Muscarinic and NMDA Receptors in the Substantia Nigra Play a Role in Reward-Related Learning.

BACKGROUND: Reward-related learning, where animals form associations between rewards and stimuli (i.e., conditioned stimuli [CS]) that predict or accompany those rewards, is an essential adaptive function for survival. METHODS: In this study, we investigated the mechanisms underlying the acquisition and performance of conditioned approach learning with a focus on the role of muscarinic acetylcholine (mACh) and NMDA glutamate receptors in the substantia nigra (SN), a brain region implicated in reward and motor processes. RESULTS: Using RNAscope in situ hybridization assays, we found that dopamine neurons of the SN express muscarinic (mACh5), NMDA2a, NMDA2b, and NMDA2d receptor mRNA but not mACh4. NMDA, but not mACh5, receptor mRNA was also found on SN GABA neurons. In a conditioned approach paradigm, rats were exposed to 3 or 7 conditioning sessions during which light/tone (CS) presentations were paired with delivery of food pellets, followed by a test session with CS-only presentations. Intra-SN microinjections of scopolamine (a mACh receptor antagonist) or AP-5 (a NMDA receptor antagonist) were made either prior to each conditioning session (to test their effects on acquisition) or prior to the CS-only test (to test their effects on expression of the learned response). Scopolamine and AP-5 produced dose-dependent significant reductions in the acquisition, but not performance, of conditioned approach. CONCLUSIONS: These results suggest that SN mACh and NMDA receptors are key players in the acquisition, but not the expression, of reward-related learning. Importantly, these findings redefine the role of the SN, which has traditionally been known for its involvement in motor processes, and suggest that the SN possesses attributes consistent with a function as a hub of integration of primary reward and CS signals.

The Impact of Heroin Self-Administration and Environmental Enrichment on Ventral Tegmental CRF1 Receptor Expression.

BACKGROUND: There is a strong link between chronic stress and vulnerability to drug abuse and addiction. Corticotropin releasing factor (CRF) is central to the stress response that contributes to continuation and relapse to heroin abuse. Chronic heroin exposure can exacerbate CRF production, leading to dysregulation of the midbrain CRF-dopamine-glutamate interaction. METHODS: Here we investigated the role of midbrain CRF1 receptors in heroin self-administration and assessed neuroplasticity in CRF1 receptor expression in key opioid addiction brain regions. RESULTS: Infusions of antalarmin (a CRF1 receptor antagonist) into the ventral tegmental area (VTA) dose dependently reduced heroin self-administration in rats but had no impact on food reinforcement or locomotor activity in rats. Using RNAscope in situ hybridization, we found that heroin, but not saline, self-administration upregulated CRF1 receptor mRNA in the VTA, particularly on dopamine neurons. AMPA GluR1 and dopamine reuptake transporter mRNA in VTA neurons were not affected by heroin. The western-blot assay showed that CRF1 receptors were upregulated in the VTA and nucleus accumbens. No significant changes in CRF1 protein expression were detected in the prefrontal cortex, insula, dorsal hippocampus, and substantia nigra. In addition, we found that 15 days of environmental enrichment implemented after heroin self-administration does not reverse upregulation of VTA CRF1 receptor mRNA but it downregulates dopamine transporter mRNA. CONCLUSIONS: Overall, these data suggest that heroin self-administration requires stimulation of VTA CRF1 receptors and upregulates their expression in brain regions involved in reinforcement. Such long-lasting neuroadaptations may contribute to continuation of drug use and relapse due to stress exposure and are not easily reversed by EE exposure.

Low-dose polypharmacology targeting dopamine D1 and D3 receptors reduces cue-induced relapse to heroin seeking in rats.

