Repeated blast model of mild traumatic brain injury alters oxycodone self‐administration and drug seeking

Each year, traumatic brain injuries (TBI) affect millions worldwide. Mild TBIs (mTBI) are the most prevalent and can lead to a range of neurobehavioral problems, including substance abuse. A single blast exposure, inducing mTBI alters the medial prefrontal cortex, an area implicated in addiction, for at least 30 days post injury in rats. Repeated blast exposures result in greater physiological and behavioral dysfunction than single exposure; however, the impact of repeated mTBI on addiction is unknown. In this study, the effect of mTBI on various stages of oxycodone use was examined. Male Sprague Dawley rats were exposed to a blast model of mTBI once per day for 3 days. Rats were trained to self‐administer oxycodone during short (2 h) and long (6 h) access sessions. Following abstinence, rats underwent extinction and two cued reinstatement sessions. Sham and rbTBI rats had similar oxycodone intake, extinction responding and cued reinstatement of drug seeking. A second group of rats were trained to self‐administer oxycodone with varying reinforcement schedules (fixed ratio (FR)‐2 and FR‐4). Under an FR‐2 schedule, rbTBI‐exposed rats earned fewer reinforcers than sham‐exposed rats. During 10 extinction sessions, the rbTBI‐exposed rats exhibited significantly more seeking for oxycodone than the sham‐injured rats. There was a positive correlation between total oxycodone intake and day 1 extinction drug seeking in sham, but not in rbTBI‐exposed rats. Together, this suggests that rbTBI‐exposed rats are more sensitive to oxycodone‐associated cues during reinstatement than sham‐exposed rats and that rbTBI may disrupt the relationship between oxycodone intake and seeking.     

Antagonism of the dopamine D1‐like receptor in mesocorticolimbic nuclei attenuates acute food deprivation‐induced reinstatement of heroin seeking in rats

The neurotransmitter dopamine (DA) plays a critical role in both priming‐and cue‐induced reinstatement of extinguished drug‐seeking behavior, but its role in stress‐induced reinstatement is less clear. Our laboratory has recently demonstrated that systemic administration of the DA D1‐like receptor antagonist, SCH 23390, attenuates acute food deprivation (FD) stress‐induced reinstatement. The current study was designed to elucidate the brain regions critical to the effect of SCH 23390 on FD stress‐induced reinstatement. Rats were trained to press a lever to self‐administer heroin (0.1 mg/kg/inf) over a period of 10 days. Following training, heroin was removed leading to an extinction of lever pressing. Next, rats were tested for reinstatement twice, under extinction conditions: once following 21–48 h FD; and once under sated conditions. Prior to testing, SCH 23390 was administered into the nucleus accumbens (NAc) shell (0.0, 0.3, 0.6 μg/side), NAc core (0.0, 0.3, 0.6 μg/side), dorsomedial prefrontal cortex (dmPFC; 0.0, 0.2, 2.0 μg/side), ventromedial prefrontal cortex (vmPFC; 0.0, 2.0 μg/side) or basolateral amygdala (BLA; 0.0, 1.0, 2.0 μg/side). An attenuation of FD‐induced reinstatement of heroin seeking was seen in rats injected with SCH 23390 into the NAc shell, dmPFC or BLA, but not into the NAc core or the vmPFC. These findings support the hypothesis that DA transmission through the DA D1‐like receptors plays a critical role in stress‐induced reinstatement of heroin seeking.     

Exercise during abstinence normalizes ultrastructural synaptic plasticity associated with nicotine‐seeking following extended access self‐administration

Nicotine‐craving progressively increases, or incubates, over abstinence following extended access self‐administration. While not yet examined for nicotine, the incubation of cocaine‐seeking is accompanied by changes in synaptic plasticity in the nucleus accumbens. Here, we determined whether such changes also accompany enhanced nicotine‐seeking following extended access self‐administration and abstinence, and whether exercise, a potential intervention for nicotine addiction, may exert its efficacy by normalizing these changes. Given that in humans, tobacco/nicotine use begins during adolescence, we used an adolescent‐onset model. Nicotine‐seeking was assessed in male rats following extended access nicotine or saline self‐administration (23‐hr/day, 10 days) and 10 days of abstinence, conditions known to induce the incubation of nicotine‐seeking, using a within‐session extinction/cue‐induced reinstatement procedure. A subset of rats had 2‐hr/day access to a running wheel during abstinence. Ultrastructural alterations of synapses in the nucleus accumbens core and shell were examined using electron microscopy. Nicotine‐seeking was elevated following extended access self‐administration and abstinence (in sedentary group), and levels of seeking were associated with an increase in the density of asymmetric (excitatory) and symmetric (inhibitory) synapses onto dendrites in the core, as well as longer asymmetric synapses onto spines, a marker of synaptic potentiation, in both the core and shell. Exercise normalized each of these changes; however, in the shell, exercise and nicotine similarly increased the synapse length. Together, these findings indicate an association between nicotine‐seeking and synaptic plasticity in the nucleus accumbens, particularly the core, and indicate that the efficacy of exercise to reduce nicotine‐seeking may be mediated by reversing these adaptations.     

