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e). In the last decades, brain imaging studies have provided important insights into the changes induced by acute and chronic intake of drugs (\u003cspan additionalcitationids=\"CR14 CR15 CR16 CR17 CR18 CR19\" citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e) and the neuroadaptations that persist after discontinuation of drug use which could be associated with incubation of craving and long term risks of relapse (\u003cspan additionalcitationids=\"CR22 CR23 CR24 CR25\" citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e). In particular, using microPET imaging in rats, we have shown that extended access to cocaine self-administration produces persistent changes in brain metabolic activity that strikingly parallel those described in humans (\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e). Cocaine-exposed rats show decreased activity in cortical regions involved in executive function and decision-making, and increased activity in regions involved in motivation, memory, stress and emotional processing (\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e). Despite our growing understanding of cocaine-induced brain changes, developing effective treatments for addiction remains challenging.\u003c/p\u003e \u003cp\u003eAmong the novel strategies to help recovering from addiction, environmental enrichment (EE) has emerged as a promising therapeutic approach (\u003cspan additionalcitationids=\"CR28 CR29\" citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e). EE involves housing animals in stimulating environments that provide opportunities for social interaction, physical activity, and cognitive engagement (\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e). When implemented during periods of drug abstinence, EE consistently reduces drug seeking and taking behaviors in animal models of addiction (\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e, \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e). The therapeutic benefits of EE likely stem from its ability to engage natural reward systems and promote adaptive neuroplasticity (\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e). While a few studies have investigated neurobiological mechanisms associated with the beneficial effects of EE on relapse (\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e, \u003cspan additionalcitationids=\"CR33\" citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e), the mechanisms by which environmental enrichment produces its therapeutic effects on the brain and their dynamics remain poorly understood.\u003c/p\u003e \u003cp\u003eIn this study, we used FDG microPET imaging to investigate how exposure to environmental enrichment during abstinence affects cocaine-induced changes in basal brain metabolic activity. We first allowed male rats to self-administer cocaine under extended access conditions that lead to escalation of intake (\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e) and then housed them in either enriched or standard environments during four weeks of abstinence. By comparing brain activity patterns across groups and time points, we aimed to better understand how environmental enrichment influences the brain changes associated with cocaine addiction and abstinence.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\n \u003ch2\u003eSubjects and experimental design\u003c/h2\u003e\n \u003cp\u003eAdult (11\u0026ndash;12 weeks of age) male Sprague-Dawley rats (Janvier, France), experimentally naive at the start of the study, were housed in a temperature- and humidity-controlled room and maintained on a 12-h light/dark cycle (light on at 7.00 AM). All experiments were conducted in accordance with European Union directives (2010/63/EU) for the care of laboratory animals approved by the local ethics committee (COMETHEA, CEEA Val de Loire, #2015033014555999).\u003c/p\u003e\n\u003c/div\u003e\n\u003ch3\u003eHousing Conditions\u003c/h3\u003e\n\u003cp\u003eOn arrival, rats were housed two per cage for about 1 week before intrajugular catheterization surgery. Rats were anaesthetized using isoflurane (5% induction, 2.5% surgery) and implanted with catheters into the right jugular vein. Animals were allowed to recover for 5\u0026ndash;7 days before the self-administration sessions began. After surgery, rats were housed individually during the entire period of active self-administration. At the end of the last self-administration session, rats were pseudo-randomly divided into two groups, one housed in standard environments (SE) and the other in EE conditions assuring that similar levels of drug self-administration were observed in the two groups. SE and EE conditions were the same as previously used (\u003cspan class=\"CitationRef\"\u003e36\u003c/span\u003e\u0026ndash;\u003cspan class=\"CitationRef\"\u003e38\u003c/span\u003e). For SE, rats were housed in groups of three in cages sized 60 \u0026times; 38 \u0026times; 20 cm. For EE, rats were housed in groups of three in cages sized 80 \u0026times; 50 \u0026times; 100 cm. Each EE cage contained a house, a running wheel, three floors connected by ramps or tunnels and four toys that were changed once per week. Rats were housed in the animal facility in SE or EE housing conditions during a 4-week period of abstinence. Four experimental groups were obtained: NaiveSE, NaiveEE, CocSE and CocEE. The timeline of the experiment, including the different experimental groups, is illustrated in Fig. \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003eA.