Chemical compounds that target dopamine (DA) D1 or D3 receptors have shown promise as potential interventions in animal models of cue-induced relapse. However, undesirable side effects or pharmacodynamic profiles have limited the advancement of new compounds in preclinical studies when administered as independent treatments. In this series of experiments, we explored the effects of coadministration of a D1-receptor partial agonist (SKF 77434) and a D3-receptor antagonist (NGB 2904) in heroin-seeking rats within a "conflict" model of abstinence and cue-induced relapse. Rats were first trained to press a lever to self-administer heroin, and drug delivery was paired contingently with cues (e.g., light and pump noise). Self-initiated abstinence was facilitated by applying electrical current to the flooring in front of the levers. Lastly, a relapse response was provoked by noncontingent presentation of conditioned cues. Prior to provocation, rats received a systemic injection of SKF 77434, NGB 2904, or a combination of both compounds to assess treatment effects on lever pressing. Results indicated that the coadministration of low (i.e., independently ineffective) doses of both compounds was more effective in reducing cue-induced relapse to heroin seeking than either compound alone, with some evidence of drug synergism. Follow-up studies indicated that this reduction was not due to motoric impairment nor enhanced sensitivity to the electrified flooring and that this treatment did not significantly affect motivation for food. Implications for the treatment of opiate use disorder and recommendations for further research are discussed.

CaMKII antagonism in the ventral tegmental area impairs acquisition of conditioned approach learning in rats.

This study investigated the role of calcium2+/calmodulin-dependent protein kinase II (CaMKII), a protein in the second messenger pathway of NMDA receptors, in the ventral tegmental area (VTA) in the acquisition and performance of conditioned approach learning. Male Long-Evans rats (N = 79) were exposed to 3 (to test acquisition) or 7 (to test performance) conditioning sessions in which they received 30 paired presentations of a light stimulus (CS) and a food pellet (US) on a random time schedule. These conditioning sessions were then followed by one 30-min session without the CS or US and lastly by a CS-only test session, where only the light stimulus was presented (without food) according to the same schedule as the conditioning sessions. Bilateral intra-VTA injections of KN93 (vehicle, 3.0, 4.5 or 6.0 µg/0.5 µL), a CaMKII inhibitor, were administered prior to each conditioning session to test effects on the acquisition of conditioned approach or prior to the CS-only test session to test effects on the performance of conditioned approach. KN93, when given prior to conditioning sessions, significantly reduced the number of conditioned approach responses emitted during CS presentations in the CS-only test. When KN93 was given prior to the CS-only test it had no effect. These results suggest that CaMKII activation in the VTA is necessary for the acquisition, but not the performance, of reward-related learning.

Aberrations in Incentive Learning and Responding to Heroin in Male Rats After Adolescent or Adult Chronic Binge-Like Alcohol Exposure.

BACKGROUND AND PURPOSE: Binge drinking is a serious problem among adolescents and young adults despite its adverse consequences on the brain and behavior. One area that remains poorly understood concerns the impact of chronic intermittent ethanol (CIE) exposure on incentive learning. METHODS: Here, we examined the effects of CIE exposure during different developmental stages on conditioned approach and conditioned reward learning in rats experiencing acute or protracted withdrawal from alcohol. Two or 21 days after adolescent or adult CIE exposure, male rats were exposed to pairings of a light stimulus (CS) and food pellets for 3 consecutive daily sessions (30 CS-food pellet pairings per session). This was followed by conditioned approach testing measuring responses (food trough head entries) to the CS-only presentations and by conditioned reward testing measuring responses on a lever producing the CS and on another producing a tone. We then measured behavioral sensitization to repeated injections of heroin (2 mg/kg/d for 9 days). RESULTS: Adolescent and adult alcohol-treated rats showed significantly impaired conditioned reward learning regardless of withdrawal period (acute or prolonged). We found no evidence of changes to conditioned approach learning after adolescent or adult exposure to CIE. Finally, in addition to producing long-term impairments in incentive learning, CIE exposure enhanced locomotor activity in response to heroin and had no effect on behavioral sensitization to heroin regardless of age and withdrawal period. CONCLUSIONS: Our work sets a framework for identifying CIE-induced alterations in incentive learning and inducing susceptibility to subsequent opioid effects.

