Intrinsic Functional Connectivity between the Anterior Insular and Retrosplenial Cortex as a Moderator and Consequence of Cocaine Self-Administration in Rats.

While functional brain imaging studies in humans suggest that chronic cocaine use alters functional connectivity (FC) within and between key large-scale brain networks, including the default mode network (DMN), the salience network (SN), and the central executive network (CEN), cross-sectional studies in humans are challenging to obtain brain FC prior to cocaine use. Such information is critical to reveal the relationship between individual's brain FC and the subsequent development of cocaine dependence and brain changes during abstinence. Here, we performed a longitudinal study examining functional magnetic resonance imaging (fMRI) data in male rats (n = 7), acquired before cocaine self-administration (baseline), on 1 d of abstinence following 10 d of cocaine self-administration, and again after 30 d of experimenter-imposed abstinence. Using repeated-measures analysis of variance (ANOVA) with network-based statistics (NBS), significant connectivity changes were found between anterior insular cortex (AI) of the SN, retrosplenial cortex (RSC) of the DMN, somatosensory cortex, and caudate-putamen (CPu), with AI-RSC FC showing the most robust changes between baseline and 1 d of abstinence. Additionally, the level of escalated cocaine intake is associated with AI-RSC and AI-CPu FC changes between 1 d and 30 d of abstinence; further, the subjects' AI-RSC FC prior to cocaine intake is a significant moderator for the AI-RSC changes during abstinence. These results provide novel insights into the roles of AI-RSC FC before and after cocaine intake and suggest this circuit to be a potential target to modulate large-scale network and associated behavioral changes in cocaine use disorders.

Stimuli predicting high-calorie reward increase dopamine release and drive approach to food in the absence of homeostatic need.

Animals and humans are motivated to consume high-fat, high-calorie foods by cues predicting such foods. The neural mechanisms underlying this effect are not well understood.Objective: We tested the hypothesis that cues paired with a food reward, as compared to explicitly unpaired cues, increase rats' food-seeking behavior by potentiating dopamine release in the nucleus accumbens, and that this effect would be less evident under satiety.Methods: We used a simple discriminative stimulus task and electrochemical recordings of dopamine release in freely moving rats.Results: We found that both food-predictive cue and hunger increased conditioned approaches to the receptacle (food-seeking behavior indicated by movement to the food receptacle). In addition, we observed dopamine release when the food-predictive cue (but not the unpaired cue) was presented, independent of hunger or satiety. Finally, we found a positive correlation between dopamine release amplitude and the number of conditioned approaches to the food receptacle in the sated condition, but not in the hungry condition.Discussion: Our results suggest that dopamine could drive seeking behavior for calorie-dense food in absence of homeostatic need, a core aspect of binge eating disorders.

Mifepristone Decreases Chronic Voluntary Ethanol Consumption in Rhesus Macaques.

The efficacy of short-term treatment with mifepristone (MIFE), a high-affinity, nonselective glucocorticoid receptor antagonist, to reduce ethanol drinking was tested in a rhesus macaque model. Stable individual daily ethanol intakes were established, ranging from 1.6 to 4.0 g/kg per day (n = 9 monkeys). After establishment of chronic ethanol intake, a MIFE dosing regimen that modeled a study of rodent drinking and human alcohol craving was evaluated. Three doses of MIFE (17, 30, and 56 mg/kg per day) were each administered for four consecutive days. Both 30 and 56 mg/kg decreased ethanol intake compared with baseline drinking levels without a change in water intake. The dose of 56 mg/kg per day of MIFE produced the largest reduction in ethanol self-administration, with the average intake at 57% of baseline intakes. Cortisol was elevated during MIFE dosing, and a mediation analysis revealed that the effect on ethanol drinking was fully mediated through cortisol. During a forced abstinence phase, access to 1.5 g/kg ethanol resulted in relapse in all drinkers and was not altered by treatment with 56 mg/kg MIFE. Overall, these results show that during active drinking MIFE is efficacious in reducing heavy alcohol intake in a monkey model, an effect that was related to MIFE-induced increase in cortisol. However, MIFE treatment did not eliminate ethanol drinking. Further, cessation of MIFE treatment resulted in a rapid return to baseline intakes, and MIFE was not effective in preventing a relapse during early abstinence. SIGNIFICANCE STATEMENT: Mifepristone reliably decreases average daily ethanol self-administration in a nonhuman primate model. This effect was mediated by cortisol, was most effective during open-access conditions, and did not prevent or reduce relapse drinking.

