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Acetylcholine and appetitive behavior
Running Head: ACETYLCHOLINE AND APPETITIVE BEHAVIOR
Nucleus accumbens acetylcholine regulates appetitive learning and
motivation for food via activation of muscarinic receptors
Wayne E. Pratt and Ann E. Kelley
University of Wisconsin-Madison Medical School
Pratt, W. E., & Kelley, A. E. (2004). Nucleus accumbens acetylcholine regulates appetitive
learning and motivation for food via activation of muscarinic receptors. Behavioral
Neuroscience, 118(4), 730-739. (doi: 10.1037/0735-7044.118.4.730)
© 2009 American Psychological Association
Note: This article may not exactly replicate the final version published in the APA journal. It is
not the copy of record.
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Acetylcholine and appetitive behavior
Abstract
These experiments tested whether nucleus accumbens muscarinic or nicotinic acetylcholine
receptor activation is required for rats to learn to lever press for sucrose. Muscarinic blockade
with scopolamine (1 or 10 g/side), but not nicotinic antagonism with mecamylamine (10
g/side), inhibited learning and performance when applied to the core or shell. Further
experiments showed that acute accumbens scopolamine treatment increased locomotor activity
and reduced sucrose consumption. However, microanalyses of behavioral events in the
instrumental chamber revealed that reductions of lever press performance during muscarinic
blockade were not due to gross motor dysfunction. Accumbens core scopolamine was
subsequently shown to reduce the amount of work rats would expend under a progressive-ratio
paradigm. These novel results implicate nucleus accumbens muscarinic receptors in the
modulation of appetitive learning, performance, and motivation for food.
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Nucleus accumbens acetylcholine regulates appetitive learning and
motivation for food via activation of muscarinic receptors
The nucleus accumbens is generally believed to be an important substrate for reward and
motivational processes, and is critical for the acquisition of multiple forms of appetitive
behaviors, reinforced by natural or drug rewards (e.g., Kelley, 1999; Koob, 1992). It receives
direct glutamatergic projections from cortical regions important for learning and memory
processes (e.g., McGeorge & Faull, 1989), as well as dopaminergic inputs from the tegmentum
argued to convey predictive reward information (Schultz, 1998). The medium spiny output
neurons of the nucleus accumbens subsequently project to ventral pallidum and the substantia
nigra (e.g., Heimer, Zahm, Churchill, Kalivas, & Wohltmann, 1991; Nauta & Domesick, 1984),
regions that subsequently modulate motor output. Pharmacological blockade of either
glutamatergic or dopaminergic receptors within the nucleus accumbens has been shown to impair
appetitive learning (Kelley, Smith-Roe, & Holahan, 1997; Smith-Roe & Kelley, 2000).
Recently, intrinsic cholinergic interneurons have also been implicated in mechanisms of
striatal plasticity. Although these large aspiny interneurons comprise only about 5% of the
striatum’s neuronal population (Graveland & DiFiglia, 1985), they are its primary source of
acetylcholine. Cholinergic modulation of striatum-based neural and behavioral plasticity is
supported by converging evidence from both in vitro and in vivo experiments that has shown
influences of muscarinic and nicotinic blockade on cellular plasticity and behavior (Calabresi,
Centonze, Gubellini, Pisani, & Bernardi, 1998; Centonze, Gubellini, Bernardi, & Calabresi,
1999; Partridge, Apparsundaram, Gerhardt, Ronesi, & Lovinger, 2002). Early behavioral studies
demonstrated that dorsal striatal cholinergic mechanisms are involved in learning of both leverpressing and active avoidance paradigms (for review see Prado-Alcala, 1985). Additionally,
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selective lesions of striatal cholinergic neurons in mice have been shown to impair rewardrelated learning (Kitabatake, Hikida, Watanabe, Pastan, & Nakanishi, 2003). Tonically active
neurons in monkeys, that are argued to be cholinergic based on similar firing patterns to
morphologically identified neurons in vitro, show flexible alterations in firing rate at the
presentation of primary reward or aversive stimuli (e.g., Matsumoto, Minamimoto, Graybiel, &
Kimura, 2001; Ravel, Legallet, & Apicella, 2003; Sardo, Ravel, Legallet, & Apicella, 2000).
Thus, the cholinergic interneurons of the striatum are capable of dynamic changes in relation to
salient environmental stimuli.
Despite the known role of the nucleus accumbens in appetitive learning paradigms, to our
knowledge no study has systematically examined the influence of intra-accumbens cholinergic
blockade on behavioral plasticity. These experiments explored the effects of muscarinic and
nicotinic antagonism within the rat nucleus accumbens upon instrumental learning in a leverpress task for sucrose reward. We report that muscarinic blockade of either nucleus accumbens
core or shell, but not nicotinic receptor antagonism, attenuates instrumental learning and
performance. Additional experiments examined acute locomotor and feeding effects of drug
treatment, as well as its impact on how hard rats would work to obtain sucrose under a
progressive ratio schedule. Together, the results from these experiments suggest a possible role
for nucleus accumbens cholinergic activation in mediating the motivational salience of sucrose
reward.
Method
Subjects and Housing
A total of 109 Male Sprague-Dawley rats (Harlan, Madison, WI) were dually housed in
clear plastic cages. The colony room was maintained at ~21 C on a 12:12 light-dark cycle
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(lights on at 7 am). Following 2 days acclimation to the laboratory, rats were handled daily for a
minimum of one week prior to surgeries. During this time, standard rat chow and water was
available ad libitum. All procedures and animal care were performed in accordance with NIH
guidelines for the ethical use of animals in research.
Surgery
Rats were anesthetized with a Ketamine-Xylazine cocktail (100 mg/kg-10mg/kg).
Standard aseptic procedures were utilized to implant indwelling stainless steel guide cannula (23
gauge) bilaterally above the nucleus accumbens core (flat skull; 1.3 mm anterior and 1.7 mm
lateral to bregma, 5.3 mm ventral to skull surface) or shell (tooth bar 5 mm above interaural zero;
3.1 mm anterior and 1 mm lateral to bregma, 5.3 mm ventral to skull surface). Guide cannula
were affixed to the skull with the use of screws and dental acrylic. Rats recovered for at least 7
days prior to food restriction and behavioral testing.
Drugs and Microinfusions
The competitive muscarinic acetylcholine receptor antagonist scopolamine methyl
bromide and the noncompetitive nicotinic acetylcholine receptor antagonist mecamylamine
(purchased from Sigma, St. Louis, MO) were dissolved in saline for these experiments. Drug
solutions were mixed at the beginning of each experiment, and stored in 60 l aliquots at 4 C
until used.