Dopamine and reward seeking: the role of ventral tegmental area.

Reward seeking is controlled by conditioned stimuli (CSs). There is a positive relation between mesocorticolimbic dopamine (DA) and the performance of learned reward-directed behavior. The mechanisms by which reward-, including drug-, associated stimuli come to acquire the capacity to activate the DA systems are not fully understood. In this review, we discuss the possible neurochemical mechanisms within the ventral tegmental area that may be involved in how CSs acquire the capacity to activate ventral tegmental area (VTA) DA neurons based on principles of long-term potentiation in the VTA and the role of mesocorticolimbic DA in reward-related learning. We propose that CSs function as such because they acquire the capacity to activate VTA DA neurons. Furthermore, CSs come to acquire this control of VTA DA cells when there is coincident N-methyl-d-aspartate receptor stimulation on VTA DA cells and strong depolarization of VTA DA cells, possibly by muscarinic acetylcholine receptor stimulation on these cells. This coincident activity leads to the strengthening of CS-associated glutamatergic synapses and the control by CSs of mesocorticolimbic DA systems and reward-directed behavior.

The D3 receptor antagonist SR 21502 reduces cue-induced reinstatement of methamphetamine-seeking in rats.

There is as of yet no FDA-approved medication for methamphetamine use disorder. Although dopamine D3 receptor antagonists have been shown to be useful in reducing methamphetamine seeking in animal models their translation to the clinic has been hindered because currently tested compounds can produce dangerously high blood pressure. Thus, it is important to continue to explore other classes of D3 antagonists. We report here the effects of SR 21502, a selective D3 receptor antagonist, on cue-induced reinstatement (i.e., relapse) of methamphetamine-seeking in rats. In Experiment 1, rats were trained to self-administer methamphetamine under a fixed ratio schedule of reinforcement followed by extinction of the response. Then, animals were tested with one of several doses of SR 21502 on cue-induced reinstatement of responding. SR 21502 significantly reduced cue-induced reinstatement of methamphetamine-seeking. In Experiment 2, animals were trained to lever press for food under a PR schedule and tested with the lowest dose of SR 21502 that caused a significant reduction in Experiment 1. These animals responded on average 8 times more than the vehicle-treated rats in Experiment 1, eliminating the possibility that SR 21502-treated rats in Experiment 1 responded less because they were incapacitated. In summary, these data suggest that SR 21502 may selectively inhibit methamphetamine-seeking and may constitute a promising pharmacotherapeutic agent for methamphetamine or other drug use disorders.

Environmental enrichment facilitates electric barrier induced heroin abstinence after incubation of craving in male and female rats.

BACKGROUND: Treatment strategies that aim to promote abstinence to heroin use and reduce vulnerability to drug-use resumption are limited in sustainability and long-term efficacy. We have previously shown that environmental enrichment (EE), when implemented after drug self-administration, reduces drug-seeking and promotes abstinence to cocaine and heroin in male rats. Here, we tested the effects of EE on abstinence in an animal conflict model in males and females, and after periods where incubation of craving may occur. METHODS: Male and female rats were trained to self-administer heroin followed by 3 or 21 days of a no-event-interval (NEI). Following NEI, rats were permanently moved to environmental enrichment (EE) or new standard (nEE) housing 3 days prior to resuming self-administration in the presence of an electric barrier adjacent to the drug access lever. Electric barrier current was increased daily until rats ceased self-administration. RESULTS: We found that 21 days of NEI led to significantly greater heroin self-administration and a trend toward shorter latencies to emit the first active lever press in the first abstinence session compared to 3 days of NEI. EE, when compared to nEE, led to longer latencies in the first abstinence session. Also, EE groups of both sexes and in both NEIs achieved abstinence criteria in significantly fewer numbers of sessions. CONCLUSIONS: EE facilitates abstinence in males and females and after periods where incubation of craving may occur. This suggests that EE may benefit individuals attempting to abstain from heroin use and may aid in the development of long term treatment strategies.