Behavioral Flexibility in Alcohol-Drinking Monkeys: The Morning After.

BACKGROUND: Heavy alcohol drinking has aspects of inflexible behavior. This study addressed the consequences of chronic alcohol drinking on cognitive and sensory-motor domains of behavioral flexibility in rhesus monkeys. METHODS: Behavioral flexibility was assessed in 12 monkeys (n = 9, ethanol [EtOH] drinkers) with a set-shifting visual discrimination procedure before alcohol self-administration and while maintaining consumption of 1.5 g/kg/d EtOH. Task performance was assessed in the morning after ~18 hours of drinking 1.5 g/kg, and 1 hour before the next day's drinking session began. The first 10 set-shifting sessions had the original (preethanol) test parameters and were used to determine retention of preethanol performance. Then, an effect of sensory-motor challenge (60% reduction in the size of the discriminative stimuli) on performance was assessed during 10 additional sessions. RESULTS: There were no average group-dependent differences in the performance between control and EtOH groups at the preethanol time-point. The daily consumption of 1.5 g/kg/d produced binge alcohol intakes in 7 of 9 monkeys (blood EtOH concentration [BEC ≥ 80 mg/dl]). Chronic daily intakes of 1.5 g/kg had no effect on retention of the task in the sober state. However, when challenged with a reduction in the size of the stimuli, daily 1.5 g/kg EtOH resulted in a decrement in performance due to an increase in the number of errors. CONCLUSIONS: Rhesus monkeys consuming 1.5 g/kg alcohol daily perform equally as could as control monkeys in retention of a well-learned cognitive task. However, this pattern of daily alcohol intake robustly decreased the ability to flexibly adjust behavior when confronted with novel changes to perceptual stimuli.

Ethanol Exposure History and Alcoholic Reward Differentially Alter Dopamine Release in the Nucleus Accumbens to a Reward-Predictive Cue.

BACKGROUND: Conditioned stimuli (CS) that predict reward delivery acquire the ability to induce phasic dopamine release in the nucleus accumbens (NAc). This dopamine release may facilitate conditioned approach behavior, which often manifests as approach to the site of reward delivery (called "goal-tracking") or to the CS itself (called "sign-tracking"). Previous research has linked sign-tracking in particular to impulsivity and drug self-administration, and addictive drugs may promote the expression of sign-tracking. Ethanol (EtOH) acutely promotes phasic release of dopamine in the accumbens, but it is unknown whether an alcoholic reward alters dopamine release to a CS. We hypothesized that Pavlovian conditioning with an alcoholic reward would increase dopamine release triggered by the CS and subsequent sign-tracking behavior. Moreover, we predicted that chronic intermittent EtOH (CIE) exposure would promote sign-tracking while acute administration of naltrexone (NTX) would reduce it. METHODS: Rats received 14 doses of EtOH (3 to 5 g/kg, intragastric) or water followed by 6 days of Pavlovian conditioning training. Rewards were a chocolate solution with or without 10% (w/v) alcohol. We used fast-scan cyclic voltammetry to measure phasic dopamine release in the NAc core in response to the CS and the rewards. We also determined the effect of NTX (1 mg/kg, subcutaneous) on conditioned approach. RESULTS: Both CIE and alcoholic reward, individually but not together, associated with greater dopamine to the CS than control conditions. However, this increase in dopamine release was not linked to greater sign-tracking, as both CIE and alcoholic reward shifted conditioned approach from sign-tracking behavior to goal-tracking behavior. However, they both also increased sensitivity to NTX, which reduced goal-tracking behavior. CONCLUSIONS: While a history of EtOH exposure or alcoholic reward enhanced dopamine release to a CS, they did not promote sign-tracking under the current conditions. These findings are consistent with the interpretation that EtOH can stimulate conditioned approach, but indicate that the conditioned response may manifest as goal-tracking.

Use of fast-scan cyclic voltammetry to assess phasic dopamine release in rat models of early postpartum maternal behavior and neglect.