For all experiments, rats were habituated to injection procedures for three days prior to
experimental infusion. Each rat received two mock injections (during which injection cannula
were lowered only to the bottom of the guide cannulae and the infusion pump was activated
without solutions present) and one saline injection. For saline and drug infusions, microinfusion
cannula (30 gauge) were lowered to the desired target site (2.5 mm below the bottom of the 10
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mm guide cannula). These cannula were connected to a syringe via polyethylene tubing, and 0.5
l of solution was delivered (at a rate of .32 l per min) by a Harvard Apparatus (Holliston, MA)
microinfusion pump (infusion time = 1.33 min). Rats were gently restrained by the experimenter
during the infusions. Following one min diffusion time, the injection cannula were removed and
the wire stylets replaced. Rats were returned to their home cage for approximately five min,
after which they were transferred to the testing apparatus. Individual rats received no more than
seven infusion sessions, each separated by at least 24 hr.
Instrumental shaping procedure
Following recovery from surgery, food was restricted to allow for gradual reduction of
body weight to 85% of free-feeding levels. During food restriction, rats were given four grams
of sucrose (45 mg pellets, BioServ, Frenchtown, NJ) with their daily chow to prevent neophobia
during testing.
Rats were trained in commercially constructed instrumental chambers (Coulbourn
Instruments, Allentown, PA), equipped with two retractable levers, a house light, a row of
stimulus lights, and a food tray into which 45 mg sucrose pellets could be dispensed. The
chambers interfaced with a computer that recorded the time of each experimental event and
controlled all reinforcement contingencies. Rats were habituated to the chamber during the three
days of mock and saline injections noted above. Following the mock infusions, they were placed
into the chamber for 15 min. During these sessions, sucrose pellets were randomly delivered to
the food tray in intervals averaging 15 (RT-15”, mock day 1) or 30 (RT-30”, mock day 2 and
saline infusion days) seconds, allowing the rats to associate the reward with delivery and the
food tray.
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Rats were assigned to one of four drug treatment groups for the duration of the
experiment. In general, experimental groups comprised eight animals (actual numbers of all
groups are given in Method). These treatment groups received the following, injected bilaterally
in the volume of 0.5 l: 10 g mecamylamine, 10 g scopolamine, 1.0 g scopolamine, or saline
vehicle. These amounts of drug were determined from previous reports demonstrating efficacy
of these doses on behavioral assessments in other paradigms (e.g., Hildebrand, Panagis,
Svensson, & Nomikos, 1999; Ragozzino, Jih, & Tzavos, 2002).
On the first instrumental training session, rats were given the first drug treatment, and
five minutes later placed into the operant chamber. The right lever was projected into the
chamber and bar presses were reinforced on a FR1 (one sucrose pellet delivery per lever press)
schedule of reinforcement. A conjoint RT-30” schedule was maintained for the first 2
instrumental training sessions (Drug Days 1 and 2) to encourage exploratory behaviors. On the
third day of instrumental training (and continuing until the end of the experiment), the RT
schedule was removed, and upon completion of the 50th response in the session, the FR1
contingency was changed to an RR2 schedule of reinforcement (each response resulted in a
reinforcer delivery at a probability of 0.5). This limited the effects of satiety sometimes seen
with FR1 schedules, allowing for easier differentiation of treatment effects. All sessions were 15
min. Rats received drug treatments for the first 5 days of instrumental training, and a subsequent
injection on either day 11 or 14, once all groups had achieved an asymptotic level of
performance (see results). No infusions were given on intervening days. The primary dependent
variables were lever presses and nose pokes into the feeding chamber across days.
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Examination of unconditioned drug effects on locomotion and feeding
Separate groups of rats were handled, operated upon, and food restricted in the same
manner as for instrumental training. During mock infusion days, rats were taken to a novel room
and placed into a plastic cage similar to their home cage. Full access was given to water and
sucrose pellets during 15 min sessions. On drug treatment days, animals received a drug
infusion five minutes prior to placement in the cage. Feeding duration, feeding bouts, latency to
feed, total food intake, locomotion, rearing, and drinking was measured by an experimenter blind
to drug conditions via a keypad connected to a computer. These were within-subjects
experiments; each rat received each drug treatment across the course of several days. Infusions
were separated by at least 24 h, and presentation of the drug treatments were randomized for
each rat. Drugs and doses were identical to those used in the instrumental shaping paradigm.
Progressive ratio procedure
In this experiment, training preceded surgery. Following acclimation, handling, and
food-deprivation, eight naïve rats were habituated to the operant chambers with 3 daily sessions
of a RT-30” reinforcement schedule (with no lever present) for 30 min. On the subsequent and
following days, the lever was inserted into the chamber, and training proceeded for three sessions
each on FR1, FR3, and FR5 schedules of reinforcement, at which point all animals had achieved
reliable responding on the lever. Rats were then switched to a progressive ratio 2 (PR-2)
schedule of reinforcement for seven sessions. In this schedule, the rat was reinforced for the first
lever press, and then was required to increase the number of responses by two lever presses for
each subsequent pellet delivery. Thus, progressively more effort was required to earn each
reinforcer. The number of responses required in the final completed ratio determined the break
point, a well-validated measure reflecting the strength of the reinforcer and the motivational state
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of the animal (Arnold & Roberts, 1997; Hodos, 1961). At the end of seven days with the PR-2
schedule, all animals had achieved high levels of lever responding. They were returned to ad lib
feeding and underwent surgery to implant guide cannula above the nucleus accumbens core.
One week following surgery, the non-food deprived animals were returned to the
chambers, and the session length for the PR-2 schedule was increased to 1 hr. Once the rats had
achieved stable break points, they were habituated to the injection procedure (as above). Each
rat subsequently received five treatments of scopolamine (at 0, 0.1, 1.0, 5.0 and 10 g/side in 0.5
l saline) five minutes prior to the experimental sessions. The order of the drug presentation was
randomized for each animal, and drug treatments were separated by at least two days of
additional PR-2 training to stabilize baseline performance. Dependent measures were total
number of bar presses and the last completed reinforcement ratio (break point).
Statistical Analysis
Standard parametric statistics were applied to the data from these experiments. For the
instrumental learning paradigm, mixed design analyses of variance (ANOVAs) were conducted
with treatment as the between subjects factor and day of training as the within subjects factor.
Such ANOVAs were run for the entire training phase of the experiment to assess drug effects on
learning (i.e., for most experiments, days 1-13). Significant results of the ANOVA (p < .05)
were followed up by analyses of simple main and interaction effects, run between each treatment
group and the saline control group. A separate mixed-design ANOVA was run on the final two
days of the experiment (i.e., the final day of training and the final drug test) to assess the effects
of the drug on performance after the task was learned. Post-hoc comparisons of performance
utilized the Tukey HSD statistic. A group of rats was considered to be reliably lever-pressing on
the first day that the group’s lower-bound 95% confidence interval exceeded zero.