Withdrawal from extended‐access cocaine self‐administration results in dysregulated functional activity and altered locomotor activity in rats

Much work has focused on determining the consequences of cocaine self‐administration on specific neurotransmitter systems, thus neglecting the global changes that occur. Previous imaging studies have focused on the effects of cocaine self‐administration in the presence of high blood levels of cocaine, but have not determined the functional effects of cocaine self‐administration after cocaine has cleared. Extended‐access cocaine self‐administration, where animals administer cocaine for 6 h each day, results in escalation in the rate of cocaine intake and is believed to model the transition from recreational use to addiction in humans. We aimed to determine the functional changes following acute (48 h) withdrawal from an extended‐access, defined‐intake self‐administration paradigm (5 days, 40 injections/day, 6 h/day), a time point when behavioral changes are present. Using the 2‐[¹⁴C]deoxyglucose method to measure rates of local cerebral glucose metabolism, an indicator of functional activity, we found reductions in circuits related to learning and memory, attention, sleep, and reward processing, which have important clinical implications for cocaine addiction. Additionally, lower levels of functional activity were found in the dorsal raphe and locus coeruleus, suggesting that cocaine self‐administration may have broader effects on brain function than previously noted. These widespread neurochemical reductions were concomitant with substantial behavioral differences in these animals, highlighted by increased vertical activity and decreased stereotypy. These data demonstrate that behavioral and neurochemical impairments following cocaine self‐administration are present in the absence of drug and persist after cocaine has been cleared.     

Effects of environmental enrichment on ERK1/2 phosphorylation in the rat prefrontal cortex following nicotine‐induced sensitization or nicotine self‐administration

Rats raised in an enriched condition (EC) exhibit alterations in the neurobiological and behavioral response to nicotine compared with rats reared in an impoverished condition (IC) or a standard condition (SC). The current study determined whether environmental enrichment differentially regulates extracellular signal‐regulated kinase1/2 (ERK1/2) activity in the prefrontal cortex in rats following nicotine sensitization or nicotine self‐administration. Under the saline control condition, EC rats displayed diminished baseline activity and greater sensitization to repeated administration of nicotine compared with IC and SC rats. After repeated saline injections, the basal levels of phosphorylated ERK1/2 (pERK1/2) were higher in EC compared with IC and SC rats, which was negatively correlated with their respective baseline activities. Repeated nicotine (0.35 mg/kg) injections induced pERK1/2 to similar levels in SC and IC rats; however, the induction of pERK1/2 in EC rats by nicotine was not significantly different from saline controls, owing to their high baseline. In the self‐administration paradigm, EC rats self‐administered less nicotine (0.03 mg/kg/infusion) relative to IC or SC rats on a fixed ratio‐1 schedule of reinforcement. Accordingly, no differences in pERK1/2 were found between EC and IC rats self‐administering saline, whereas nicotine self‐administration resulted in an increase in pERK1/2 in IC rats but not in EC rats. Furthermore, the levels of pERK1/2 in EC and IC rats were positively correlated with their respective total number of nicotine infusions. Thus, these findings suggest that environmental enrichment alters the basal and nicotine‐mediated pERK1/2, which may contribute to enrichment‐induced behavioral alterations in response to nicotine.     