\u003c/p\u003e\n\u003ch3\u003eCocaine self-administration apparatus and procedure\u003c/h3\u003e\n\u003cp\u003e\u003cstrong\u003eCatheter implantation\u003c/strong\u003e. Rats were prepared for cocaine self-administration by surgical catheterization of the right jugular vein. Briefly, rats were anesthetized with isoflurane (5% induction, 2.5% maintenance) in O2 and administered with the nonsteroidal anti-inflammatory ketoprofen (2.5mg/kg, s.c.). Animals were placed on a heating pad from the induction of the anesthesia to the end of the surgery. A handmade silastic catheter was inserted into the jugular vein and the distal end was led to the back between the scapulae. Rats were allowed to recover for 7 days and flushed daily with 0.1ml sterile saline (0.9%), gentamicin (20mg/ml) and heparin (100 UI/mL) in sterile saline to help protect against infection and catheter occlusion.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eApparatus.\u003c/strong\u003e Experiments were conducted in MedAssociates operant-conditioning chambers, equipped with retractable levers as operanda, a cue-light above the active lever, a house light and controlled by MedAssociates interfaces and MED-PC IV software (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ewww.medassociates.com\u003c/span\u003e\u003c/span\u003e).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCocaine self-administration procedure\u003c/strong\u003e. Rats were allowed to self-administer cocaine (Cooper, France; 6g/L in saline solution 0.9%; 0.75mg/kg/infusion) for 6h/day for 25 sessions, using a Fixed Ratio 1 (FR1) schedule of reinforcement. A single press on the active lever resulted in one intravenous (i.v.) cocaine infusion with the concomitant activation of the light that remained on for 5s and then pulsed for 5s, followed by a 5s time-out. Inactive lever presses were recorded but did not produce any consequences. Naive control animals were age-matched rats that were house similarly to cocaine rats but did not undergo surgery and self-administration procedures.\u003c/p\u003e\n\u003cp\u003eAt the end of the last self-administration session, all rats were transferred from the animal facility at the University of Poitiers to the animal facility at University of Tours by an authorized transporter, placed in the SE or EE previously described and underwent abstinence for 4 weeks (EE: N\u0026thinsp;=\u0026thinsp;8 and SE: N\u0026thinsp;=\u0026thinsp;8). Metabolic imaging using FDG was performed in the same rats after 6\u0026ndash;8 days (1 week) and 27\u0026ndash;29 days (4 weeks) of abstinence. Naive rats of the same age and with analogous housing conditions but that did not undergo self-administration were used as controls (EE: N\u0026thinsp;=\u0026thinsp;7 and SE: N\u0026thinsp;=\u0026thinsp;8).\u003c/p\u003e\n\u003ch3\u003eBrain Imaging\u003c/h3\u003e\n\u003cp\u003eLocal uptake of FDG reflects cerebral metabolic rates of glucose utilization and allows the investigation of regional brain metabolic status (\u003cspan class=\"CitationRef\"\u003e39\u003c/span\u003e). Metabolic imaging using FDG was performed under basal conditions using the same procedure as previously described (\u003cspan class=\"CitationRef\"\u003e24\u003c/span\u003e). Briefly, rats were habituated to the PET experimental procedures for 4 days before each scan and fasted overnight before each scan. The day of brain-imaging acquisition, awake rats were injected with FDG (18.5 MBq/100 g i.p.; Cyclopharma, Tours), and placed in the habituation cage for 45 min. This timing was chosen based on previous studies that used similar approaches to investigate brain activity associated with behavioral performance (\u003cspan class=\"CitationRef\"\u003e40\u003c/span\u003e) activity before anesthesia. Then, they were anesthetized using isoflurane 4% (Baxter, Maurepas, France), placed on a heating pad (Minerve, Esternay, France) and centered in the field of view of the Explore VISTA-CT microPET camera (GE Healthcare, Velizy, France). CT-scan was performed for attenuation correction of PET images and a list-mode PET acquisition of 30 min started 60 min after FDG injection. After data reconstruction using a 2-D OSEM algorithm, all images were co-registered and normalized for tissue activity in the whole brain. Quantitative results were expressed as mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD and were presented on Z-score maps (for more details, refer to (\u003cspan class=\"CitationRef\"\u003e41\u003c/span\u003e)). Analyses focused on brain areas known to be key nodes in addiction and that exhibited changes in FDG uptake during cocaine abstinence: the cingulate (Cg), orbitofrontal (OFC), prelimbic/infralimbic (PrL/IL), insular and motor cortices, in addition to the dorsal striatum (DStr), nucleus accumbens (NAc), substantia nigra/ventral tegmental area (SN/VTA), amygdala (Amyg), and hippocampus (Hipp) (\u003cspan class=\"CitationRef\"\u003e24\u003c/span\u003e, \u003cspan class=\"CitationRef\"\u003e42\u003c/span\u003e).