Download The subthalamic nucleus exerts opposite control on cocaine and

© 2005 Nature Publishing Group http://www.nature.com/natureneuroscience
ARTICLES
The subthalamic nucleus exerts opposite control
on cocaine and ‘natural’ rewards
Christelle Baunez1, Carine Dias2, Martine Cador2 & Marianne Amalric1
A challenge in treating drug addicts is preventing their pathological motivation for the drug without impairing their general
affective state toward natural reinforcers. Here we have shown that discrete lesions of the subthalamic nucleus greatly decreased
the motivation of rats for cocaine while increasing it for food reward. The subthalamic nucleus, a key structure controlling basal
ganglia outputs, is therefore able to oppositely modulate the effect of ‘natural’ rewards and drugs of abuse on behavior.
Modulating the activity of the subthalamic nucleus might prove to be a new target for the treatment of cocaine addiction.
Drug dependence is a major worldwide public health problem. In drug
addicts, one of the main unsolved challenges is eradicating the
pathological motivation for the drug without impairing the general
affective state of subjects and their motivation for ‘natural’ reinforcers.
As drugs of abuse and natural reinforcers mostly recruit common
biological substrates, the possibility of obtaining such dissociation
remains an open issue.
Structures of the basal ganglia classically considered motor structures
can be involved in coding reward1,2 and in the reinforcing effects of
cocaine3,4. Among these structures, the subthalamic nucleus (STN) has
received much attention as a therapeutic target because its inactivation
by high-frequency stimulation has been used successfully for the
treatment of Parkinson disease5. Although inactivation of the STN is
used mainly to improve motor deficits5–9, preliminary experimental
and clinical observations have suggested that this structure
may also have considerable involvement in the modulation of motivation10–15. Accordingly, this structure seems to be sensitive to drugs of
abuse, as repeated injections of cocaine result in a decreased metabolic
activity in the STN16,17.
To address whether the STN could differentially affect motivation for
conventional (‘natural’) or pharmacological (‘drug’) reinforcers, we
have studied the effects of excitotoxic lesions of this nucleus, which
have effects functionally similar to high-frequency stimulation7,18. We
analyzed motivation for food and cocaine rewards using two complementary behavioral approaches: an ‘operant paradigm’ (progressive
ratio) that measures the ability of the animals to provide an increased
rate of responding to obtain food or drugs, and a ‘place-conditioning
schedule’ that measures preference or aversion for an environment
specifically paired with food or cocaine.
The combined results of these two approaches indicate that excitotoxic lesions of the STN increase the motivation of rats for a food
reward while greatly decreasing the motivation for cocaine. These data
demonstrate that motivation for natural rewards and drugs of abuse
can be oppositely modulated in part via the STN. This possibility
indicates the STN may be a potential target for developing specific and
efficient treatments for drug abusers.
RESULTS
Histology
Only rats with a bilateral lesion of the STN and an injection site in the
STN were included in the data analysis. As in previous studies8,15, the
bilateral STN lesion selectively damaged all sectors of the STN
bilaterally, sparing only a few cells in the most lateral portions of the
nucleus (Fig. 1). We found no damage in the lateral hypothalamus. We
eliminated from the behavioral data analyses eight rats with lesions
above the nucleus. The location of the injection cannula was in the STN
in four rats and in three it was located outside the STN. The
performance of the eliminated rats was similar to that of control rats
(‘sham-lesioned’ rats), thus confirming that the behavioral effects
noted were specifically due to STN lesions or infusions in the STN.
Behavioral results
To assess the effects of STN lesions on food or cocaine intake and to
ensure that STN-lesioned rats could acquire drug self-administration,
we first tested the rats with a continuous reinforcement schedule (fixed
ratio 1), in which each lever press was followed by the reward. Access to
the reward was thus fairly easy and required a minimum of work. In
this task, whatever the reward (a sucrose pellet or an intravenous
cocaine injection), STN-lesioned rats obtained as many rewards as the
sham-lesioned control rats did (approximately 100 pellets per session
and 25 cocaine injections per session; P 4 0.05; Fig. 2a,b). In both
procedures, with food and with cocaine reinforcement, there was no
significant difference between the sham-lesioned and STN-lesioned
groups in the rate of lever presses (P 4 0.05; Fig. 2c,d). The typical
response records showed that both the sham-lesioned and STNlesioned groups had a stable pattern of evenly spaced responses on
1Laboratoire de Neurobiologie de la Cognition, Unité Mixte de Recherche (UMR) 6155 Centre National de la Recherche Scientifique Université de Provence, 13402
Marseille Cedex 20, France. 2Laboratoire de Neuropsychobiologie des Désadaptations, UMR5541 Centre National de la Recherche Scientifique, Université de Bordeaux 2,
33076 Bordeaux Cedex, France. Correspondence should be addressed to C.B. ([email protected]).
Published online 27 March 2005; doi:10.1038/nn1429
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Figure 1 Frontal sections at the level of the STN, stained with cresyl violet.