Environmental enrichment, administered after establishment of cocaine self-administration, reduces lever pressing in extinction and during a cocaine context renewal test.

The objective of this study was to test the hypothesis that environmental enrichment (EE) administered to rats previously trained to self-administer cocaine would reduce responding in extinction and in a cocaine-context renewal test. Long-Evans male rats were trained to press an active lever reinforced by cocaine (1.0 mg/kg/injection) under a fixed-ratio 1 schedule of reinforcement (inactive lever presses produced no consequences). After stable responding was established, all rats were given a 10-day break from the operant chambers followed by random assignment to EE (larger cages equipped with visual and auditory stimuli) or control (standard housing) group conditions in which they lived for the remainder of the experiment. Ten days after this move, rats were exposed to 10 extinction-responding sessions in a context different from the one in which self-administration occurred, followed by a context-renewal session occurring in the original self-administration context. The EE group responded significantly less in both the extinction and context-renewal sessions compared with the control group. These results suggest that EE reduces the ability of cocaine-associated stimuli to control cocaine-related responding.

Neurobiology of reward-related learning.

A major goal in psychology is to understand how environmental stimuli associated with primary rewards come to function as conditioned stimuli, acquiring the capacity to elicit similar responses to those elicited by primary rewards. Our neurobiological model is predicated on the Hebbian idea that concurrent synaptic activity on the primary reward neural substrate-proposed to be ventral tegmental area (VTA) dopamine (DA) neurons-strengthens the synapses involved. We propose that VTA DA neurons receive both a strong unconditioned stimulus signal (acetylcholine stimulation of DA cells) from the primary reward capable of unconditionally activating DA cells and a weak stimulus signal (glutamate stimulation of DA cells) from the neutral stimulus. Through joint stimulation the weak signal is potentiated and capable of activating the VTA DA cells, eliciting a conditioned response. The learning occurs when this joint stimulation initiates intracellular second-messenger cascades resulting in enhanced glutamate-DA synapses. In this review we present evidence that led us to propose this model and the most recent evidence supporting it.

Environmental enrichment reduces heroin seeking following incubation of craving in both male and female rats.

BACKGROUND: Contemporary treatments for heroin use disorder demonstrate only limited efficacy when the goals are long term abstinence and prevention of relapse. We have demonstrated that environmental enrichment (EE) reduces cue-induced heroin reinstatement in male rats. The present study is an attempt to extend the "anti-relapse" effects of EE to female rats and to periods where incubation of craving is hypothesized to occur. METHODS: This experiment implemented a 3-phase procedure. In Phase 1, male and female rats were trained to self-administer heroin for 15 days. Phase 2 consisted of a 3- or 15-day forced abstinence (FA) period. In Phase 3 half of the rats were placed into EE and the other half in non-EE housing and subsequently tested for responding in extinction (no heroin or cues) for 15 days followed by a cue-induced reinstatement test. RESULTS: We found that rats in the 15 days FA condition showed significantly enhanced drug seeking during extinction, irrespective of sex. We also found that EE significantly reduced this effect. During reinstatement, EE significantly reduced drug seeking in male and female rats and in both 3- and 15-day FA groups. CONCLUSIONS: EE, with or without prolonged FA, effectively reduced heroin seeking in male and female rats. These findings indicate that EE can reduce drug-seeking in males and females and when putative incubation of craving (i.e., prolonged abstinence period) has occurred and suggest that it may aid in the development of future long-term behavioral treatments for individuals at risk for heroin relapse.

Behavioral activating effects of selective AMPA receptor antagonism in the ventral tegmental area.

