Adolescent Ethanol Exposure Alters Cholinergic Function and Apical Dendritic Branching Within the Orbital Frontal Cortex.

During adolescence, heavy binge-like ethanol consumption can lead to frontocortical structural and functional impairments. These impairments are likely driven by adolescence being a critical time point for maturation of brain regions associated with higher-order cognitive functioning. Rodent models of heavy binge-like ethanol exposure show consistent disruptions to the typical development of the prefrontal cortex (PFC). All deep cortical layers receive cholinergic projections that originate from the Nucleus basalis of Meynert (NbM) complex. These cholinergic projections are highly involved in learning, memory, and attention. Adolescent intermittent ethanol exposure (AIE) induces cholinergic dysfunction as a result of an epigenetic suppression of the genes that drive the cholinergic phenotype. The current study used a model of AIE to assess structural and functional changes to the frontal cortex and NbM following binge-like ethanol exposure in adolescence. Western blot analysis revealed long-term disruptions of the cholinergic circuit following AIE: choline acetyltransferase (ChAT) was suppressed in the NbM and vesicular acetylcholine transporter (VAChT) was suppressed in the orbitofrontal cortex (OFC). In vivo microdialysis for acetylcholine efflux during a spatial memory task determined changes in cholinergic modulation within the PFC following AIE. However, AIE spared performance on the spatial memory task and on an operant reversal task. In a second study, Golgi-Cox staining determined that AIE increased apical dendritic complexity in the OFC, with sex influencing whether the increase in branching occurred near or away from the soma. Spine density or maturity was not affected, likely compensating for a disruption in neurotransmitter function following AIE.

Persistent Alterations of Accumbal Cholinergic Interneurons and Cognitive Dysfunction after Adolescent Intermittent Ethanol Exposure.

Adolescent binge drinking renders young drinkers vulnerable to alcohol use disorders in adulthood; therefore, understanding alcohol-induced brain damage and associated cognitive dysfunctions is of paramount importance. Here we investigated the effects of binge-like adolescent intermittent ethanol (AIE) exposure on nonspatial working memory, behavioral flexibility and cholinergic alterations in the nucleus accumbens (NAc) in male and female rats. On postnatal days P25-57 rats were intubated with water or ethanol (at a dose of 5 g/kg) on a 2-day-on/2-day-off cycle and were then tested in adulthood on social recognition and probabilistic reversal learning tasks. During the social recognition task AIE-treated rats spent similar amounts of time interacting with familiar and novel juveniles, indicating an impaired ability to sustain memory of the familiar juvenile. During probabilistic reversal learning, AIE-treated male and female rats showed behavioral inflexibility as indicated by a higher number of trials needed to complete three reversals within a session, longer response latencies for lever selection, and for males, a higher number of errors as compared to water-treated rats. AIE exposure also reduced the number of cholinergic interneurons in the NAc in males and females. These findings indicate AIE-related pathologies of accumbal cholinergic interneurons and long lasting cognitive-behavioral deficits, which may be associated with cortico-striatal hypofunction.

The strength of reward-related learning depends on the degree of activation of ventral tegmental area dopamine neurons.

We tested whether (1) the capacity of a reward-associated conditioned stimulus (CS) to cause conditioned activation of ventral tegmental area (VTA) dopamine (DA) neurons is associated with its capacity to elicit conditioned approach responses and (2) whether the acquisition of these capacities by a CS requires N-methyl-D-aspartate (NMDA) and muscarinic acetylcholine (mACh) receptor stimulation. Rats were trained to emit a conditioned approach response to a light CS that was previously paired with food and were treated systemically with scopolamine (a mACh receptor antagonist) or MK-801 (an NMDA receptor antagonist) either prior to each conditioning session (during which animals experienced paired CS and food presentations) or prior to the conditioned approach (CS-only) test. Brains were harvested after the CS-only test and processed for tyrosine hydroxylase (TH) (DA cells) and c-fos in the VTA. When animals received scopolamine or MK-801 treatment prior to conditioning sessions we observed significantly fewer TH-labeled (i.e., DA) cells in the VTA that expressed c-fos and significantly less conditioned approach responding during the CS-only test. Further analysis showed a correlation between the number of VTA DA cells activated and the number of conditioned approach responses. Treatments made prior to the CS-only test did not affect responding. Altogether these results suggest that the degree to which a CS elicits conditioned approach depends partially on the degree to which the CS activates VTA DA cells and that the acquisition of both of these capacities by a CS requires mACh and NMDA receptor stimulation.

Blockade of muscarinic acetylcholine receptors in the ventral tegmental area blocks the acquisition of reward-related learning.

In the present study we investigated whether stimulation of muscarinic acetylcholine (mACh) receptors in the ventral tegmental area (VTA) plays a role in the acquisition of food-based conditioned approach learning. Rats were exposed to 3 (in Experiment 1) or 7 (in Experiment 2) conditioning sessions in which 30, randomly presented light (CS) presentations were paired with delivery of food pellets (US), followed by one session with no light or food and finally one CS-only test session with only light stimulus presentations. Bilateral microinjections of scopolamine (a mACh receptor antagonist) were made either prior to each conditioning session (Experiment 1; to test effects on acquisition) or prior to the CS-only test (Experiment 2; to test effects on performance of the learned response). Scopolamine produced a dose-related significant reduction in the acquisition of conditioned approach but had no effect on its performance. These results suggest that mACh receptor stimulation in the VTA plays a necessary role in the acquisition of reward-related learning.