Maternal behavior (MB) is a complex response to infant cues, orchestrated by postpartum neurophysiology. Although mesolimbic dopamine contributes toward MB, little is known about real-time dopamine fluctuations during the postpartum period. Thus, we used fast-scan cyclic voltammetry to measure individual dopamine transients in the nucleus accumbens of early postpartum rats and compared them with dopamine transients in virgins and in postpartum females exposed to cocaine during pregnancy, which is known to disrupt MB. We hypothesized that dopamine transients are normally enhanced postpartum and support MB. In anesthetized rats, electrically evoked dopamine release was larger and clearance was faster in postpartum females than in virgins and gestational cocaine exposure blocked the change in clearance. In awake rats, control mothers showed more dopamine transients than cocaine-exposed mothers during MB. Salient pup-produced stimuli may contribute toward differences in maternal phasic dopamine by evoking dopamine transients; supporting the feasibility of this hypothesis, urine composition (glucose, ketones, and leukocytes) differed between unexposed and cocaine-exposed infants. These data, resulting from the novel application of fast-scan cyclic voltammetry to models of MB, support the hypothesis that phasic dopamine signaling is enhanced postpartum. Future studies with additional controls can delineate which aspects of gestational cocaine reduce dopamine clearance and transient frequency.

Anatomical and pharmacological characterization of catecholamine transients in the medial prefrontal cortex evoked by ventral tegmental area stimulation.

Voltammetric measurements of catecholamines in the medial prefrontal cortex (mPFC) are infrequent because of lack of chemical selectivity between dopamine and norepinephrine and their overlapping anatomical inputs. Here, we examined the contribution of norepinephrine to the catecholamine release in the mPFC evoked by electrical stimulation of the ventral tegmental area (VTA). Initially, electrical stimulation was delivered in the midbrain at incremental depths of -5 to -9.4 mm from bregma while catecholamine release was monitored in the mPFC. Although catecholamine release was observed at dorsal stimulation sites that may correspond to the dorsal noradrenergic bundle (DNB, containing noradrenergic axonal projections to the mPFC), maximal release was evoked by stimulation of the VTA (the source of dopaminergic input to the mPFC). Next, VTA-evoked catecholamine release was monitored in the mPFC before and after knife incision of the DNB, and no significant changes in the evoked catecholamine signals were found. These data indicated that DNB fibers did not contribute to the VTA-evoked catecholamine release observed in the mPFC. Finally, while the D2-receptor antagonist raclopride significantly altered VTA-evoked catecholamine release, the α2-adrenergic receptor antagonist idazoxan did not. Specifically, raclopride reduced catecholamine release in the mPFC, opposite to that observed in the striatum, indicating differential autoreceptor regulation of mesocortical and mesostriatal neurons. Together, these findings suggest that the catecholamine release in the mPFC arising from VTA stimulation was predominately dopaminergic rather than noradrenergic.

Ethanol reduces evoked dopamine release and slows clearance in the rat medial prefrontal cortex.

BACKGROUND: Ethanol (EtOH) intoxication affects cognitive performance, contributing to attentional deficits and poor decision making, which may occur via actions in the medial prefrontal cortex (mPFC). mPFC function is modulated by the catecholamines dopamine and norepinephrine. In this study, we examine the acute effects of EtOH on electrically evoked dopamine release and clearance in the mPFC of anesthetized rats naïve to alcohol or chronically exposed to alcohol during adolescence. METHODS: Dopamine release and clearance was evoked by electrical stimulation of the ventral tegmental area (VTA) and measured in the mPFC of anesthetized rats with fast-scan cyclic voltammetry. In Experiments 1 and 2, effects of a high dose of EtOH (4 g/kg, intraperitoneally) on dopamine neurotransmission in the mPFC of EtOH-naïve rats and rats given EtOH exposure during adolescence were investigated. Effects of cumulative dosing of EtOH (0.5 to 4 g/kg) on the dopamine release and clearance were investigated in Experiment 3. Experiment 4 studied effects of EtOH locally applied to the VTA on the dopamine neurotransmission in the mPFC of EtOH-naïve rats. RESULTS: A high dose of EtOH decreased evoked dopamine release within 10 minutes of administration in EtOH-naïve rats. When tested via cumulative dosing from 0.5 to 4 g/kg, both 2 and 4 g/kg EtOH inhibited evoked dopamine release in the mPFC of EtOH-naïve rats, while 4 g/kg EtOH also slowed dopamine clearance. A similar effect on electrically evoked dopamine release in the mPFC was observed after infusion of EtOH into the VTA. Interestingly, intermittent EtOH exposure during adolescence had no effect on observed changes in mPFC dopamine release and clearance induced by acute EtOH administration. CONCLUSIONS: Taken together, these data describe EtOH-induced reductions in the dynamics of VTA-evoked mPFC dopamine release and clearance, with the VTA contributing to the attenuation of evoked mPFC dopamine release induced by EtOH.