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To follow up on learning and performance effects of scopolamine treatment on the
instrumental learning paradigm, statistical analyses were supplemented by microstructural
behavior analysis. Raw data files for the last two days of each experiment were exported from
the Colbourn system into Excel®. Lever presses, nose pokes, and earned reinforcers that
occurred during each session were time-stamped by Graphic State Notation. Latencies were
calculated between three behavioral dyads: the time between each lever press and a nose poke,
the time between the delivery of a sucrose reinforcer and the subsequent nose poke, and the time
between a nose poke and the next bar press. The distribution of median latencies for each
scopolamine treatment group was compared between the final instrumental training day and the
subsequent drug test on performance. These microanalyses allowed a detailed assessment of
ongoing behavior in the operant chamber.
Standard repeated-measures ANOVAs were utilized to examine the effects of drug on
dependent measures in the locomotor and feeding control paradigm, as well as for the
progressive ratio experiment. Post-hoc comparisons were made with the Tukey HSD for
locomotion and food intake measures; planned contrasts were used to evaluate the effect of each
drug dose on break point and lever pressing measures (vs. the saline control) within the
progressive ratio experiment.
Histology
Once the experiments were complete, rats were deeply anesthetized with sodium
pentobarbital and perfused through the heart with a 0.9% buffered NaCl solution, followed by
10% formalin. Brains were removed and allowed to sink in 10% sucrose formalin. Sixty
micrometer frozen sections were then sliced through the penetrated area with a cryostat.
Sections were stained with cresyl violet. The tips of the cannula were confirmed by light
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microscopy, and charted in reference to (Paxinos & Watson, 1998). Eight animals were
excluded from additional analysis due to cannula misplacement. Representative
photomicrographs of cannula placements, as well as the charted locations of nucleus accumbens
core and shell placements for three experimental groups each, are shown in Figure 1.
Results
Effects of scopolamine and mecamylamine infusions into nucleus accumbens core or shell on
learning and performance of instrumental conditioning for sucrose reward
In general, intra-nucleus accumbens treatments with high (10 g/side) or low (1.0
g/side) doses of the muscarinic antagonist scopolamine impaired learning when infused into
either shell or core (Fig. 2). High doses of scopolamine also reduced performance on Day 14
following learning in both regions; the low dose impaired performance only in the nucleus
accumbens shell. Infusions of the nicotinic receptor antagonist mecamylamine had no effect on
learning or performance of the operant response. Similar systematic decreases were observed for
nose poke frequencies during muscarinic blockade (data not shown). Statistical presentation of
the bar press data for each experiment are detailed in subsequent sections for nucleus accumbens
core and shell manipulations.
Nucleus accumbens core. Examination of drug effects in the nucleus accumbens core
was comprised of two experiments; each tested scopolamine or mecamylamine treatment,
respectively, against a concurrent saline control group. For scopolamine-treated animals,
repeated measures ANOVA across days revealed significant effects for treatment group (F2,19 =
4.62, p < .05), day of training (F12,288 = 61.86, p < .001), and the treatment X day interaction
(F24,228 = 3.515, p < .001). As seen in Figure 2A, these effects were the result of a dosedependent learning impairment in the scopolamine groups during the initial five days of drug
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treatment, which followed through into the subsequent training trials. These effects were
verified with single effects analyses (utilizing corrected F values) comparing both scopolamine
groups with the control group; simple main effects were significant for both low (F1,19 = 6.59, p
< .05) and high dose (F1,19 = 10.12, p < .01) drug treatments. Treatment X day interactions also
achieved significance for both groups (p < .01). The control group and the 1.0 g/side group
showed reliable bar-pressing on day 3, although the rate of subsequent learning increased less
rapidly in the low dose group. The rats that received the high dose of the drug did not
demonstrate consistent bar-pressing behavior until day 9.
Comparison of lever pressing rates across the final two days of the experiment revealed
an interaction effect between days and treatment (F2,19 = 9.13, p < .01), as well as a significant
main effect of treatment day (F1,19 = 23.82, p < .01). Tukey’s HSD statistic was used to compare
the number of bar presses between the final training day and the performance test within each
treatment group. Although all groups pressed less on the treatment day compared to the previous
training day, the decreases in lever pressing were not reliably significant for saline or low-dose
scopolamine (p > .05); the high dose scopolamine group demonstrated impaired performance
following drug treatment (p < .01).
There was no effect of mecamylamine treatment in the nucleus accumbens core on the
learning of the instrumental task (treatment main effect: F1,15 = 0.06, p > .10; treatment X day
interaction: F9,135 = 0.94, p >.10). Both saline and mecamylamine animals were reliably lever
pressing on day 3 of training. Neither was there an effect of mecamylamine treatment on
performance (treatment X day interaction: F1,15 = 0.56, p > .10).
Nucleus accumbens shell. Muscarinic and nicotinic blockade in the nucleus accumbens
shell were run together in two separate cohorts of animals, each with a full control group.
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Controls were combined for the final analysis (Ncontrol group = 15; 8 rats were in each drug group).
Figure 2b shows the results of this experiment. Once again, injections of scopolamine, but not
mecamylamine, impaired acquisition of the instrumental response. Repeated measures
ANOVAs revealed significant effects for treatment group (F3,35 = 4.02, p < .05), day of training
(F12,420 = 103.04, p < .001), and the treatment X day interaction (F36,420 = 3.14, p < .001). This
result was limited to impairment of learning caused by the scopolamine groups; corrected F
values comparing the saline and mecamylamine conditions showed no effect of treatment group
(F1,35 = 0.56, p > .10) or treatment X day interaction (F12,420 = 1.37, p > .10). Similar analyses
revealed treatment X day interaction effects for both the low (F12,420 = 2.47, p < .01) and high
dose (F12,420 = 4.81, p < .01) scopolamine treatments versus the control group. The main effect
of group was significant only for the low dose group (F1,35 = 7.34, p < .05). Mecamylaminetreated animals began reliably pressing on day 3, as did controls. Reliable lever pressing did not
occur until day 6 for the low dose scopolamine group, nor until day 8 in the high dose condition.
Thus, scopolamine treatments in the shell, as in the nucleus accumbens core, impaired
instrumental conditioning in a dose-dependent manner. It should be noted that the higher
variability and lower asymptotic performance in the low dose scopolamine group was the result
of two animals that never learned the task.
A significant interaction of treatment X day was observed on the final two days of testing
(F3,35 = 10.78, p < .001). Tukey HSD comparison of group means between days suggested that
both the low (p < .05) and high dose (p < .01) of scopolamine impaired post-training
performance, whereas there was no effect of saline or mecamylamine infusion (p > .05).