\u003c/p\u003e\n\u003cdiv id=\"Sec7\" class=\"Section2\"\u003e\n \u003ch2\u003eStatistical analysis\u003c/h2\u003e\n \u003cp\u003eFor self-administration, data were analyzed by two-way repeated measures ANOVA with time designated as a within-subject factor and future environment exposure (Coc SE or Coc EE) as a between-subject factor.\u003c/p\u003e\n \u003cp\u003eFor micro-PET data, a voxel-based analysis was used to assess the differences in cerebral FDG uptake between the averaged brains of cocaine vs control rats at each stage of abstinence for rats housed in EE vs SE. The regions of interest were derived from Schiffer\u0026rsquo;s templates (\u003cspan class=\"CitationRef\"\u003e43\u003c/span\u003e) using PMOD v3.2 software (PMOD Technologies Ltd, Switzerland) and applied to Z-score and effect size maps to obtain the Z-score and \u003cem\u003ed\u003c/em\u003e values in these areas (\u003cspan class=\"CitationRef\"\u003e41\u003c/span\u003e, \u003cspan class=\"CitationRef\"\u003e44\u003c/span\u003e). Statistical analyses focused on inter-group comparisons investigating differences in metabolic activity between (i) cocaine and naive rats housed in each environment at each time point, and (ii) EE vs SE in cocaine and naive rats at each time point using a two-tail unpaired Student t-test. Cohen d values were calculated to evaluate effect size of the differences. Notably, the effect of time of abstinence was not included as a factor in statistical analysis to avoid excessive comparisons which could inflate alpha errors. However, data in naive SE rats demonstrate that apart from a reduction in the activity in the dorsal striatum, brain activity was not significantly affected by testing at different times (Fig. \u003cspan class=\"InternalRef\"\u003eS1\u003c/span\u003e), suggesting that time per se and repeated testing produce limited effects on brain metabolic activity.\u003c/p\u003e\n \u003cp\u003eDifferences were considered significant when p\u0026thinsp;\u0026lt;\u0026thinsp;0.01 for signals of at least 50 contiguous voxels, with effect sizes showing Cohen \u003cem\u003ed\u003c/em\u003e values of at least 0.80.\u003c/p\u003e\n\u003c/div\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003eCocaine self-administration\u003c/h2\u003e \u003cp\u003ePrior to being assigned to different housing conditions, all rats showed similar patterns of cocaine self-administration (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eB, C). Statistical analysis revealed no significant difference between future SE and future EE rats in the number of active lever presses (two-way ANOVA: time, F\u003csub\u003e(24,360)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;5.81, p\u0026thinsp;\u0026lt;\u0026thinsp;0.05 ; group SE/EE, F\u003csub\u003e(1,14)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.1150, p\u0026thinsp;=\u0026thinsp;0.7395 ; interaction, F\u003csub\u003e(24,336)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.006, p\u0026thinsp;=\u0026thinsp;0.4582) or in cocaine intake (two-way ANOVA: time, F\u003csub\u003e(24,336)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;5.777,p\u0026thinsp;\u0026lt;\u0026thinsp;0.05 ; group SE/EE, F\u003csub\u003e(1,14)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.1147, p\u0026thinsp;=\u0026thinsp;0.7399 ; interaction, F\u003csub\u003e(24,336)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.7573, p\u0026thinsp;=\u0026thinsp;0.7896).\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eEffects of Environmental Enrichment on Brain Activity\u003c/h3\u003e\n\u003cp\u003eFirst, we investigated the effects of EE in naive rats (NaiveEE vs NaiveSE). One week of EE produced specific changes in brain metabolic activity (Supplementary Table\u0026nbsp;1 and Fig. S2A). Decreased activity was observed in the anterior cingulate cortex (ACC) (Cohen d\u0026thinsp;=\u0026thinsp;1.14, p\u0026thinsp;=\u0026thinsp;0.0149), insula (Ins) (Cohen d\u0026thinsp;=\u0026thinsp;1.24, p\u0026thinsp;=\u0026thinsp;0.0019), motor cortex (Mot) (Cohen d\u0026thinsp;=\u0026thinsp;1.13, p\u0026thinsp;=\u0026thinsp;0.0018), hippocampus (Hipp) (Cohen d\u0026thinsp;=\u0026thinsp;1.11, p\u0026thinsp;=\u0026thinsp;0.0104), and substantia nigra/ventral tegmental area (SN/VTA) (Cohen d\u0026thinsp;=\u0026thinsp;1.22, p\u0026thinsp;=\u0026thinsp;0.0002). Increased activity was found in prelimbic/infralimbic (PrL/IL) (Cohen d\u0026thinsp;=\u0026thinsp;1.16, p\u0026thinsp;=\u0026thinsp;0.0047), dorsomedial striatum (DMStr) (Cohen d\u0026thinsp;=\u0026thinsp;1.21, p\u0026thinsp;=\u0026thinsp;0.0011), dorsolateral striatum (DLStr) (Cohen d\u0026thinsp;=\u0026thinsp;1.19, p\u0026thinsp;=\u0026thinsp;0.0021), and posterior hippocampus (Hipp) (dorsal: Cohen d\u0026thinsp;=\u0026thinsp;1.15, p\u0026thinsp;=\u0026thinsp;0.0043; ventral: Cohen d\u0026thinsp;=\u0026thinsp;1.11, p\u0026thinsp;=\u0026thinsp;0.0067). After four weeks (Supplementary Table\u0026nbsp;1 and Fig. S2B), most changes normalized except for persistent alterations in motor cortex (increased, Cohen d\u0026thinsp;=\u0026thinsp;1.09, p\u0026thinsp;=\u0026thinsp;0.0081), insula (decreased, Cohen d\u0026thinsp;=\u0026thinsp;1.10, p\u0026thinsp;=\u0026thinsp;0.0220), ventral posterior hippocampus (increased, Cohen d\u0026thinsp;=\u0026thinsp;1.16, p\u0026thinsp;=\u0026thinsp;0.0011), and SN/VTA (decreased, Cohen d\u0026thinsp;=\u0026thinsp;1.09, p\u0026thinsp;=\u0026thinsp;0.0085).