Acute alcohol induces greater dose-dependent increase in the lateral cortical network functional connectivity in adult than adolescent rats.

Alcohol misuse and, particularly adolescent drinking, is a major public health concern. While evidence suggests that adolescent alcohol use affects frontal brain regions that are important for cognitive control over behavior little is known about how acute alcohol exposure alters large-scale brain networks and how sex and age may moderate such effects. Here, we employ a recently developed functional magnetic resonance imaging (fMRI) protocol to acquire rat brain functional connectivity data and use an established analytical pipeline to examine the effect of sex, age, and alcohol dose on connectivity within and between three major rodent brain networks: defaul mode, salience, and lateral cortical network. We identify the intra- and inter-network connectivity differences and establish moderation models to reveal significant influences of age on acute alcohol-induced lateral cortical network connectivity. Through this work, we make brain-wide isotropic fMRI data with acute alcohol challenge publicly available, with the hope to facilitate future discovery of brain regions/circuits that are causally relevant to the impact of acute alcohol use.

Low cognitive flexibility as a risk for heavy alcohol drinking in non-human primates.

Chronic alcohol abuse is frequently considered a habitual or inflexible behavior; however, measures of pre-existing cognitive flexibility prior to initiation of alcohol use are usually not available. This study used rhesus monkeys and an attentional set-shifting task to investigate whether pre-existing cognitive flexibility would predict increased risk for heavy alcohol drinking. As previously reported, monkeys were given 30 daily set-shifting sessions prior to alcohol access. These sessions consisted of the same sequence of eight unique visual discriminations (sets) of two objects that varied on two dimensions (shapes and colors). The ratio of errors per trials, session duration, and maximum set reached were primary dependent variables from each session and were used to compose a session performance index (PI) that ranged from a low performance PI of 31 to an optimal performance PI of 247. Here, animals underwent an alcohol induction period followed by 22 weeks of daily (22-h) self-administration sessions with free access to water and alcohol. Based on average daily alcohol intake during 22 weeks of 22-h/day access, the monkeys were categorized as non-heavy (mean = 2.0 ± 0.3 g/kg/day; n = 3) and heavy (mean = 3.3 ± 0.5 g/kg/day; n = 6) drinkers. The two groups diverged in performance on the set-shifting task across the 30 pre-alcohol sessions, and at the end of the pre-alcohol testing, the group average PI was 216 ± 27 and 137 ± 71 for the future non-heavy and heavy drinkers, respectively. The data show that low cognitive flexibility assessed with a set-shifting procedure was predictive of future classification as a heavy alcohol drinker. The data highlight individual differences in both cognitive flexibility and in alcohol self-administration in this population of rhesus monkeys.

Chronic Alcohol Drinking Slows Brain Development in Adolescent and Young Adult Nonhuman Primates.

The transition from adolescence to adulthood is associated with brain remodeling in the final stages of developmental growth. It is also a period when a large proportion of this age group engages in binge alcohol drinking (occasional consumption of four to five drinks leading to intoxication) and heavy alcohol drinking (binge drinking on ≥5 d in a month). Here we report on magnetic resonance imaging of developmental changes in the brain occurring during late adolescence and early adulthood (3.5-7.5 years of age) in a rhesus macaque model of alcohol self-administration. Monkeys were imaged prior to alcohol exposure, and following ∼6 and ∼12 months of daily (22 h/d) access to ethanol and water. The results revealed that the brain volume increases by 1 ml/1.87 years throughout the late adolescence and early adulthood in controls. Heavy alcohol drinking reduced the rate of brain growth by 0.25 ml/year per 1 g/kg daily ethanol. Cortical volume increased throughout this period with no significant effect of alcohol drinking on the cortical growth rate. In subcortical regions, age-dependent increases in the volumes of globus pallidus, thalamus, brainstem, and cerebellum were observed. Heavy drinking attenuated the growth rate of the thalamus. Thus, developmental brain volume changes in the span of late adolescence to young adulthood in macaques is altered by excessive alcohol, an insult that may be linked to the continuation of heavy drinking throughout later adult life.