Pattern of neural activation following yohimbine‐induced reinstatement of alcohol seeking in rats

Alcohol use disorders represent an extensive socioeconomic burden, yet effective treatment options are suboptimal. A major hurdle in treating alcohol use disorders is the high rate of relapse. Stress is a major factor that promotes relapse in abstinent drug users; therefore, understanding neural mechanisms that underpin the effects of stress on alcohol seeking is critical. In rodent models of stress‐induced relapse, the α₂‐adrenoceptor antagonist, yohimbine, is a widely used chemical stressor to elicit reinstatement of drug/alcohol seeking. However, the exact mechanism how yohimbine precipitates reinstatement of alcohol seeking and the pattern of neural activation associated with yohimbine‐induced reinstatement is poorly understood. Therefore, we counted Fos‐protein positive nuclei across 42 brain regions in alcohol‐experienced alcohol preferring rats that received either yohimbine in the home‐cage (1 mg/kg i.p.) or following yohimbine‐induced reinstatement of alcohol seeking. The number of Fos‐protein positive nuclei was increased in the prefrontal cortex and extended amygdala after home‐cage yohimbine compared to naïve‐ and vehicle‐treated rats. Yohimbine‐induced reinstatement increased the number of Fos‐protein expressing nuclei in multiple other regions including the thalamus, hypothalamus and hippocampus. We then examined inter‐regional correlations in Fos‐protein expression for all 42 brain regions, which showed Fos expression was more strongly positively correlated following yohimbine‐induced reinstatement of alcohol seeking, compared to home‐cage yohimbine. These data suggest low‐dose yohimbine in a non‐drug‐associated context activates stress/impulsivity centres within the brain, whereas yohimbine in the drug‐associated context recruits additional brain regions to drive alcohol seeking.     

The effects of methamphetamine self‐administration on cortical monoaminergic deficits induced by subsequent high‐dose methamphetamine administrations

Preclinical models suggest that repeated high‐dose methamphetamine (METH) exposures, administered in a “binge‐like” pattern, acutely decrease norepinephrine (NE), and acutely and persistently decrease serotonin (5‐hydroxytryptamine; 5HT) content in the frontal cortex. However, the impact of METH self‐administration on this region is unknown. Because of the importance of the monoaminergic neurons in the frontal cortex to a variety of cognitive and addictive processes, effects of METH self‐administration on cortical NE and 5HT content were assessed. Results revealed several novel findings. First, METH self‐administration decreased cortical NE content as assessed 24 h after last exposure. Consistent with previous preclinical reports after a binge METH regimen, this decrease was reversed 8 days after the final METH exposure. Second, and in contrast to our previous reports involving the hippocampus or striatum, METH self‐administration caused persistent decreases in 5HT content as assessed 8 days after the final METH exposure. Of note, the magnitude of this decrease (～20%) was less than that observed typically after a binge METH treatment. Third, prior METH self‐administration attenuated METH‐induced serotonergic deficits as assessed 7 days, but not 1 h, following a neurotoxic METH regimen. No protection was observed when the binge exposure occurred 15 days after the last self‐administration session. Taken together, these data demonstrate important and selective alterations in cortical serotonergic neuronal function subsequent to METH self‐administration. These data provide a foundation to investigate complex questions involving “resistance” to the persistent deficits caused by neurotoxic METH exposure and frontal cortical function. Synapse 67:875–881, 2013 . © 2013 Wiley Periodicals, Inc.     

Reversible inactivation of the basolateral amygdala,but not the dorsolateral caudate putamen,attenuates consolidation of cocaine‐cue associative learning in a reinstatement model of drug‐seeking

Previous research has shown that the basolateral amygdala (BLA) mediates stimulus‐reward learning, including drug‐cue associations, whereas the dorsolateral caudate putamen (dlCPu) primarily mediates stimulus‐response (habit) learning. Recent evidence has indicated that the dlCPu may be critical in cocaine‐seeking following extended self‐administration, but it remains unknown whether the dlCPu plays a role in the early formation of drug‐cue associations. The current study used a model of Pavlovian learning to compare the roles of the BLA and dlCPu in the consolidation of cocaine‐cue associations that maintain cocaine‐seeking during cue‐induced reinstatement. Male Sprague‐Dawley rats self‐administered cocaine (0.2 mg/50 μL infusion, i.v.) in the absence of cues for 6 days (2 h/day). Immediately following a single 1‐h classical conditioning session in which passive cocaine infusions were paired with a light/tone cue, animals received bilateral infusions of the GABA receptor agonists, baclofen/muscimol (1.0/0.1 mm ), or vehicle into the BLA or dlCPu. Following additional cocaine self‐administration (5 days) and subsequent extinction (no cocaine or cues, 7 days), the ability of the previously cocaine‐paired cues to reinstate cocaine‐seeking was assessed. Inactivation of the BLA, but not the dlCPu, immediately following the classical conditioning session impaired the consolidation of cocaine‐cue associations as seen by decreased cue‐induced reinstatement. These results extend previous findings that the BLA mediates the consolidation of learned associations that drive cocaine‐seeking during subsequent reinstatement and indicate that the dlCPu does not play a role during initial stimulus‐drug associative learning.     