\u003c/p\u003e \u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003eEffects of Cocaine on Brain Metabolic Activity During Abstinence\u003c/h2\u003e \u003cp\u003eThen, we verified that we could replicate our findings of cocaine-induced changes in metabolic activity (\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e). Consistent with our previous findings, in rats housed in standard environments (CocSE vs NaiveSE), one week of abstinence was characterized by widespread decreases in metabolic activity (Supplementary Table\u0026nbsp;2 and Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eA), particularly in cortical regions including the ACC (Cohen d\u0026thinsp;=\u0026thinsp;1.14, p\u0026thinsp;=\u0026thinsp;0.0018), motor cortex (Cohen d\u0026thinsp;=\u0026thinsp;1.22, p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001), OFC (Cohen d\u0026thinsp;=\u0026thinsp;1.26, p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001), PrL/IL (Cohen d\u0026thinsp;=\u0026thinsp;1.12, p\u0026thinsp;=\u0026thinsp;0.0011), and insula (Cohen d\u0026thinsp;=\u0026thinsp;1.17, p\u0026thinsp;=\u0026thinsp;0.0017). The NAc also showed decreased activity (Cohen d\u0026thinsp;=\u0026thinsp;1.22, p\u0026thinsp;=\u0026thinsp;0.002). In contrast, increased activity was observed in the dorsomedial striatum (Cohen d\u0026thinsp;=\u0026thinsp;1.00, p\u0026thinsp;=\u0026thinsp;0.0242), dorsal posterior hippocampus (Cohen d\u0026thinsp;=\u0026thinsp;1.05, p\u0026thinsp;=\u0026thinsp;0.0155), ventral posterior hippocampus (Cohen d\u0026thinsp;=\u0026thinsp;1.05, p\u0026thinsp;=\u0026thinsp;0.0038), and SN/VTA (Cohen d\u0026thinsp;=\u0026thinsp;1.04, p\u0026thinsp;=\u0026thinsp;0.0396). After four weeks of abstinence (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eB), while some brain regions showed recovery (PrL/IL, NAc, and the motor cortex), persistent decreases were observed in the ACC (Cohen d\u0026thinsp;=\u0026thinsp;1.00, p\u0026thinsp;=\u0026thinsp;0.0362), OFC (Cohen d\u0026thinsp;=\u0026thinsp;1.21, p\u0026thinsp;=\u0026thinsp;0.0002), insula (Cohen d\u0026thinsp;=\u0026thinsp;1.17, p\u0026thinsp;=\u0026thinsp;0.0018), and dorsolateral striatum (Cohen d\u0026thinsp;=\u0026thinsp;1.15, p\u0026thinsp;=\u0026thinsp;0.012). The ventral posterior hippocampus (Cohen d\u0026thinsp;=\u0026thinsp;1.14, p\u0026thinsp;=\u0026thinsp;0.0008) and SN/VTA (Cohen d\u0026thinsp;=\u0026thinsp;1.00, p\u0026thinsp;=\u0026thinsp;0.0325) maintained increased activity. Finally, the anterior hippocampus showed switch from decrease to increase in metabolic activity (Cohen d\u0026thinsp;=\u0026thinsp;0.98, p\u0026thinsp;=\u0026thinsp;0.0074).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003eEffects of Environmental Enrichment on Cocaine-Induced Brain Changes\u003c/h2\u003e \u003cp\u003eWhen cocaine-exposed rats were housed in enriched environments (CocEE vs NaiveEE), changes in metabolic activity were found only in a few regions (Supplementary Table\u0026nbsp;3 and Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). Indeed, after one week (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eA), changes were limited to decreases in the ACC (Cohen d\u0026thinsp;=\u0026thinsp;1.04, p\u0026thinsp;=\u0026thinsp;0.0160), PrL/IL (Cohen d\u0026thinsp;=\u0026thinsp;1.12, p\u0026thinsp;=\u0026thinsp;0.0104), amygdala (Cohen d\u0026thinsp;=\u0026thinsp;1.16, p\u0026thinsp;=\u0026thinsp;0.0035), and anterior hippocampus (Cohen d\u0026thinsp;=\u0026thinsp;1.13, p\u0026thinsp;=\u0026thinsp;0.0071), with increases in NAc (Cohen d\u0026thinsp;=\u0026thinsp;1.12, p\u0026thinsp;=\u0026thinsp;0.0006) and posterior hippocampus (dorsal: Cohen d\u0026thinsp;=\u0026thinsp;1.12, p\u0026thinsp;=\u0026thinsp;0.0032; ventral: Cohen d\u0026thinsp;=\u0026thinsp;1.13, p\u0026thinsp;=\u0026thinsp;0.0138). After four weeks (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eB), only significant increases in ventral posterior hippocampus (Cohen d\u0026thinsp;=\u0026thinsp;1.11, p\u0026thinsp;=\u0026thinsp;0.0176) and significant decreases in the ACC (Cohen d\u0026thinsp;=\u0026thinsp;1.15, p\u0026thinsp;=\u0026thinsp;0.0049) and in the motor cortex (Cohen d\u0026thinsp;=\u0026thinsp;1.19, p\u0026thinsp;=\u0026thinsp;0.0004).