Dashed lines outline the STN in a sham-lesioned rat (a) and an STN-lesioned
rat (b). Inset, coronal section of the rat brain at the level of the subthalamic
nucleus, from ref. 49. CP, cerebral peduncle.
the active lever with a short delay before the first active injection in
STN-lesioned rats (Fig. 2e). As all rats reached similar levels of intake
with the same rate during acquisition (data not shown), the primary
reinforcing properties of food and cocaine seemed to be identical in
both sham-lesioned and STN-lesioned rats.
As both groups worked similarly for both rewards when they could
easily be obtained, the next issue was to determine whether STN lesions
could differentially affect the behavior of the rats in a highly demanding
task such as those with progressive ratio schedules of reinforcement, in
which the work demand required to obtain the reward is progressively
increased. When no difference is found in a continuous reinforcement
schedule, a difference can be unmasked using a progressive ratio19,20.
With progressive ratio schedules of reinforcement, the measure of
the last ratio reached (also called the ‘breaking point’) allows assessment of the amount of effort a rat is willing to expend to obtain the
reinforcer21. With this schedule, rats with bilateral STN lesions reached
a higher breaking point than did sham-lesioned control rats and thus
obtained a significantly higher number of reinforcers when working for
one or two sucrose pellets (group effect: F1,13 ¼ 6.07 and 7.05, P o 0.05;
Fig. 3a). We found a similar effect after intra-STN infusion of
muscimol (3 ng/side) in rats working for one sucrose pellet (drug
effect: F2,6 ¼ 20.04, P o 0.01; Fig. 3b). In contrast, when the reinforcer
was a single intravenous injection of cocaine (250 mg/injection),
Figure 2 Effects of bilateral STN lesions on continuous reinforcement
tasks for food and cocaine. Each lever press is reinforced by one sucrose
pellet (a,c) or by an intravenous injection of 250 mg cocaine (b,d). Each dot
represents the mean number of pellets or injections during the 30-minute
(a) or 120-minute (b) session (7 s.e.m.) for the last five daily sessions with
this schedule (abscissa) or the mean rate of lever presses during the same
sessions (c,d). Data are for sham-lesioned control rats (n ¼ 8 (a,c) and
n ¼ 9 (b,d)) and for STN-lesioned lesion rats (n ¼ 7 (a,c) and n ¼ 9 (b,d)).
(e) Kinetics of cocaine injections for a representative 2-hour session using
the ‘fixed ratio 1’ schedule for one representative rat of each group. Each bar
represents a cocaine infusion.
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STN-lesioned rats worked less than sham-lesioned rats. They reached
a lower breaking point and gained a lower number of injections (group
effect: F1,16 ¼ 8.69, P o 0.01) (Fig. 3c). The amount of locomotor
activity recorded during the sessions for cocaine self-administration
was equivalent in both groups (P 4 0.05; data not shown), ruling out
the possibility of a motor effect of cocaine at this dose on the
performance of the rats.
As STN-lesioned rats showed decreased motivation to obtain one
cocaine injection when the work load was increased, we then progressively decreased the cocaine dose in the continuous reinforcement
schedule. In these conditions, we were able to evaluate any shift in the
sensitivity of the rats to the drug and how much they were able to work
to adjust the concentration of cocaine in their blood circulation. Each
dose was available for a 1-h period. As classically described, the first
10 min correspond to the ‘loading period’ during which the animals
adjust to the dose available. When the doses of cocaine were decreased,
sham-lesioned rats consequently increased their number of injections as
a function of the dose during these first 10 min (dose effect: F3,15 ¼ 30.93,
P o 0.01; Fig. 4a). The STN-lesioned group showed a poorer
adaptation to the lower doses of cocaine and obtained significantly
fewer injections than control rats (group dose interaction: F3,30 ¼ 5.47,
P o 0.01) during the first 10 min of the schedule, as demonstrated by
the response pattern (Fig. 4c). However, when given the 1-h period of
access, STN-lesioned rats were able to adapt their response
to the diminished effect of cocaine reward value in the long term, as
the total number of injections obtained during the session was not
significantly different for the sham-lesioned and STN-lesioned groups
(P 4 0.05; Fig. 4b). STN-lesioned rats clearly showed a slower rate of
injection for each dose except the standard one (250 mg), especially
during the first minutes (Fig. 4c). The fact that there was no difference
between the groups at this last dose rules out the possibility of a motor
impairment produced by the lesion.