Dorsomedial and dorsolateral striatum exhibit distinct phasic neuronal activity during alcohol self‐administration in rats

The development of alcoholism may involve a shift from goal‐directed to habitual drinking. These action control systems are distinct in the dorsal striatum, with the dorsomedial striatum (DMS) important for goal‐directed behavior and the dorsolateral striatum (DLS) required for habit formation. Goal‐directed behavior can be modeled in rats with a fixed ratio (FR) reinforcement schedule, while a variable interval (VI) schedule promotes habitual behavior (e.g. insensitivity to contingency degradation). Using extracellular recordings from chronically implanted electrodes, we investigated how DMS and DLS neurons encoded lever‐press responses and conditioned cues during operant alcohol self‐administration in these two models. In rats self‐administering 10% alcohol on an FR schedule, the DMS neuronal population showed increased firing at the onset of start‐of‐session stimuli. During self‐administration, the most prominent phasic firing patterns in the DMS occurred at the time of reinforcement and reinforcement‐associated cues, while the most prominent phasic activity in the DLS surrounded the lever response. Neural recordings from an additional cohort of rats trained on a VI schedule revealed a similar pattern of results; however, phasic changes in firing were smaller and differences between the medial and lateral dorsal striatum were less marked. In summary, the DMS and DLS exhibited overlapping but specialized phasic firing patterns: DMS excitations were typically time‐locked to reinforcement, while DLS excitations were generally associated with lever responses. Furthermore, the regional specificities and magnitudes of phasic firing differed between reinforcement schedules, which may reflect differences in behavioral flexibility, reward expectancy and the action sequences required to procure reinforcement.     

Effects of isoflurane anesthesia and intravenous morphine self‐administration on regional glucose metabolism ([18F]FDG‐PET) of male Sprague‐Dawley rats

Although certain drugs of abuse are known to disrupt brain glucose metabolism (BGluM), the effects of opiates on BGluM are not well characterized. Moreover, preclinical positron emission tomography (PET) studies anesthetize animals during the scan, which limits clinical applications. We investigated the effects of (i) isoflurane anesthesia and (ii) intravenous morphine self‐administration (MSA) on BGluM in rats. Jugular vein cannulated adult male Sprague‐Dawley rats self‐administered either saline (SSA) or morphine (0.5 mg/kg/infusion, 4 h/day for 12 days). All animals were scanned twice with [¹⁸F]‐fluoro‐deoxy‐glucose (FDG)‐PET/CT at a baseline and at 2‐day withdrawal from self‐administration. After the IV injection of FDG, one batch of animals (n = 14) was anesthetized with isoflurane and the other batch (n = 16) was kept awake during the FDG uptake (45 min). After FDG uptake, all animals were anesthetized in order to perform a PET/CT scan (30 min). Isoflurane anesthesia, as compared to the awake condition, reduced BGluM in the olfactory, cortex, thalamus, and basal ganglia, while increasing BGluM in the midbrain, hypothalamus, hippocampus, and cerebellum. Morphine self‐administered animals exhibited withdrawal signs (piloerection and increased defecation), drug seeking, and locomotor stimulation to morphine (0.5 mg/kg) during the 2 day withdrawal. The BGluM in the striatum was increased in the MSA group as compared to the SSA group; this effect was observed only in the isoflurane anesthesia, not the awake condition. These findings suggest that the choice of the FDG uptake condition may be important in preclinical PET studies and increased BGluM in the striatum may be associated with opiate seeking in withdrawal.     