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Unconditioned effects of cholinergic receptor blockade on locomotor and feeding behavior
In order to examine possible non-specific effects of the drug treatments on locomotion
and food intake, separate groups of food-deprived animals were subjected to a 15 minute sucrose
intake session under the influence of the same drug conditions that were utilized within the
instrumental training paradigm. Tables 1 and 2 demonstrate the effect of drug doses on fooddirected and locomotor activity into the core and shell, respectively. All behavioral measures
were evaluated by repeated-measures ANOVAs performed across treatments, with significance
set at
= .05. Post-hoc comparisons of drug treatments versus saline infusions utilized the
Tukey HSD. Values of each ANOVA are not reported for purposes of simplicity; all statistically
significant differences are denoted in the tables. High dose infusions of scopolamine (but not
mecamylamine) into either the nucleus accumbens core or shell decreased feeding behavior and
increased locomotor activity. Low dose scopolamine reduced food consumption but did not alter
locomotor behavior. Rats showed equal latencies to initiate the first feeding bout in all drug
conditions, and scopolamine-treated rats approached the food at least as often as control animals
(the only significant change was with high dose scopolamine treatment in the core, which
increased approaches to the food). However, total time spent feeding was reduced in the high
dose condition in both shell and core, and scopolamine-treated animals consumed less food than
the control group in both infusion sites. Mean feeding bout length was significantly decreased in
response to high dose scopolamine in the accumbens core (relative to the saline group); there
was a trend for a similar decrease in the shell as well (p < .10). Injections of the 10 g
scopolamine into the nucleus accumbens core increased the time taken to initiate the first
drinking bout, but did not significantly alter the number of drinking bouts or time spent drinking
(data not shown).
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Repeated-measures ANOVAs on the number of center-crossings and rearing events for
each site also showed a significant effect of treatment in both nucleus accumbens sites. Post-hoc
comparisons demonstrated that this effect was due to increases in both of these measures during
intra-accumbens infusions of the high dose scopolamine treatment, but not for the low dose.
There was no effect of mecamylamine treatment on any of the locomotor or feeding measures
utilized here.
Microanalysis of post-training performance under muscarinic blockade
Given that we observed an increase in locomotor activity in the locomotor and feeding
control paradigm, one possible interpretation of these data are that scopolamine grossly impairs
the abilities of rats to perform the required behaviors in the chamber. Although those increases
in activity were limited to the high dose of the drug in the locomotor paradigm, it is possible that
even the low dose of the drug could have resulted in impairments of performance of the intricate
movements required to press the lever and retrieve the sucrose pellet in the instrumental
condition. To address this issue, we further analyzed the raw data from the instrumental learning
experiments (above), and calculated the time between each lever press and nose poke, the time it
took the animal to perform a nose poke (presumably in the process of retrieval of the sugar
pellet) once a reinforcer had been earned, and the time between each nose poke and its
subsequent lever press (see Method for additional details). The median time for each of these
three “dyads” was calculated for each animal in the control, low, and high dose scopolamine on
the final two days of each experiment. The distributions of these medians were subsequently
plotted for the treatment groups in both nucleus accumbens core and shell (Figure 3). Very little
difference was seen in these median distributions, even in the high dose condition, at a time that
the scopolamine-treated rats pressed an average of half as many times within the session.
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The effect of nucleus accumbens core scopolamine treatment in the progressive ratio condition
In order to test possible effects of muscarinic receptor blockade on the motivational value
of the reinforcer, we trained eight animals in a one-hour progressive ratio paradigm (see
Methods). We then examined the effect of increasing doses of nucleus accumbens core
scopolamine on the amount of work a rat would maintain to continue receiving sucrose reward.
The results of this experiment are shown in Figure 4. Repeated measures ANOVAs performed
across treatments demonstrated trends towards significance for break point (F4,28 = 2.69, p =
.051) and total lever presses (F4,28 = 2.54, p = .063). This trend is likely due to the decrease of
responding for the rats on days that they received the two higher doses (5.0 and 10.0 g/side) of
scopolamine. Planned contrasts performed on break point revealed a significant decrease for the
5.0 g scopolamine dose (p < .05), and a similar trend for the 10.0 g dose (p < .10) Contrasts
comparing saline infusions versus the lower doses of scopolamine (0.1 and 1.0 g) were nonsignificant (p >.1)
Discussion
To our knowledge, this report is the first to demonstrate a significant role for nucleus
accumbens cholinergic receptors in appetitive learning. Blockade of nucleus accumbens
muscarinic receptors in the core or shell, but not nicotinic receptors, dose-dependently impaired
instrumental learning. This learning impairment appears to have been the result of a
scopolamine-induced reduction of the rewarding value of the reinforcer, in this case sucrose.
This interpretation is consistent with the overall pattern of observations from these
experiments. As lever pressing was reduced following acquisition in response to high-dose
scopolamine challenge in both the shell and core (as well as the low dose group in the shell), it is
likely that muscarinic blockade mediated neural processes present during both learning and
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performance of this task. Although acute high dose scopolamine treatment in both nucleus
accumbens sites caused increases in locomotion (as measured in the locomotor and feeding
experiments), post-training muscarinic blockade did not increase the rats’ median latencies
between the necessary motor acts required to earn and collect the sucrose reward within the
operant chamber (see Figure 3). Therefore, gross locomotor impairments per se cannot be
argued to be the source of performance effects in the high dose group, and are unlikely to explain
the learning deficits seen in the low dose scopolamine condition (as 1 g caused no change in
locomotion or post-training latencies between behavioral dyads). Additionally, in the locomotor
and feeding experiments, sucrose intake was significantly reduced in response to both high and
low dose scopolamine treatment in the shell and core. Thus, on the short term, hungry animals
reduced their consumption of freely available sucrose pellets while under the influence of
nucleus accumbens muscarinic blockade.
Additionally, we directly tested the effects of muscarinic blockade of the nucleus
accumbens core upon the amount of work animals would undertake to obtain sucrose, as
measured by break point within a progressive ratio paradigm. In contrast to break point
increases that we have observed with nucleus accumbens infusions of amphetamine or opiates
(M. Zhang, Balmadrid, & Kelley, 2003), we found a dose-dependent trend for scopolamine to
reduce break point. This reduction supports the hypothesis that accumbens muscarinic blockade
reduces the reinforcing value of the sucrose. That the trend just missed statistical significance (p
= .051) may be the result of the relative level of overtraining the progressive ratio animals
received prior to drug treatments, as compared to the instrumental learning condition. It is also
possible that the reward-reducing effects of scopolamine would have been more evident had we
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used a reinforcement schedule that elicited more vigorous responding in the vehicle condition; in
this progressive ratio 2 task, rats earned an average of only 16 pellets during vehicle infusion.