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eDirect comparison between cocaine-experienced rats in EE versus SE (CocEE vs CocSE; Supplementary table 4 and Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e) revealed that after one week (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eA), EE rats showed higher metabolic activity in the ACC (Cohen d\u0026thinsp;=\u0026thinsp;1.13, p\u0026thinsp;=\u0026thinsp;0.0042), OFC (Cohen d\u0026thinsp;=\u0026thinsp;1.13, p\u0026thinsp;=\u0026thinsp;0.0003), insula (Cohen d\u0026thinsp;=\u0026thinsp;1.10, p\u0026thinsp;=\u0026thinsp;0.0017), dorsolateral striatum (Cohen d\u0026thinsp;=\u0026thinsp;1.06, p\u0026thinsp;=\u0026thinsp;0.0092), and NAc (Cohen d\u0026thinsp;=\u0026thinsp;1.11, p\u0026thinsp;=\u0026thinsp;0.0005), lower activity in the amygdala (Cohen d\u0026thinsp;=\u0026thinsp;1.15, p\u0026thinsp;=\u0026thinsp;0.0052), anterior hippocampus (Cohen d\u0026thinsp;=\u0026thinsp;1.05, p\u0026thinsp;=\u0026thinsp;0.0288), and SN/VTA (Cohen d\u0026thinsp;=\u0026thinsp;0.99, p\u0026thinsp;=\u0026thinsp;0.0485). After four weeks (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eB), differences persisted in the OFC (Cohen d\u0026thinsp;=\u0026thinsp;1.13, p\u0026thinsp;=\u0026thinsp;0.0028) and dorsolateral striatum (Cohen d\u0026thinsp;=\u0026thinsp;1.10, p\u0026thinsp;=\u0026thinsp;0.0039), while the ACC showed decreased activity (Cohen d\u0026thinsp;=\u0026thinsp;0.97, p\u0026thinsp;=\u0026thinsp;0.0257) and new differences emerged in ventral posterior hippocampus (Cohen d\u0026thinsp;=\u0026thinsp;1.03, p\u0026thinsp;=\u0026thinsp;0.0379).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003eFocus on the metabolic responses to cocaine in the OFC and the NAc as a function of exposure to SE and EE\u003c/b\u003e \u003c/p\u003e \u003cp\u003eBecause the effects of EE appeared particularly robust in the OFC and NAc, we focused our final comparison on these two regions. To better highlight differences between groups, we plotted the normalized FDG uptake values for individual rats. After one week of abstinence CocSE rats exhibited significantly lower uptake in both the OFC (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eA) and the NAc (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eB) compared with NaiveSE rats (OFC, p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; NAc, p\u0026thinsp;\u0026lt;\u0026thinsp;0.001). After four weeks, this reduction persisted in the OFC (p\u0026thinsp;\u0026lt;\u0026thinsp;0.001, Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eC) but was no longer apparent in the NAc (p\u0026thinsp;=\u0026thinsp;0.0777, Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eD). Compared to CocSE, exposure to CocEE showed increased normalized FDG uptake in the OFC both at 1 and 4 weeks (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eA, \u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eC) and in the NAc 1 week (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eC). In contrast, EE did not alter NAc uptake at 4 weeks (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eD, p\u0026thinsp;=\u0026thinsp;0.6737).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eOur findings reveal that cocaine self-administration produces persistent changes in brain metabolic activity that environmental enrichment can partially counteract. Specifically, cocaine decreased activity in cortical regions involved in executive function and the nucleus accumbens while increasing activity in emotional circuits. Environmental enrichment normalized activity in regions crucial for decision-making and reward processing, though some cocaine-induced changes remained unaffected.\u003c/p\u003e \u003cp\u003eCocaine addiction has been shown to be associated with long-lasting neuroadaptations in brain structures and functions that are believed to play a role in the persistent risks of relapse (\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e, \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e). These changes evolve throughout the course of abstinence: some emerge during the early stages and disappear after prolonged abstinence, others persist and some only manifest after several weeks of abstinence (\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e, \u003cspan additionalcitationids=\"CR47\" citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e). Importantly, whereas the temporary changes may reflect tolerance/withdrawal processes, those persisting after long periods of abstinence could contribute to the phenomenon of incubation of craving (\u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e, \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e) and may be more relevant to relapse after treatment and detoxification (\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e, \u003cspan additionalcitationids=\"CR47\" citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e). Consistent with our previous work (\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e), the changes we observed in cocaine-exposed rats closely parallel those reported in human addiction studies and align with the I-RISA model (\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e). During early abstinence, we found decreased metabolic activity in the OFC, ACC, and