Local µ-Opioid Receptor Antagonism Blunts Evoked Phasic Dopamine Release in the Nucleus Accumbens of Rats.

µ-opioid receptors (MORs) in the nucleus accumbens (NAc) can regulate reward-related behaviors that are dependent on mesolimbic dopamine, but the precise mechanism of this MOR regulation is unknown. We hypothesized that MORs within the NAc core regulate dopamine release. Specifically, we infused the MOR antagonist CTAP (d-Phe-Cys-Tyr-d-Trp-Arg-Thr-Pen-Thr-NH2) into the NAc core while dopamine release was evoked by electrical stimulation of the ventral tegmental area and measured by fast-scan cyclic voltammetry. We report that CTAP dose-dependently inhibited evoked dopamine release, with full blockade achieved with the 8 µg infusion. In contrast, evoked dopamine release increased after nomifensine infusion and was unchanged after vehicle infusion. These findings demonstrate profound local control of dopamine release by MORs within the NAc core, which has implications for regulation of reward processing.

Correction to: Acute phenylalanine/tyrosine depletion of phasic dopamine in the rat brain.

After publication of this paper, the authors determined an error in the calculation of the norepinephrine standard concentrations for the HPLC calibration curves.

Ranking Cognitive Flexibility in a Group Setting of Rhesus Monkeys with a Set-Shifting Procedure.

Attentional set-shifting ability is an executive function underling cognitive flexibility in humans and animals. In humans, this function is typically observed during a single experimental session where dimensions of playing cards are used to measure flexibility in the face of changing rules for reinforcement (i.e., the Wisconsin Card Sorting Test (WCST)). In laboratory animals, particularly non-human primates, variants of the WCST involve extensive training and testing on a series of dimensional discriminations, usually in social isolation. In the present study, a novel experimental approach was used to assess attentional set-shifting simultaneously in 12 rhesus monkeys. Specifically, monkeys living in individual cages but in the same room were trained at the same time each day in a set-shifting task in the same housing environment. As opposed to the previous studies, each daily session began with a simple single-dimension discrimination regardless of the animal's performance on the previous session. A total of eight increasingly difficult, discriminations (sets) were possible in each daily 45 min session. Correct responses were reinforced under a second-order schedule of flavored food pellet delivery, and criteria for completing a set was 12 correct trials out of a running total of 15 trials. Monkeys progressed through the sets at their own pace and abilities. The results demonstrate that all 12 monkeys acquired the simple discrimination (the first set), but individual differences in the ability to progress through all eight sets were apparent. A performance index (PI) that encompassed progression through the sets, errors and session duration was calculated and used to rank each monkey's performance in relation to each other. Overall, this version of a set-shifting task results in an efficient assessment of reliable differences in cognitive flexibility in a group of monkeys.

Adolescent binge-like alcohol alters sensitivity to acute alcohol effects on dopamine release in the nucleus accumbens of adult rats.

UNLABELLED: Rationale: Early onset of alcohol drinking has been associated with alcohol abuse in adulthood. The neurobiology of this phenomenon is unclear, but mesolimbic dopamine pathways, which are dynamic during adolescence, may play a role. OBJECTIVES: We investigated the impact of adolescent binge-like alcohol on phasic dopaminergic neurotransmission during adulthood. METHODS: Rats received intermittent intragastric ethanol, water, or nothing during adolescence. In adulthood, electrically evoked dopamine release and subsequent uptake were measured in the nucleus accumbens core at baseline and after acute challenge of ethanol or saline. RESULTS: Adolescent ethanol exposure did not alter basal measures of evoked dopamine release or uptake. Ethanol challenge dose-dependently decreased the amplitude of evoked dopamine release in rats by 30­50 % in control groups, as previously reported, but did not alter evoked release in ethanol-exposed animals. To address the mechanism by which ethanol altered dopamine signaling, the evoked signals were modeled to estimate dopamine efflux per impulse and the velocity of the dopamine transporter. Dopamine uptake was slower in all exposure groups after ethanol challenge compared to saline, while dopamine efflux per pulse of electrical stimulation was reduced by ethanol only in ethanol-naive rats. CONCLUSIONS: The results demonstrate that exposure to binge levels of ethanol during adolescence blunts the effect of ethanol challenge to reduce the amplitude of phasic dopamine release in adulthood. Large dopamine transients may result in more extracellular dopamine after alcohol challenge in adolescent-exposed rats and may be one mechanism by which alcohol is more reinforcing in people who initiated drinking at an early age.