In contrast to the effects of muscarinic receptor blockade, 10 g of the nicotinic
antagonist mecamylamine did not impair any of the behaviors measured. Although it is possible
that a higher dose of mecamylamine might have been effective in reducing learning and/or
performance, this dose is high in consideration of the literature; when infused into the ventral
tegmentum, 9 g of mecamylamine consistently increases behavioral measures of withdrawal in
animals with chronic nicotine exposure (Hildebrand et al., 1999). Furthermore, the dose chosen
has previously been shown to impair spatial working memory when infused in similar amounts
and volumes into hippocampus (i.e. Kim & Levin, 1996; Ohno, Yamamoto, & Watanabe, 1993).
Therefore, despite evidence linking nicotinic receptor activation to cellular plasticity in the slice
and dopamine outflow in the striatum (e.g., Partridge et al., 2002; Wonnacott, Kaiser, Mogg,
Soliakov, & Jones, 2000; Zhou, Liang, & Dani, 2001), it is muscarinic receptors that appear to
selectively affect instrumental learning and the value of the sucrose reward in these instrumental
and feeding tasks.
The data presented here are consistent with multiple other studies that have also
suggested a role for accumbens circuitry in motivational processes. For example, manipulations
of dopaminergic, opioid, and GABAergic receptors within nucleus accumbens have been shown
to modulate feeding and motivation for food reward (MacDonald, Billington, & Levine, 2003;
Ragnauth, Znamensky, Moroz, & Bodnar, 2000; Stratford & Kelley, 1997; M. Zhang,
Balmadrid, & Kelley, 2003). Differential concentrations of dopamine and acetylcholine within
the nucleus accumbens may mediate, or vary in relation to, satiety-related processing (Helm,
Rada, & Hoebel, 2003). It has been also been repeatedly demonstrated that tonically active
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neurons (which are believed to be the cholinergic interneurons of striatum) in monkeys respond
to motivationally relevant stimuli (Matsumoto et al., 2001; Ravel et al., 2003; Sardo et al., 2000).
These cells have recently been shown to be capable of changing responses to aversive stimuli
when they are subsequently paired with reward presentation. Specifically, as monkeys learned
that a sound previously associated with an aversive outcome (i.e. an air puff) came to predict a
juice reward, recorded striatal interneurons altered their firing characteristics in a manner
consistent with the presentation of a rewarding (rather than aversive) stimulus (Ravel et al.,
2003). The authors argue that tonically active neurons may mediate the motivational
significance of learned stimuli. Our data are consistent with such an interpretation.
The present findings are also in line with previous reports of learning impairments caused
by cholinergic manipulations in other striatal regions. For instance, behavioral impairments
result from dorsal striatal infusions of cholinergic antagonists during acquisition of appetitive
lever-pressing and aversive avoidance paradigms (see Prado-Alcala, 1985 for review, also
Tikhonravov, Shapovalova, & Dyubkacheva, 1997). More recently, medial dorsal striatal
injections of scopolamine have been shown to impair rats’ flexibility to switch responses when
reward is moved on the t-maze (Ragozzino et al., 2002). Selective lesions of striatal cholinergic
neurons in mice impair animals’ abilities to perform spatial alternation or learn to associate
different tones with the appropriate reward locations on the t-maze (Kitabatake et al., 2003).
Such lesions did not, however, produce deficits on the Morris water maze, in contextual or cued
fear conditioning paradigms, or on the rota-rod motor learning task. Kitabataki and colleagues
suggested that striatal cholinergic mechanisms are important for reward-related tasks. The data
we present here provides the novel insight that selective inhibition of ventral striatal cholinergic
Acetylcholine and appetitive behavior
20
mechanisms is sufficient to alter reward-related learning and performance, and that this effect is
likely the result of reinforcer devaluation.
We have previously shown that dopaminergic or glutamatergic antagonism within the
accumbens core selectively impairs instrumental learning (Kelley et al., 1997; Smith-Roe &
Kelley, 2000). Also, low doses of dopaminergic and glutamatergic antagonists, that do not
produce a deficit when injected alone, block learning when infused together (Smith-Roe &
Kelley, 2000). Interestingly, unlike the feeding effects seen as the result of muscarinic receptor
blockade, dopaminergic and glutamatergic receptor antagonism (with SCH-22390, AP-5, or
both) did not alter either overall food intake or time spent feeding when tested in a similar
locomotor and feeding paradigm to the one used here (Baldo, Sadeghian, Basso, & Kelley, 2002;
Kelley et al., 1997; Smith-Roe & Kelley, 2000). This suggests that nucleus accumbens
cholinergic receptors play a unique, if complementary role, in accumbens-based neural
processing.
The nucleus accumbens has long been argued to provide a means for limbic information
to interface with motor processes (Mogenson, Jones, & Yim, 1980). Environmental contextual
information, which presumably arrives from cortical limbic inputs into the nucleus accumbens,
may come to elicit behavioral repertoires based on the reinforcement history of the context
(Mizumori, Pratt, & Ragozzino, 1999). Dopaminergic signals filter glutamatergic signals from
limbic cortex (e.g., O'Donnell, 2003; Yang & Mogenson, 1984; Yim & Mogenson, 1982), and
may provide a means for predictive reward information (e.g., Schultz, 1998) to affect ongoing
neural activation patterns in the accumbens. The distribution of cholinergic receptors within the
striatum suggests multiple roles for acetylcholine in presynaptically mediating accumbens
glutamate and dopamine outflow (e.g. Contant, Umbriaco, Garcia, Watkins, & Descarries, 1996;
Acetylcholine and appetitive behavior
21
Hersch, Gutekunst, Rees, Heilman, & Levey, 1994; Izzo & Bolam, 1988), and further affects
medium spiny neuron output via postsynaptic mechanisms. Thus, blocking muscarinic receptors
with scopolamine in this experiment may have disrupted normal striatal function on multiple
levels. It is known, for example, that presynaptic M1 receptor activation decreases calciumdependent glutamate release, while increasing dopaminergic output (De Klippel, Sarre, Ebinger,
& Michotte, 1993; Rawls & McGinty, 1998; Rawls, McGinty, & Terrian, 1999; Xu, Mizobe,
Yamamoto, & Kato, 1989). Although scopolamine affects all muscarinic receptor subtypes,
blockade of M1 receptors in our experiments may have impaired normal accumbens function by
effectively reversing normal dopaminergic filtering of cortical inputs. Furthermore, a possible
reduction of dopaminergic outflow during the performance test could have resulted in decreased
lever-pressing, as striatal treatment with dopaminergic antagonists has been shown to diminish
responding for food reward (Beninger & Ranaldi, 1993; Smith-Roe & Kelley, 2000).