Temporal integration as “common currency” of brain and self‐scale‐free activity in resting‐state EEG correlates with temporal delay effects on self‐relatedness

The self is a multifaceted phenomenon that integrates information and experience across multiple time scales. How temporal integration on the psychological level of the self is related to temporal integration on the neuronal level remains unclear. To investigate temporal integration on the psychological level, we modified a well‐established self‐matching paradigm by inserting temporal delays. On the neuronal level, we indexed temporal integration in resting‐state EEG by two related measures of scale‐free dynamics, the power law exponent and autocorrelation window. We hypothesized that the previously established self‐prioritization effect, measured as decreased response times or increased accuracy for self‐related stimuli, would change with the insertion of different temporal delays between the paired stimuli, and that these changes would be related to temporal integration on the neuronal level. We found a significant self‐prioritization effect on accuracy in all conditions with delays, indicating stronger temporal integration of self‐related stimuli. Further, we observed a relationship between temporal integration on psychological and neuronal levels: higher degrees of neuronal integration, that is, higher power‐law exponent and longer autocorrelation window, during resting‐state EEG were related to a stronger increase in the self‐prioritization effect across longer temporal delays. We conclude that temporal integration on the neuronal level serves as a template for temporal integration of the self on the psychological level. Temporal integration can thus be conceived as the “common currency” of neuronal and psychological levels of self.     

Electrophysiological evidence of mediolateral functional dichotomy in the rat accumbens during cocaine self‐administration: tonic firing patterns

Given the increasing research emphasis on putative accumbal functional compartmentation, we sought to determine whether neurons that demonstrate changes in tonic firing rate during cocaine self‐administration are differentially distributed across subregions of the NAcc. Rats were implanted with jugular catheters and microwire arrays targeting NAcc subregions (core, dorsal shell, ventromedial shell, ventrolateral shell and rostral pole shell). Recordings were obtained after acquisition of stable cocaine self‐administration (0.77 mg/kg/0.2mL infusion; fixed‐ratio 1 schedule of reinforcement; 6‐h daily sessions). During the self‐administration phase of the experiment, neurons demonstrated either: (i) tonic suppression (or decrease); (ii) tonic activation (or increase); or (iii) no tonic change in firing rate with respect to rates of firing during pre‐ and post‐drug phases. Consistent with earlier observations, tonic decrease was the predominant firing pattern observed. Differences in the prevalence of tonic increase firing were observed between the core and the dorsal shell and dorsal shell–core border regions, with the latter two areas exhibiting a virtual absence of tonic increases. Tonic suppression was exhibited to a greater extent by the dorsal shell–core border region relative to the core. These differences could reflect distinct subregional afferent processing and/or differential sensitivity of subpopulations of NAcc neurons to cocaine. Ventrolateral shell firing topographies resembled those of core neurons. Taken together, these observations are consistent with an emerging body of literature that differentiates the accumbens mediolaterally and further advances the likelihood that distinct functions are subserved by NAcc subregions in appetitive processing.     

Acquired self‐control of insula cortex modulates emotion recognition and brain network connectivity in schizophrenia

Real‐time functional magnetic resonance imaging (rtfMRI) is a novel technique that has allowed subjects to achieve self‐regulation of circumscribed brain regions. Despite its anticipated therapeutic benefits, there is no report on successful application of this technique in psychiatric populations. The objectives of the present study were to train schizophrenia patients to achieve volitional control of bilateral anterior insula cortex on multiple days, and to explore the effect of learned self‐regulation on face emotion recognition (an extensively studied deficit in schizophrenia) and on brain network connectivity. Nine patients with schizophrenia were trained to regulate the hemodynamic response in bilateral anterior insula with contingent rtfMRI neurofeedback, through a 2‐weeks training. At the end of the training stage, patients performed a face emotion recognition task to explore behavioral effects of learned self‐regulation. A learning effect in self‐regulation was found for bilateral anterior insula, which persisted through the training. Following successful self‐regulation, patients recognized disgust faces more accurately and happy faces less accurately. Improvements in disgust recognition were correlated with levels of self‐activation of right insula. RtfMRI training led to an increase in the number of the incoming and outgoing effective connections of the anterior insula. This study shows for the first time that patients with schizophrenia can learn volitional brain regulation by rtfMRI feedback training leading to changes in the perception of emotions and modulations of the brain network connectivity. These findings open the door for further studies of rtfMRI in severely ill psychiatric populations, and possible therapeutic applications. Hum Brain Mapp, 2013. © 2011 Wiley Periodicals, Inc.     