insula, accompanied by disrupted activity in the NAc and SN/VTA. Many of these alterations persisted through prolonged abstinence, while new changes emerged in the dorsolateral striatum and hippocampus, consistent with the development of habits and enhanced emotional processing associated with addiction (\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e, \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIn this study, environmental enrichment produced temporally distinct patterns of brain activity in naive versus cocaine-exposed animals. In naive rats, EE initially triggered widespread adaptations including decreased activity in the ACC, motor cortex, and insula, coupled with increased activity in striatal regions and hippocampus. Most of these changes normalized by four weeks, suggesting they represent acute adaptations to environmental stimulation rather than permanent circuit reorganization. These results are generally in agreement with previous studies using brain imaging approaches to investigate changes in brain activity following exposure to EE in mice (\u003cspan additionalcitationids=\"CR53 CR54\" citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e55\u003c/span\u003e) and in rats (\u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e) and with the known molecular neuroadaptations induced by EE (\u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e, \u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e). In cocaine-exposed animals, however, EE demonstrated markedly different effects. In particular, EE selectively normalized cocaine-induced disruptions in specific circuits. Interesting effects were seen in the OFC and NAc, where cocaine-induced decreases in metabolic activity were prevented when animals were housed in enriched conditions. These regions are crucial for value-based decision making and reward processing (\u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e59\u003c/span\u003e, \u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e60\u003c/span\u003e), suggesting EE may restore the ability to properly evaluate natural rewards and make adaptive choices. Importantly, these therapeutic-like effects showed circuit specificity. While executive and reward circuits showed substantial recovery, other cocaine-induced alterations persisted despite enrichment. Most notably, the increased metabolic activity in the hippocampus remained largely unchanged, suggesting that EE may not affect all addiction-related neural adaptations equally. This selective pattern of effects provides insight into which aspects of addiction may be most amenable to environmental interventions and which may require additional therapeutic approaches.\u003c/p\u003e \u003cp\u003eThe effects of environmental enrichment reveal distinct phases of therapeutic action. In early abstinence, EE rapidly normalized activity in reward-related circuits, particularly the NAc and in regions involved in interoception such as the insular cortex. This rapid normalization likely reflects immediate effects on reward processing and stress response systems. The quick restoration of NAc function could explain the rapid effects of EE on drug seeking (\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e), possibly by enhancing the salience of natural rewards provided by enriched environment. In contrast, whereas changes in cortical regions like the OFC and in the and dorsolateral striatum were sustained from early to late abstinence, changes in the ACC evolved from increased metabolic activity at the beginning of abstinence to a decrease after four weeks of abstinence. This response suggests involvement of sustained neuroadaptive processes, including structural plasticity through dendritic remodeling and synaptogenesis, alterations in gene expression patterns, circuit-level reorganization, and possible restoration of normal neurotransmitter function (\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e, \u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e, \u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e). This temporal pattern suggests that successful treatment requires initial targeting of reward and stress systems while maintaining enrichment long enough for executive function recovery, accounting for different temporal windows of plasticity across circuits.\u003c/p\u003e \u003cp\u003eWhereas most research has investigated the neurobiological consequences of drug taking, considerably less research has been dedicated to the investigation of the mechanisms associated with recovery from addiction. Our study addresses this knowledge gap by examining the neurobiological mechanisms underlying one specific pathway to recovery\u0026mdash;environmental enrichment. Brain imaging studies in humans and rats have demonstrated that long-term abstinence from cocaine is associated with heightened prefrontal cortical activity compared to early abstinence periods (\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e), which corresponds with our findings of progressive normalization of prefrontal function in EE animals. Furthermore, as discussed