Postsynaptically, M1 receptor activation decreases current through NMDA channels on
medium spiny neurons, and muscarinic receptor antagonism (particularly of the M1 subtype)
reduces plasticity within the striatal slice preparation (Calabresi, Centonze, Gubellini, &
Bernardi, 1999; Calabresi et al., 1998). Our data do not preclude the possibility that blockade of
these receptors in vivo may have directly impaired plasticity during the acquisition phase of this
experiment. M2 receptors are found chiefly on the cholinergic interneurons themselves, and
have been argued to autoregulate cholinergic output (Hersch et al., 1994, but see W. Zhang,
Yamada, Gomeza, Basile & Wess, 2002). Less is known about the physiological role of the
remaining muscarinic receptor subtypes in the modulation of striatal transmitters and plasticity,
although recent evidence suggests that M3 and M5 receptors also mediate dopaminergic actions
Acetylcholine and appetitive behavior
22
within striatum (W. Zhang et al., 2002). Future studies will delineate the effects of specific
acetylcholine receptor blockade within the experimental paradigms utilized here.
Given the known interactions between acetylcholine and other striatal components, it is
surprising that there has not been more research into the role of nucleus accumbens acetylcholine
in mediating learning and behavior. Recent experiments have made it clear that both cortical and
striatal acetylcholine serves an important role in learning and memory processes. Relative levels
of cholinergic activation between regions may serve as an important indicator of task
involvement, as it has been recently argued that cholinergic levels in hippocampus and dorsal
striatum reflect strategies selected by rats on the T-maze (Chang & Gold, 2003; McIntyre,
Marriott, & Gold, 2003). However, much of the cortex receives cholinergic innervation from the
basal forebrain regions, whereas the striatum’s cholinergic influence arises predominantly from
its own interneurons. Understanding the influence of these neurons in striatal function is
imperative to the understanding of basal ganglia functions involved in cognitive as well as
movement processes. The experiments reported here suggest that muscarinic acetylcholine
receptors within the nucleus accumbens shell and core are important for plasticity underlying
appetitive instrumental learning, most likely by modulating the motivational value of the
reinforcer.
Acetylcholine and appetitive behavior
23
References
Arnold, J. M., & Roberts, D. C. (1997). A critique of fixed and progressive ratio schedules used
to examine the neural substrates of drug reinforcement. Pharmacology, Biochemistry,
and Behavior, 57(3), 441-447.
Baldo, B. A., Sadeghian, K., Basso, A. M., & Kelley, A. E. (2002). Effects of selective dopamine
D1 or D2 receptor blockade within nucleus accumbens subregions on ingestive behavior
and associated motor activity. Behav Brain Res, 137(1-2), 165-177.
Beninger, R. J., & Ranaldi, R. (1993). Microinjections of flupenthixol into the caudate-putamen
but not the nucleus accumbens, amygdala or frontal cortex of rats produce intra-session
declines in food-rewarded operant responding. Behavioural Brain Research, 55(2), 203212.
Calabresi, P., Centonze, D., Gubellini, P., & Bernardi, G. (1999). Activation of M1-like
muscarinic receptors is required for the induction of corticostriatal LTP.
Neuropharmacology, 38(2), 323-326.
Calabresi, P., Centonze, D., Gubellini, P., Pisani, A., & Bernardi, G. (1998). Endogenous ACh
enhances striatal NMDA-responses via M1-like muscarinic receptors and PKC activation.
European Journal of Neuroscience, 10(9), 2887-2895.
Centonze, D., Gubellini, P., Bernardi, G., & Calabresi, P. (1999). Permissive role of interneurons
in corticostriatal synaptic plasticity. Brain Research Reviews, 31(1), 1-5.
Chang, Q., & Gold, P. E. (2003). Switching memory systems during learning: changes in
patterns of brain acetylcholine release in the hippocampus and striatum in rats. Journal of
Neuroscience, 23(7), 3001-3005.
Acetylcholine and appetitive behavior
24
Contant, C., Umbriaco, D., Garcia, S., Watkins, K. C., & Descarries, L. (1996). Ultrastructural
characterization of the acetylcholine innervation in adult rat neostriatum. Neuroscience,
71(4), 937-947.
De Klippel, N., Sarre, S., Ebinger, G., & Michotte, Y. (1993). Effect of M1- and M2-muscarinic
drugs on striatal dopamine release and metabolism: an in vivo microdialysis study
comparing normal and 6-hydroxydopamine-lesioned rats. Brain Research, 630(1-2), 5764.
Graveland, G. A., & DiFiglia, M. (1985). The frequency and distribution of medium-sized
neurons with indented nuclei in the primate and rodent neostriatum. Brain Research,
327(1-2), 307-311.
Heimer, L., Zahm, D. S., Churchill, L., Kalivas, P. W., & Wohltmann, C. (1991). Specificity in
the projection patterns of accumbal core and shell in the rat. Neuroscience, 41(1), 89-125.
Helm, K. A., Rada, P., & Hoebel, B. G. (2003). Cholecystokinin combined with serotonin in the
hypothalamus limits accumbens dopamine release while increasing acetylcholine: a
possible satiation mechanism. Brain Research, 963(1-2), 290-297.
Hersch, S. M., Gutekunst, C. A., Rees, H. D., Heilman, C. J., & Levey, A. I. (1994). Distribution
of m1-m4 muscarinic receptor proteins in the rat striatum: light and electron microscopic
immunocytochemistry using subtype-specific antibodies. Journal of Neuroscience, 14(5
Pt 2), 3351-3363.
Hildebrand, B. E., Panagis, G., Svensson, T. H., & Nomikos, G. G. (1999). Behavioral and
biochemical manifestations of mecamylamine-precipitated nicotine withdrawel in the rat:
Role of nicotinic receptors in the ventral tegmental area. Neuropsychopharmacology,
21(4), 560-574.
Acetylcholine and appetitive behavior
25
Hodos, W. (1961). Progressive ratio as a measure of reward strength. Science, 134, 943-944.
Izzo, P. N., & Bolam, J. P. (1988). Cholinergic synaptic input to different parts of spiny
striatonigral neurons in the rat. Journal of Comparative Neurology, 269(2), 219-234.
Kelley, A. E. (1999). Functional specificity of ventral striatal compartments in appetitive
behaviors. Annals of the New York Academy of the Sciences, 877, 71-90.
Kelley, A. E., Smith-Roe, S. L., & Holahan, M. R. (1997). Response-reinforcement learning is
dependent on N-methyl-D-aspartate receptor activation in the nucleus accumbens core.
Procedings of the National Academy of Sciences of the United States of America, 94(22),
12174-12179.