No evidence that environmental enrichment during rearing protects against cocaine behavioral effects but as an intervention reduces an already established cocaine conditioned place preference.

OBJECTIVES: Environmental enrichment (EE) produces differential effects on psychostimulant-related behaviors. Therefore, we investigated whether the timing of EE exposure - during rearing and before cocaine exposure versus in adulthood and after cocaine exposure might be a determining factor. METHODS: In Experiment 1, rats reared with EE or not (non-EE) were conditioned with cocaine (5, 10 or 20mg/kg) in one compartment of a CPP apparatus and saline in the other, and later tested for cocaine CPP. In Experiment 2, locomotor activity in response to repeated injections of saline or cocaine was measured in rats raised with EE or non-EE. In Experiment 3 we measured the effects of EE or non-EE during rearing on food-based conditioned approach learning. In Experiment 4, rats were exposed to cocaine CPP conditioning then underwent 60days of EE or non-EE treatment after which they were tested for cocaine CPP. RESULTS: Our results show that rearing in EE did not reduce cocaine CPP or cocaine-induced locomotor activity (Experiments 1 and 2) but significantly facilitated conditioned approach learning (Experiment 3). On the other hand, EE treatment introduced after cocaine conditioning significantly reduced the expression of cocaine CPP (Experiment 4). CONCLUSIONS: These findings suggest that EE does not protect against cocaine's rewarding and stimulant effects but can reduce already established cocaine effects, suggesting that EE might be an effective treatment for cocaine addiction-related behaviors.

Dopamine D1 and D3 receptor interactions in cocaine reward and seeking in rats.

RATIONALE: Animal research has demonstrated a role of dopamine D1 and D3 receptors in cocaine reward and seeking. PURPOSE AND METHODS: Here, we investigated the potential interaction of these two dopamine receptors in cue-induced reinstatement of cocaine seeking, cocaine conditioned place preference (CPP), and cocaine self-administration in rats. RESULTS: The co-administration of a D3 receptor antagonist, NGB 2904 and a D1 partial agonist, SKF 77434, of doses which when administered individually produced no significant effects, prior to reinstatement or CPP tests significantly reduced lever pressing and time spent in the cocaine-paired environment, suggesting synergistic effects of the combined compounds on cocaine seeking. When given to rats self-administering cocaine under a progressive ratio schedule of reinforcement doses of NGB 2904 which were ineffective alone significantly enhanced the break point-reducing effects of SKF 77434. CONCLUSIONS: Our results indicate that the combined treatment with a D1 receptor partial agonist and D3 receptor antagonist produces robust decreases in cocaine seeking and reward. This suggests an interaction between dopamine D1 and D3 receptors in cocaine-related behaviors.

Environmental enrichment as a potential intervention for heroin seeking.

BACKGROUND: Heroin-related cues can trigger craving and relapse in addicts or heroin seeking in rats. In the present study we investigated whether environmental enrichment (EE) implemented after heroin exposure can reduce cue-induced reinstatement of heroin seeking and expression of heroin conditioned place preference. METHODS: In Experiment 1, male Long Evans rats that already acquired a heroin self-administration habit, were housed in enriched or non-enriched environments, underwent extinction training and later were tested for cue-induced reinstatement of heroin seeking. In Experiment 2, rats were conditioned with heroin in one compartment of a CPP apparatus and saline in the other, exposed to 30days of enrichment or no enrichment and were later tested for heroin CPP. RESULTS: The results showed that exposure to EE significantly reduced responding during the reinstatement test (Experiment 1) and prevented the expression of heroin CPP (Experiment 2). CONCLUSION: Our findings suggest that EE can be an effective behavioral approach to diminish the effects of conditioned cues on heroin seeking.

Treatment with a muscarinic acetylcholine receptor antagonist impairs the acquisition of conditioned reward learning in rats.

The neural mechanisms whereby a reward-associated stimulus gains reinforcing properties and comes to function as a conditioned reward (CR) are not understood. We propose that muscarinic acetylcholine (mACh) receptor stimulation is necessary for this type of learning. Here we tested the hypothesis that mACh receptor antagonism, with scopolamine, would attenuate the acquisition by a food-related stimulus of the capacity to function as a CR. Rats were exposed to 5 pre-exposure sessions during which two levers were present, one producing a light and the other a tone when pressed. This was followed by 3 conditioning sessions in which the levers were absent and the rats were presented with 30 light-food pairings delivered randomly. In the test session, the levers were present and presses on both levers were recorded. Different groups of rats received intraperitoneal injections of scopolamine (0, 0.375, 0.75 and 1mg/kg) either prior to each conditioning session or prior to the test session. All groups showed significantly greater responding on the light lever in the test compared to the pre-exposure sessions, demonstrating the CR effect. In animals treated prior to conditioning the scopolamine groups pressed significantly less on the light lever than the vehicle group. In animals treated prior to the test the increased lever pressing for light was similar for all groups. These data suggest that scopolamine impaired the acquisition of CR but not its expression. The results support the hypothesis that mACh receptor stimulation is important for the acquisition by reward-associated stimuli of the ability to function as CRs.