Translational Research on Habit and Alcohol.

Habitual actions enable efficient daily living, but they can also contribute to pathological behaviors that resistant change, such as alcoholism. Habitual behaviors are learned actions that appear goal-directed but are in fact no longer under the control of the action's outcome. Instead, these actions are triggered by stimuli, which may be exogenous or interoceptive, discrete or contextual. A major hallmark characteristic of alcoholism is continued alcohol use despite serious negative consequences. In essence, although the outcome of alcohol seeking and drinking is dramatically devalued, these actions persist, often triggered by environmental cues associated with alcohol use. Thus, alcoholism meets the definition of an initially goal-directed behavior that converts to a habit-based process. Habit and alcohol have been well investigated in rodent models, with comparatively less research in non-human primates and people. This review focuses on translational research on habit and alcohol with an emphasis on cross-species methodology and neural circuitry.

Acute phenylalanine/tyrosine depletion of phasic dopamine in the rat brain.

RATIONALE: Dopamine plays a critical role in striatal and cortical function, and depletion of the dopamine precursors phenylalanine and tyrosine is used in humans to temporarily reduce dopamine and probe the role of dopamine in behavior. This method has been shown to alter addiction-related behaviors and cognitive functioning presumably by reducing dopamine transmission, but it is unclear what specific aspects of dopamine transmission are altered. OBJECTIVES: We performed this study to confirm that administration of an amino acid mixture omitting phenylalanine and tyrosine (Phe/Tyr[-]) reduces tyrosine tissue content in the prefrontal cortex (PFC) and nucleus accumbens (NAc), and to test the hypothesis that Phe/Tyr[-] administration reduces phasic dopamine release in the NAc. METHODS: Rats were injected with a Phe/Tyr[-] amino acid mixture, a control amino acid mixture, or saline. High-performance liquid chromatography was used to determine the concentration of tyrosine, dopamine, or norepinephrine in tissue punches from the PFC and ventral striatum. In a separate group of rats, phasic dopamine release was measured with fast-scan cyclic voltammetry in the NAc core after injection with either the Phe/Tyr[-] mixture or the control amino acid solution. RESULTS: Phe/Tyr[-] reduced tyrosine content in the PFC and NAc, but dopamine and norepinephrine tissue content were not reduced. Moreover, Phe/Tyr[-] decreased the frequency of dopamine transients, but not their amplitude, in freely moving rats. CONCLUSIONS: These results indicate that depletion of tyrosine via Phe/Tyr[-] decreases phasic dopamine transmission, providing insight into the mechanism by which this method modifies dopamine-dependent behaviors in human imaging studies.

Regional variation in phasic dopamine release during alcohol and sucrose self-administration in rats.

While dopamine input to the dorsal striatum is well-known to be critical for action selection, including alcohol-motivated behaviors, it is unknown whether changes in phasic dopamine accompany these behaviors. Long-term alcohol abuse is believed to promote alterations in the neurocircuitry of reward learning in both ventral and dorsal striatum, potentially through increasing dopamine release. Using fast-scan cyclic voltammetry, we measured phasic dopamine release in the dorsal and ventral striatum during alcoholic and nonalcoholic reward-seeking behavior and reward-related cues in rats trained on a variable-interval schedule of reinforcement. We observed robust phasic dopamine release in the dorsolateral striatum after reinforced lever presses and inconsistent dopamine release in the dorsomedial striatum. Contrary to our expectations, alcohol did not enhance dopamine release in rats drinking alcoholic rewards. Cue-induced dopamine release was also observed in the nucleus accumbens core of rats drinking the reward solutions. These data demonstrate that alcoholic and nonalcoholic reward self-administration on a variable-interval schedule of reinforcement in rats is accompanied by phasic dopamine release time-locked to reinforcement in the dorsolateral striatum and the nucleus accumbens, but not the dorsomedial striatum.