Kim, J. S., & Levin, E. D. (1996). Nicotinic, muscarinic and dopaminergic actions in the ventral
hippocampus and the nucleus accumbens: effects on spatial working memory in rats.
Brain Research, 725(2), 231-240.
Kitabatake, Y., Hikida, T., Watanabe, D., Pastan, I., & Nakanishi, S. (2003). Impairment of
reward-related learning by cholinergic cell ablation in the striatum. Procedings of the
National Academy of Sciences of the United States of America, 100(13), 7965-7970.
Koob, G. F. (1992). Neural mechanisms of drug reinforcement. Annals of the New York Academy
of the Sciences, 654, 171-191.
MacDonald, A. F., Billington, C. J., & Levine, A. S. (2003). Effects of the opioid antagonist
naltrexone on feeding induced by DAMGO in the ventral tegmental area and in the
nucleus accumbens shell region in the rat. American Journal of Physiology. Regulatory,
Integrative, and Comparative Physiology, 285(5), R999-R1004.
Acetylcholine and appetitive behavior
26
Matsumoto, N., Minamimoto, T., Graybiel, A. M., & Kimura, M. (2001). Neurons in the
thalamic CM-Pf complex supply striatal neurons with information about behaviorally
significant sensory events. Journal of Neurophysiology, 85(2), 960-976.
McGeorge, A. J., & Faull, R. L. (1989). The organization of the projection from the cerebral
cortex to the striatum in the rat. Neuroscience, 29(3), 503-537.
McIntyre, C. K., Marriott, L. K., & Gold, P. E. (2003). Patterns of brain acetylcholine release
predict individual differences in preferred learning strategies in rats. Neurobiology of
Learning and Memory, 79(2), 177-183.
Mizumori, S. J. Y., Pratt, W. E., & Ragozzino, K. E. (1999). Function of the nucleus accumbens
within the context of the larger striatal system. Psychobiology, 27(2), 214-224.
Mogenson, G. J., Jones, D. L., & Yim, C. Y. (1980). From motivation to action: functional
interface between the limbic system and the motor system. Progress in Neurobiology,
14(2-3), 69-97.
Nauta, W. J., & Domesick, V. B. (1984). Afferent and efferent relationships of the basal ganglia.
CIBA Foundation Symposium, 107, 3-29.
O'Donnell, P. (2003). Dopamine gating of forebrain neural ensembles. European Journal of
Neuroscience, 17(3), 429-435.
Ohno, M., Yamamoto, T., & Watanabe, S. (1993). Blockade of hippocampal nicotinic receptors
impairs working memory but not reference memory in rats. Pharmacology, Biochemistry,
and Behavior, 45(1), 89-93.
Partridge, J. G., Apparsundaram, S., Gerhardt, G. A., Ronesi, J., & Lovinger, D. M. (2002).
Nicotinic acetylcholine receptors interact with dopamine in induction of striatal long-term
depression. J Neurosci, 22(7), 2541-2549.
Acetylcholine and appetitive behavior
27
Paxinos, G., & Watson, C. (1998). The rat brain in sterotaxic coordinates (4th ed). San Diego,
CA: Academic Press.
Prado-Alcala, R. A. (1985). Is cholinergic activity of the caudate nucleus involved in memory?
Life Sciences, 37(23), 2135-2142.
Ragnauth, A., Znamensky, V., Moroz, M., & Bodnar, R. J. (2000). Analysis of dopamine
receptor antagonism upon feeding elicited by mu and delta opioid agonists in the shell
region of the nucleus accumbens. Brain Research, 877(1), 65-72.
Ragozzino, M. E., Jih, J., & Tzavos, A. (2002). Involvement of dorsomedial striatum in
behavioral flexibility: Role of muscarinic cholinergic receptors. Brain Research, 953,
205-214.
Ravel, S., Legallet, E., & Apicella, P. (2003). Responses of tonically active neurons in the
monkey striatum discriminate between motivationally opposing stimuli. Journal of
Neuroscience, 23(24), 8489-8497.
Rawls, S. M., & McGinty, J. F. (1998). Muscarinic receptors regulate extracellular glutamate
levels in the rat striatum: an in vivo microdialysis study. Journal of Pharmacology and
Experimental Therapeutics, 286(1), 91-98.
Rawls, S. M., McGinty, J. F., & Terrian, D. M. (1999). Presynaptic kappa-opioid and muscarinic
receptors inhibit the calcium-dependent component of evoked glutamate release from
striatal synaptosomes. Journal of Neurochemistry, 73(3), 1058-1065.
Sardo, P., Ravel, S., Legallet, E., & Apicella, P. (2000). Influence of the predicted time of stimuli
eliciting movements on responses of tonically active neurons in the monkey striatum.
European Journal of Neuroscience, 12(5), 1801-1816.
Acetylcholine and appetitive behavior
28
Schultz, W. (1998). Predictive reward signal of dopamine neurons. Journal of Neurophysiology,
80(1), 1-27.
Smith-Roe, S. L., & Kelley, A. E. (2000). Coincident activation of NMDA and dopamine D1
receptors within the nucleus accumbens core is required for appetitive instrumental
learning. Journal of Neuroscience, 20(20), 7737-7742.
Stratford, T. R., & Kelley, A. E. (1997). GABA in the nucleus accumbens shell participates in
the central regulation of feeding behavior. Journal of Neuroscience, 17(11), 4434-4440.
Tikhonravov, D. L., Shapovalova, K. B., & Dyubkacheva, T. A. (1997). Effects of
microinjection of scopolamine into the neostriatum of rats on performance of a food
conditioned reflex at different levels of fixation. Neuroscience and Behavioral
Physiology, 27(3), 312-318.
Wonnacott, S., Kaiser, S., Mogg, A., Soliakov, L., & Jones, I. W. (2000). Presynaptic nicotinic
receptors modulating dopamine release in the rat striatum. European Journal of
Pharmacology, 393(1-3), 51-58.
Xu, M., Mizobe, F., Yamamoto, T., & Kato, T. (1989). Differential effects of M1- and M2muscarinic drugs on striatal dopamine release and metabolism in freely moving rats.
Brain Research, 495(2), 232-242.
Yang, C. R., & Mogenson, G. J. (1984). Electrophysiological responses of neurones in the
nucleus accumbens to hippocampal stimulation and the attenuation of the excitatory
responses by the mesolimbic dopaminergic system. Brain Research, 324(1), 69-84.
Yim, C. Y., & Mogenson, G. J. (1982). Response of nucleus accumbens neurons to amygdala
stimulation and its modification by dopamine. Brain Research, 239(2), 401-415.