In recent years, the role of glutamate in drug seeking has received increasing attention. The current study tested the hypothesis that NBQX (a selective AMPA receptor antagonist) in the ventral tegmental area (VTA) would reduce cocaine seeking. Rats were trained to lever-press for intravenous cocaine (1.0 mg/kg/injection). Two test conditions were used, each preceded by bilateral intra-VTA microinjections of vehicle, 0.5, 1.0 or 2.0 µg/0.5 µl of NBQX. In the without-conditioned-stimulus (without-CS) condition, neither active (cocaine-associated) nor inactive levers produced any consequences. In the with-CS condition, the active lever turned on the stimulus light (as during training) and the infusion pump, which delivered an injection of saline. NBQX produced 2 effects: (1) an overall increase in responding, and (2) an increase in active lever-pressing at the 0.5-µg dose in the with-CS condition. A separate group of rats was allowed to press 2 levers without reinforcement, and the 0.5-µg NBQX dose enhanced responding on both levers. These results failed to support the hypothesis that AMPA receptor antagonism in VTA would reduce cocaine seeking and suggest that such antagonism causes general activating effects and may enhance the capacity of cocaine-CSs to function as conditioned reinforcers.

Therapeutic efficacy of environmental enrichment for substance use disorders.

Addiction to drug and alcohol is regarded as a major health problem worldwide for which available treatments show limited effectiveness. The biggest challenge remains to enhance the capacities of interventions to reduce craving, prevent relapse and promote long-term recovery. New strategies to meet these challenges are being explored. Findings from preclinical work suggest that environmental enrichment (EE) holds therapeutic potential for the treatment of substance use disorders, as demonstrated in a number of animal models of drug abuse. The EE intervention introduced after drug exposure leads to attenuation of compulsive drug taking, attenuation of the rewarding (and reinforcing) effects of drugs, reductions in control of behavior by drug cues, and, very importantly, relapse prevention. Clinical work also suggests that multidimensional EE interventions (involving physical activity, social interaction, vocational training, recreational and community involvement) might produce similar therapeutic effects, if implemented continuously and rigorously. In this review we survey preclinical and clinical studies assessing the efficacy of EE as a behavioral intervention for substance use disorders and address related challenges. We also review work providing empirical evidence for EE-induced neuroplasticity within the mesocorticolimbic system that is believed to contribute to the seemingly therapeutic effects of EE on drug and alcohol-related behaviors.

Contrasting effects of adolescent and early-adult ethanol exposure on prelimbic cortical pyramidal neurons.

BACKGROUND: Adolescence and early-adulthood are vulnerable developmental periods during which binge drinking can have long-lasting effects on brain function. However, little is known about the effects of binge drinking on the pyramidal cells of the prelimbic cortex (PrL) during early and protracted withdrawal periods. METHODS: In the present study, we performed whole-cell patch clamp recordings and dendritic spine staining to examine the intrinsic excitability, spontaneous excitatory post-synaptic currents (sEPSCs), and spine morphology of pyramidal cells in the PrL from rats exposed to chronic intermittent ethanol (CIE) during adolescence or early-adulthood. RESULTS: Compared to chronic intermittent water (CIW)-treated controls, the excitability of PrL-L5 pyramidal neurons was significantly increased 21 days after adolescent CIE but decreased 21 days after early-adult CIE. No changes of excitability in PrL Layer (L) 5 were detected 2 days after either adolescent or early-adulthood CIE. Interestingly, decreases in sEPSC amplitude and increases in thin spines ratio were detected 2 days after adolescent CIE. Furthermore, decreased frequency and amplitude of sEPSCs, accompanied by a decrease in the density of total spines and non-thin spines were observed 21 days after adolescent CIE. In contrast, increased frequency and amplitude of sEPSCs, accompanied by increased densities of total spines and non-thin spines were found 21 days after early adult CIE. CONCLUSION: CIE produced prolonged neuronal and synaptic alterations in PrL-L5, and the developmental stage, i.e., adolescence vs. early-adulthood when subjects receive CIE, is a key factor in determining the direction of these changes.