by Engeln \u0026amp; Ahmed (\u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e61\u003c/span\u003e), recovery can involve both the reversal of drug-induced changes and the introduction of new compensatory mechanisms that compete with drug-related circuits. These time-dependent effects of EE align with these observations, demonstrating that EE not only counteracts some drug-induced adaptations but potentially creates additional circuit-level changes that support recovery. The neuroadaptive changes we observed during EE-induced recovery likely represent just one pathway among many that can lead to remission. While our study shows that EE can counteract drug-induced neuroadaptations, particularly in the nucleus accumbens and prefrontal regions, it is important to recognize that other interventions targeting different neural mechanisms may achieve similar outcomes.\u003c/p\u003e \u003cp\u003eSeveral aspects should be considered when interpreting our results. First, our measures reflect metabolic activity under basal conditions, and therefore we cannot determine if and how environmental enrichment affects the reactivity of different brain regions to specific stimuli such as drug-related cues or stressors. Future studies using different behavioral challenges before imaging could provide insights into how environmental enrichment modifies brain responses to addiction-relevant triggers. Second, while PET imaging provides a valuable whole-brain perspective, its spatial resolution (approximately 1mm) limits our ability to distinguish small adjacent structures or subregions. For example, we analyzed the SN/VTA as a single region despite their distinct roles and could not differentiate between different amygdala nuclei or striatal subregions. More precise techniques like high-resolution fMRI or region-specific molecular analyses could complement our findings by providing finer anatomical detail. Third, to minimize the number of animals used, we performed repeated scans in the same rats. While this longitudinal approach has advantages for tracking changes over time, we cannot completely rule out potential effects of repeated manipulations such as fasting and anesthesia on brain activity. However, analysis of control rats at different time points showed limited effects of repeated testing (Fig.\u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003e). Fourth, we focused only on male rats in this study. Significant differences exist in self-administration of drugs in male and female rats (\u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e, \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e63\u003c/span\u003e). On the other hand, the beneficial effects of EE on drug-related effects appears to be similar between males and females (\u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e64\u003c/span\u003e, \u003cspan citationid=\"CR65\" class=\"CitationRef\"\u003e65\u003c/span\u003e). Future studies will be needed to determine whether the neurobiological mechanisms underlying the anti-craving effects of EE are similar in male and female rats. Fifth, the control group used in this study was naive i.e. did not undergo surgery and was not exposed to self-administration cages. While we cannot completely rule out the possibility that these manipulations contributed to differences in metabolic activity in cocaine vs control rats, the contribution of these experimental differences is likely limited compared to exposure to cocaine. Moreover, it should be noted that the main comparison of this work was between rats exposed to SE or EE during abstinence after a similar history of cocaine self-administration. Finally, while environmental enrichment in laboratory settings is well-defined, translation to human conditions is complex. Our enriched environment combines social, physical, and cognitive stimulation in ways that may not directly parallel human conventional environmental interventions. However, an accumulating body of literature suggests that specific aspects of enrichment such as physical activity (\u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e66\u003c/span\u003e, \u003cspan citationid=\"CR67\" class=\"CitationRef\"\u003e67\u003c/span\u003e), cognitive training (\u003cspan citationid=\"CR68\" class=\"CitationRef\"\u003e68\u003c/span\u003e, \u003cspan citationid=\"CR69\" class=\"CitationRef\"\u003e69\u003c/span\u003e) and social support (\u003cspan citationid=\"CR70\" class=\"CitationRef\"\u003e70\u003c/span\u003e) can be effective in the treatment of addiction. Importantly, a clinical trial is in progress to investigate the effects of combination of several of these stimulations on relapse to severe alcohol use disorder (\u003cspan citationid=\"CR71\" class=\"CitationRef\"\u003e71\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIn conclusion, this study reveals that environmental enrichment can normalize or counteract cocaine-induced brain metabolic changes through distinct temporal mechanisms. Our findings provide a neurobiological framework explaining how environmental interventions can help recovery from addiction: immediate effects may reduce craving intensity by normalizing nucleus accumbens activity, while sustained environmental stimulation gradually repairs prefrontal cortical function, potentially improving decision-making and impulse control. Our results suggest that effective addiction treatment should combine early intensive intervention targeting reward and stress systems with sustained environmental support to allow executive function recovery. This multi-phase approach mirrors clinical observations that successful recovery requires both acute crisis management and long-term lifestyle changes. By understanding these neuroadaptive processes, we can design more targeted interventions that address the circuit-specific and temporally distinct aspects of addiction recovery, ultimately improving treatment outcomes for those struggling with substance use disorders.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe thank Miriam Melis and Celine Nicolas for helpful comments on a previous version of this manuscript.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThis study has benefited from the facilities and expertise of PREBIOS platform (Université de Poitiers).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis work was supported by the Centre National pour la Recherche Scientifique, the Institut National de la Santé et de la Recherche Médicale, the University of Poitiers, the Nouvelle Aquitaine CPER 2015-2020 / FEDER 2014-2020 program “Habisan”, the Fondation pour la Recherche Medicale (FRM, DPA20140629806 grant, PI: M. Solinas) and the Institut National du Cancer (Inca, CAD-V124-023-2024-105, PI: M. Solinas).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflict of Interest Section:\u003c/strong\u003e The authors declare no conflict of interest.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor Contributions:\u0026nbsp;\u003c/strong\u003ePB, LG, MS: Designed research; PB, VL, ED, SS, SB, JB, MS: performed research; PB, ADM,CT, LG, MS: analyzed the data; PB, LG, MS: wrote the paper\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData availability:\u0026nbsp;\u003c/strong\u003eData will be made available upon request to the corresponding author\u003cstrong\u003e\u003cbr\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n \u003cli\u003eVenniro M, Banks ML, Heilig M, Epstein DH, Shaham Y. Improving translation of animal models of addiction and relapse by reverse translation. 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BMJ Open. 2023 May 12;13(5):e069249.\u003cstrong\u003e\u003cbr\u003e\u003c/strong\u003e\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-6984132/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6984132/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eEnvironmental enrichment (EE) is a promising strategy to promote recovery from addiction, but its neurobiological mechanisms remain poorly understood. In this study we used 18-fluorodeoxyglucose (FDG) microPET imaging to investigate how exposure to EE during abstinence affects brain neuroadaptations induced by voluntary intake of cocaine at different time of abstinence. After establishing escalated cocaine intake, male rats were housed in enriched or standard environments for four weeks and their brain metabolic activity was assessed after one and four weeks of abstinence. Cocaine self-administration produced widespread decreases in cortical metabolic activity, particularly in regions involved in executive function (orbitofrontal cortex, anterior cingulate), interoception (insula) and motivation (nucleus accumbens), while increasing activity in emotional circuits (ventral hippocampus) and the mesencephalon. EE selectively normalized these alterations restoring nucleus accumbens and orbitofrontal cortex activity. These findings reveal circuit-specific effects of environmental enrichment on cocaine-induced brain adaptations and suggest that effective addiction treatment requires both early interventions targeting reward circuits and sustained environmental stimulation to restore executive function, potentially reducing both immediate craving intensity and long-term relapse vulnerability.\u003c/p\u003e","manuscriptTitle":"Environmental enrichment selectively counteracts brain metabolic activity during cocaine abstinence","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-12-22 09:19:14","doi":"10.21203/rs.3.rs-6984132/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"ba39a1d7-5924-4f56-a2c3-d0ded80722ce","owner":[],"postedDate":"December 22nd, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":59159928,"name":"Health sciences/Diseases/Psychiatric disorders/Addiction"},{"id":59159929,"name":"Health sciences/Risk factors"},{"id":59159930,"name":"Biological sciences/Neuroscience/Diseases of the nervous system/Addiction"}],"tags":[],"updatedAt":"2026-02-25T19:45:52+00:00","versionOfRecord":[],"versionCreatedAt":"2025-12-22 09:19:14","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-6984132","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6984132","identity":"rs-6984132","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}