Acetylcholine and appetitive behavior
29
Zhang, M., Balmadrid, C., & Kelley, A. E. (2003). Nucleus accumbens opiod, GABAergic, and
dopaminergic modulation of palatable food motivation: Contrasting effects revealed by a
progressive ratio study in the rat. Behavioral Neuroscience, 117(2), 202-211.
Zhang, W., Yamada, M., Gomeza, J., Basile, A. S., & Wess, J. (2002). Multiple muscarinic
acetylcholine receptor subtypes modulate striatal dopamine release, as studied with M1M5 muscarinic receptor knock-out mice. Journal of Neuroscience, 22(15), 6347-6352.
Zhou, F. M., Liang, Y., & Dani, J. A. (2001). Endogenous nicotinic cholinergic activity regulates
dopamine release in the striatum. Nature Neuroscience, 4(12), 1224-1229.
Acetylcholine and appetitive behavior
30
Author note
This work was supported by NIDA grant DA04788. We would like to express our thanks
to Ken Sadeghian and Drs. Matt Andrzejewski and Brian Baldo for technical assistance during
various phases of these projects.
Acetylcholine and appetitive behavior
Table 1. Influence of nucleus accumbens core infusions on feeding and locomotion.
Behavioral measure
Saline
Mecamylamine
1 g Scopolamine
10 g Scopolamine
Latency to feed (s)
12.2 +/- 2.1
13.6 +/- 3.0
14.4 +/- 2.3
27.9 +/- 10.3
Number of feeding bouts 27.9 +/- 3.6
25.8 +/- 2.6
34.4 +/- 6.3
39.4 +/- 3.7 *
Mean bout length (s)
25.5 +/- 3.2
23.7 +/- 3.4
18.2 +/- 3.3
7.7 +/- 3.0 **
Total time feeding (s)
606.4 +/- 30.8
568.7 +/- 32.1
488.4 +/- 27.2
245.4 +/- 57.4 **
Sucrose intake (g)
5.1 +/- 0.3
4.2 +/- 0.3
3.7 +/- 0.6 *
1.0 +/- 0.3 **
Total center crossings
31.1 +/- 3.5
36.0 +/- 4.5
46.13 +/- 10.3
127.9 +/- 18.6 **
Total rearing events
31.9 +/- 3.7
36.0 +/- 3.7
37.1 +/- 6.6
131.9 +/- 24.8 **
Time spent rearing (s)
64.7 +/- 9.7
63.5 +/- 11.0
69.6 +/- 10.5
172.3 +/- 31.9 **
Note. Means +/- SEM for infusions into the nucleus accumbens core (N = 8). *p < .05; ** p < .01
31
Acetylcholine and appetitive behavior
Table 2. Influence of nucleus accumbens shell infusions on feeding and locomotion.
Behavioral measure
Saline
Mecamylamine
1 g Scopolamine
10 g Scopolamine
Latency to feed (s)
10.3 +/- 2.6
15.9 +/- 3.9
12.3 +/- 2.2
17.9 +/- 2.0
Number of feeding bouts 24.3 +/- 3.4
21.7 +/- 4.2
25.2 +/- 3.9
22.5 +/- 2.3
Mean bout length (s)
25.5 +/- 5.4
34.9 +/- 7.9
19.7 +/- 3.3
15.8 +/- 5.4
Total time feeding (s)
565.1 +/- 69.0
607.9 +/- 42.4
452.3 +/- 53.5
317.7 +/- 81.4 *
Sucrose intake (g)
5.9 +/- 0.5
6.4 +/- 0.6
3.2 +/- 0.7 **
2.1 +/- 0.5 **
Total center crossings
28.0 +/- 3.6
19.0 +/- 2.0
38.8 +/- 4.7
68.7 +/- 20.3 **
Total rearing events
31.2 +/- 5.8
19.8 +/- 3.2
34.0 +/- 3.5
63.3 +/- 15.4 *
Time spent rearing (s)
69.5 +/- 22.1
54.2 +/- 12.4
78.3 +/- 13.5
97.25 +/- 22.5
Note. Numbers +/- SEM for infusions into the nucleus accumbens shell (N = 6). *p < .05; ** p < .01
32
Acetylcholine and appetitive behavior
33
Figure Captions
Figure 1. Representative histology for these experiments. The left panel shows nucleus
accumbens core placements for the saline (white triangle), low dose (grey triangle), and high
dose (black triangle) scopolamine groups in the instrumental learning paradigm. The center
panel demonstrates the position of shell placements for eight saline (white triangle),
mecamylamine (grey circle), and high dose scopolamine (black triangle) animals in the same
condition. The two photomicrographs in the right column are representative placements in the
core (top) and shell (bottom) of the nucleus accumbens.
Figure 2. Effects of muscarinic or nicotinic antagonism of the nucleus accumbens core (A) and
shell (B) on the learning and performance of lever-pressing for sucrose reward. Drug infusions
preceeded the first five training sessions, as well as the final drug session. Muscarinic receptor
blockade with 1.0 and 10.0 g scopolamine impaired acquisition of the instrumental response;
nicotinic antagonism by 10 g mecamylamine was without effect. Lever pressing also decreased
after learning in response to high dose scopolamine treatment in the core, as well as both high
and low dose treatments in the shell. Numbers in parentheses () denote animals included in each
group; the control group for both shell treatments consisted of the same 15 animals. * p < .05,
treatment effect, ** p < .01, treatment effect, ‡, p < .01, interaction of treatment x day.
Figure 3. Box plots representing group distributions of the median time required to perform
three behavioral dyads within the instrumental chamber on the final two days of the instrumental
learning experiments. White boxes demonstrate the distribution of behavioral latencies between
behaviors on the final day of training, after which all groups had reached asymptotic
performance. Grey boxes show latencies on the following day, after animals have received a
final treatment of scopolamine. Notice that the drug treatments do not appear to increase the
Acetylcholine and appetitive behavior
34
time animals take to complete these behavioral dyads, suggesting that the decreases in
responding on the final experimental day was not the result of gross impairments in the animals’
abilities to perform the task.
Figure 4. Effects of nucleus accumbens core muscarinic blockade on break point and lever
pressing within the progressive ratio paradigm. Increasing doses of scopolamine show a trend
toward decreasing break point and total lever presses for the session. Post-hoc contrasts showed
a significant reduction of break point at the 5 g dose. These data suggest a role for muscarinic
acetylcholinergic mechanisms in mediating the reinforcing value of the sucrose.
Acetylcholine and appetitive behavior
35
Pratt & Kelley
Figure 1
Acetylcholine and appetitive behavior
36
Pratt & Kelley
Figure 2
Acetylcholine and appetitive behavior
37
Pratt & Kelley; Figure 3
Acetylcholine and appetitive behavior
38
Pratt & Kelley
Figure 4