Repeated heroin in rats produces locomotor sensitization and enhances appetitive Pavlovian and instrumental learning involving food reward.

We tested the hypothesis that sensitization to heroin enhances appetitive motivational processes involving food reward. In Experiment 1, sixteen rats were exposed to positive pairings of a light stimulus and food for 4 consecutive daily sessions. Then, the rats received either saline or heroin (2 mg/kg) injections before placement in activity monitors for 9 consecutive daily sessions. Rats were then tested in operant conditioning chambers where one lever produced the light stimulus previously paired with food and another lever produced a tone stimulus not paired with anything. Heroin produced both significant progressive increases in locomotor activity (sensitization) and significantly enhanced conditioned reinforcement of instrumental lever pressing by the food-associated stimulus. In Experiment 2, thirty-two rats were given Pavlovian discrimination training in a conditioned magazine approach task where one stimulus was associated with food and a second unpaired with food. Rats then received repeated saline or heroin injections as in Experiment 1, before being tested under extinction conditions with the two stimuli without the drug. Chronic heroin had no effect on performance in this test, but it facilitated learning of the reversed discrimination in a subsequent phase. These data suggest that sensitization to heroin enhances appetitive motivational processes involving food reward.

Environmental enrichment facilitates cocaine abstinence in an animal conflict model.

In this study, we sought to discover if housing in an enriched environment (EE) is an efficacious intervention for encouraging abstinence from cocaine seeking in an animal "conflict" model of abstinence. Sixteen Long-Evans rats were trained in 3-h daily sessions to self-administer a cocaine solution (1 mg/kg/infusion) until each demonstrated a stable pattern of drug-seeking. Afterward, half were placed in EE cages equipped with toys, obstacles, and a running wheel, while the other half were given clean, standard laboratory housing. All rats then completed daily 30-min sessions during which the 2/3 of flooring closest to the self-administration levers was electrified, causing discomfort should they approach the levers; current strength (mA) was increased after every day of drug seeking until the rat ceased activity on the active lever for 3 consecutive sessions (abstinence). Rats housed in EE abstained after fewer days and at lower current strengths than rats in standard housing. These results support the idea that EE administered after the development of a cocaine-taking habit may be an effective strategy to facilitate abstinence.

Dopamine D1 and D3 receptor polypharmacology as a potential treatment approach for substance use disorder.

In the search for efficacious pharmacotherapies to treat cocaine addiction much attention has been given to agents targeting dopamine D1 or D3 receptors because of the involvement of these receptors in drug-related behaviors. D1-like and D3 receptor partial agonists and antagonists have been shown to reduce drug reward, reinstatement of drug seeking and conditioned place preference in rodents and non-human primates. However, translation of these encouraging results to clinical settings has been limited due to a number of factors including toxicity, poor pharmacokinetic properties and extrapyramidal and sedative side effects. This review highlights the role of D1 and D3 receptors in drug reward and seeking, the discovery of D1-D3 heteromers and their potential as targets in the treatment of addiction.

Blockade of NMDA receptors blocks the acquisition of cocaine conditioned approach in rats.

Conditioned stimuli (CSs) exert motivational effects on both adaptive and pathological reward-related behaviors, including drug taking and seeking. We developed a paradigm that allows us to investigate the neuropharmacology by which previously neutral stimuli acquire the capacity to function as CSs and elicit (intravenous) cocaine conditioned approach and used this paradigm to test the role of NMDA receptor stimulation in the acquisition of cocaine conditioned approach. Rats were injected systemically with the NMDA receptor antagonist, MK-801, before the start of 4 consecutive conditioning sessions, each of which consisted of 20 randomly presented light/tone (CS) presentations paired with cocaine infusion contingent upon nose pokes. Rats later were subjected to a CS-only test. To test the role of NMDA receptor stimulation in the already established conditioned approach, rats were injected with MK-801 prior to the CS-only test that occurred after 18 CS-cocaine conditioning sessions. Blockade of NMDA receptors significantly impaired the acquisition of cocaine-conditioned approach as indicated by the emission of significantly fewer nose pokes and significantly longer latencies to nose poke during CS presentations. When MK-801 treatment was applied after the acquisition of conditioned approach responding it had no effect on these measures. These results suggest that NMDA receptor stimulation plays an important role in the acquisition of reward-related conditioned responses driven by intravenous cocaine-associated CSs.