Electrophysiological evidence of mediolateral functional dichotomy in the rat nucleus accumbens during cocaine self‐administration II: phasic firing patterns

In the cocaine self‐administering rat, individual nucleus accumbens (NAcc) neurons exhibit phasic changes in firing rate within minutes and/or seconds of lever presses (i.e. slow phasic and rapid phasic changes, respectively). To determine whether neurons that demonstrate these changes during self‐administration sessions are differentially distributed in the NAcc, rats were implanted with jugular catheters and microwire arrays in different NAcc subregions (core, dorsal shell, ventromedial shell, ventrolateral shell, or rostral pole). Neural recording sessions were typically conducted on days 13–17 of cocaine self‐administration (0.77 mg/kg per 0.2‐mL infusion; fixed‐ratio 1 schedule of reinforcement; 6‐h daily sessions). Pre‐press rapid phasic firing rate changes were greater in lateral accumbal (core and ventrolateral shell) than in medial accumbal (dorsal shell and rostral pole shell) subregions. Slow phasic pattern analysis revealed that reversal latencies of neurons that exhibited change + reversal patterns differed mediolaterally: medial NAcc neurons exhibited more early reversals and fewer progressive/late reversals than lateral NAcc neurons. Comparisons of firing patterns within individual neurons across time bases indicated that lateral NAcc pre‐press rapid phasic increases were correlated with tonic increases. Tonic decreases were correlated with slow phasic patterns in individual medial NAcc neurons, indicative of greater pharmacological sensitivity of neurons in this region. On the other hand, the bias of the lateral NAcc towards increased pre‐press rapid phasic activity, coupled with a greater prevalence of tonic increase firing, may reflect particular sensitivity of these neurons to excitatory afferent signaling and perhaps differential pharmacological influences on firing rates between regions.     

Regional specificity in the real‐time development of phasic dopamine transmission patterns during acquisition of a cue–cocaine association in rats

Drug seeking is significantly regulated by drug‐associated cues and associative learning between environmental cues and cocaine reward is mediated by dopamine transmission within the nucleus accumbens (NAc). However, dopamine transmission during early acquisition of a cue–cocaine association has never been assessed because of the technical difficulties associated with resolving cue‐evoked and cocaine‐evoked dopamine release within the same conditioning trial. Here, we used fast‐scan cyclic voltammetry to measure sub‐second fluctuations in dopamine concentration within the NAc core and shell during the initial acquisition of a cue–cocaine Pavlovian association. Within the NAc core, cue‐evoked dopamine release developed during conditioning. However, within the NAc shell, the predictive cue appeared to cause an unconditioned decrease in dopamine concentration. The pharmacological effects of cocaine also differed between sub‐regions, as cocaine increased phasic dopamine release events within the NAc shell but not the core. Thus, real‐time measurements not only revealed the initial development of a conditioned neurochemical response but also demonstrated differential phasic dopamine transmission patterns across NAc sub‐regions during the acquisition of a cue–cocaine association.     

Incubation of nicotine seeking is associated with enhanced protein kinase A‐regulated signaling of dopamine‐ and cAMP‐regulated phosphoprotein of 32 kDa in the insular cortex

A recent clinical study demonstrated that damage to the insular cortex can disrupt tobacco addiction. The neurobiological mechanisms for this effect are not yet understood. In this study we used an animal model of nicotine addiction to examine the possibility that changes in insular cortex levels of dopamine (DA)‐ and cAMP‐regulated phosphoprotein of 32 kDa (DARPP‐32), a phosphoprotein enriched in DA neurons containing DA D1 receptors, may be associated with changes in vulnerability to nicotine addiction. Once rats acquired self‐administration, they were given unlimited access to nicotine (0.01 mg/kg/infusion) for 23 h/day for a total of 10 days. Each infusion was paired with a visual cue (stimulus light) and auditory cue (sound of pump). Nicotine seeking, as assessed under a cue‐induced reinstatement paradigm, and markers of DARPP‐32 signaling, as assessed using western blot analysis, were examined in separate groups of rats at two different abstinent intervals: 1 and 7 days. Consistent with findings with other drugs of abuse, rats in the 7‐day abstinence group took longer to extinguish and responded at higher levels during reinstatement testing as compared with rats in the 1‐day reinstatement group. Relative to saline controls, rats in the 7‐day but not the 1‐day abstinence group had higher levels of DARPP‐32 phosphorylated at the protein kinase A site in the insular cortex. These results demonstrate incubation of drug seeking following extended access to nicotine self‐administration and suggest that enhanced protein kinase A signaling in the insular cortex via phosphorylation of DARPP‐32 at Thr34 is associated with this effect.