The dopamine D3 receptor antagonist, SR 21502, facilitates extinction of cocaine conditioned place preference.

BACKGROUND: Pharmacotherapeutic agents that could facilitate extinction of cocaine cues would be useful in the treatment of cocaine addiction. We tested whether SR 21502, a selective dopamine (DA) D3 receptor antagonist, can facilitate extinction of cocaine conditioned place preference (CPP) in rats. METHODS: In experiment 1, cocaine (10mg/kg) CPP was first established and then extinguished. During the extinction phase the rats were injected with SR 21502 and placed in the previously cocaine-paired compartment for four sessions and vehicle in the other compartment on four alternating sessions. The rats were then tested again for cocaine CPP. In experiment 2, different groups of rats were trained to associate SR 21502 with one compartment and saline with the other. RESULTS: In experiment 1, the animals spent significantly more time in the cocaine-paired compartment after cocaine conditioning than they did before conditioning. Subsequently, the animals treated with SR 21502 during the extinction phase spent significantly less time in the cocaine-paired compartment than the vehicle group. In experiment 2, animals conditioned with SR 21502 preferred neither side of the CPP apparatus, indicating that SR 21502 produced no effects of its own. CONCLUSIONS: These findings suggest that treatment with SR 21502, a DA D3 receptor antagonist, in the presence of cocaine cues can facilitate extinction of cocaine CPP and further suggest that this compound might be an effective cocaine addiction treatment.

Microinjections of a dopamine D1 receptor antagonist into the ventral tegmental area block the expression of cocaine conditioned place preference in rats.

Stimulation of dopamine (DA) D1 receptors in the ventral tegmental area (VTA) is involved in primary rewards. In the current study we investigated whether VTA D1 receptor stimulation likewise plays a role in mediating the rewarding effects of cocaine-associated stimuli, using the cocaine conditioned place preference (CPP) paradigm. Rats were prepared with cannulae so as to allow microinjections in the VTA and later conditioned to a cocaine-associated environment using the CPP paradigm. Prior to each conditioning session rats were injected with either saline or cocaine (10mg/kg, intraperitoneally) and then placed in one of the two sides of the CPP apparatus. Sessions lasted 30min a day over a period of eight days, such that rats alternated daily between consistently experiencing cocaine in one side and saline in the other. On the test day, which was conducted one day after conditioning, rats were given bilateral microinjections of one of four doses of the D1 antagonist, SCH 23390, (0, 2, 4 or 8µg/0.5µl) directly into the VTA and allowed free access to both sides of the apparatus. Preference for either side was measured as time spent in each side and compared to the same measures taken before conditioning. The D1 antagonist produced a dose-related, significant reduction in the preference for the cocaine-paired side compared to vehicle. These data suggest that the expression of cocaine conditioned place preference requires stimulation of VTA D1 receptors and, as such, are the first to suggest a role for VTA dendritically released DA in cocaine-, or other reward-, related learning.

The effects of the novel DA D3 receptor antagonist SR 21502 on cocaine reward, cocaine seeking and cocaine-induced locomotor activity in rats.

RATIONALE: There is a focus on developing D3 receptor antagonists as cocaine addiction treatments. OBJECTIVE: We investigated the effects of a novel selective D3 receptor antagonist, SR 21502, on cocaine reward, cocaine-seeking, food reward, spontaneous locomotor activity and cocaine-induced locomotor activity in rats. METHODS: In Experiment 1, rats were trained to self-administer cocaine under a progressive ratio (PR) schedule of reinforcement and tested with vehicle or one of three doses of SR 21502. In Experiment 2, animals were trained to self-administer cocaine under a fixed ratio schedule of reinforcement followed by extinction of the response. Then, animals were tested with vehicle or one of the SR 21502 doses on cue-induced reinstatement of responding. In Experiment 3, animals were trained to lever press for food under a PR schedule and tested with vehicle or one dose of the compound. In Experiments 4 and 5, in separate groups of animals, the vehicle and three doses of SR 21502 were tested on spontaneous or cocaine (10 mg/kg, IP)-induced locomotor activity, respectively. RESULTS: SR 21502 produced significant, dose-related (3.75, 7.5 and 15 mg/kg) reductions in breakpoint for cocaine self-administration, cue-induced reinstatement (3.75, 7.5 and 15 mg/kg) and cocaine-induced locomotor activity (3.75, 7.5 and 15 mg/kg) but failed to reduce food self-administration and spontaneous locomotor activity. CONCLUSIONS: SR 21502 decreases cocaine reward, cocaine-seeking and locomotor activity at doses that have no effect on food reward or spontaneous locomotor activity. These data suggest SR 21502 may selectively inhibit cocaine's rewarding, incentive motivational and stimulant effects.