Rostral-caudal differences in the effects of intra-VTA muscimol on cocaine self-administration.

We have found that dopamine (DA) in the ventral tegmental area (VTA) plays an important role in cocaine self-administration. DA in the VTA acts at D1-type receptors on the terminals of GABA afferents causing release of this neurotransmitter. Thus, the neurochemical pathways whereby VTA DA might be involved in cocaine self-administration may include GABA neurotransmission. In the present study, we investigated this possibility. Rats were prepared with intravenous catheters and bilateral guide cannulae positioned to allow microinjections directly into the VTA or a site 1 mm dorsal to it. The rats were then trained to self-administer cocaine (1.0 mg/kg/injection) under a fixed-ratio 1 schedule of reinforcement and tested with microinjections of muscimol (0, 0.05 and 0.1 microg/0.25 microl) or picrotoxin (0, 0.025 and 0.05 microg/0.25 microl) or trained under a progressive ratio (PR) schedule and tested with vehicle and 0.05 microg/0.25 microl muscimol. Muscimol in the VTA, but not immediately dorsal to it, significantly reduced cocaine intake under the FR1 schedule. Furthermore, when analyzed by rostral/caudal site of injection, it was found that rostral injections of muscimol significantly reduced cocaine self-administration whereas caudal injections produced non-significant decreases in self-administration. Inspection of individual records revealed no signs of non-specific behavioral effects of the muscimol treatments. Muscimol in the rostral VTA also significantly increased break points in responding under the PR schedule. Intra-VTA picrotoxin did not significantly affect cocaine self-administration. These data suggest that stimulation of GABA-A receptors in the VTA is involved in cocaine self-administration and reward and that this involvement is more pronounced in the rostral than in the caudal VTA.

Blockade of muscarinic acetylcholine receptors in the ventral tegmental area disrupts food-related learning in rats.

RATIONALE: Stimulation of ventral tegmental area (VTA) muscarinic acetylcholine receptors (mAChRs) is implicated in feeding. OBJECTIVE: To investigate the effects of mAChR blockade in the VTA on food-related learning. METHODS: In experiment 1, rats (N=12) were placed in chambers containing food and received microinjections of 0 or 5 microg/0.5 microl scopolamine prior to the first four feeding sessions and the alternate dose prior to the tenth feeding session. In experiment 2 (N=9), the effects of daily microinjections of scopolamine on lever pressing under a progressive ratio schedule of food reinforcement were tested. In experiment 3 (N=34), the effects of daily microinjections of scopolamine on lever pressing maintained by conditioned reward were investigated. RESULTS: In experiment 1, all rats demonstrated low consumption during session 1. However, pellet consumption for rats initially pretreated with the 0-microg dose rose to and stayed at maximal levels for the remaining sessions, even when pretreated with the 5-microg dose during the tenth session. Pellet consumption for rats initially pretreated with the 5-microg dose remained low, even for the first two sessions following the cessation of scopolamine pretreatment, and gradually rose to maximal levels by the eighth session. In experiment 2, scopolamine significantly decreased break points. In experiment 3, scopolamine failed to significantly decrease responding specifically on the lever producing the conditioned reward. CONCLUSIONS: Altogether, these data suggest that VTA mAChR stimulation is involved in feeding and food-related learning but may not be involved in responding maintained by conditioned reward.