To assess whether the reinforcing property of food or cocaine was
modified by the STN lesions, we used the place-conditioning paradigm
to quantify their reward efficacy. In this paradigm, the reward effects of
a given reinforcer (food or cocaine) are inferred by measurement of
the time spent in a specific environment previously paired with the
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Figure 3 Effects of bilateral STN lesions on performance in the progressive
ratio task for food and cocaine. The willingness of the rats to work for the
reward is evaluated by the mean number (7 s.e.m.) of reinforcers acquired
during each session (averaged over five sessions). (a) Mean number of pellets
obtained with one or two sucrose pellets as a reinforcer. (b) Mean number
of pellets obtained with one sucrose pellet as a reinforcer after infusion of
saline (0) or muscimol (2 or 3 ng per 0.5 ml) into the STN (n ¼ 4). (c) Mean
number of intravenous cocaine injections obtained (250 mg/injection).
Sham-lesioned control rats, n ¼ 8 (a) and n ¼ 9 (c); STN-lesioned lesion
rats, n ¼ 7 (a) and n ¼ 9 (c). *, P o 0.05, compared with the
sham-lesioned group. **, P o 0.01.
reinforcer in comparison with time spent in another environment
paired with no food or with vehicle injection. In a ‘choice’ situation and
in the absence of the reinforcer, measurement of the time spent in these
environments gives an indication of the positive or negative affective
memory that the animal has of its food or cocaine experience. Both
groups showed a preference for the environment associated with food,
with the STN-lesioned group showing a stronger preference than the
sham-lesioned group (conditioning effect: P o 0.05 (sign test); group
effect: P o 0.05 (Mann-Whitney test); Fig. 5a). In contrast,
after conditioning with two different doses of cocaine (5 mg/kg and
10 mg/kg; Fig. 5b), the sham-lesioned group showed a clear preference
for the compartment associated with cocaine whatever the dose, but
the STN-lesioned group showed a weaker preference for the compartment associated with cocaine (conditioning effect: P o 0.05 for
both doses (sign test); group effect: Po 0.01 for the 10-mg/kg dose
(Mann-Whitney test)).
DISCUSSION
In this study we have presented converging evidence showing that
bilateral lesions of the STN have opposite effects on motivation for
food and cocaine. Rats with excitotoxic lesions of the STN did indeed
work harder for a food reward but less for cocaine. In parallel, STN
lesions enhanced the preference for an environment previously associated with food while decreasing the preference for a cocaineassociated environment.
Reversible inactivation of the STN by the GABAA receptor agonist
muscimol increased the progressive ratio performance for food reinforcement to an extent similar to that of the STN excitotoxic lesion,
thus confirming the specificity of the STN site itself in mediating the
behavioral effects seen here. Either a muscimol infusion or a permanent lesion of the STN disrupts the performance of an attentional task,
emphasizing the critical involvement of the STN in mediating reward
information and attentional processes22,23.
Our results confirm involvement of the STN in food reinforcement
and further suggest that the effects of STN lesions on motivation are
reinforcer dependent. They extend previous findings15 showing that in
addition to its classical control of motor output, the STN is associated
with motivational associative processes. The effects produced by STN
lesions cannot be related to a nonspecific motor impairment, as we
found no motor disinhibition (reflecting impulsive behavior)15 or
changes in cocaine-induced locomotion in the STN-lesioned rats.
STN lesions usually produce hyperkinetic-like disorders (such as
ballism in primates) rather than motor depression and cannot therefore account for the lower number of lever presses for cocaine with the
progressive ratio schedule8,24,25.
As found here, a dissociation between natural and drug reinforcement has been demonstrated recently by electrophysiological studies.
Selective neuronal subpopulation of the nucleus accumbens (ventral
striatum) recorded in freely moving rats were found to be differentially
activated by natural reward (water and food) versus cocaine reinforcement26,27. In addition, the activity of ventral striatal neurons was
notably different for cocaine and juice reward in monkeys performing
a reaction-time task28. These findings have important functional
implications, as they suggest that different parallel microcircuits
mediate responses for natural reward versus cocaine. As the nucleus
accumbens and the STN participate in a cortico-basal ganglia-cortical
‘limbic loop’ encompassing the prefrontal cortex, nucleus accumbens,
ventral pallidum area and STN29,30, which is analogous to the ‘motor
loop’ involving the dorsal striatum31, these microcircuits are likely to be
represented at each level of the loop. Specifically, the core region of the
nucleus accumbens projects to the ventral pallidal area that projects to
the medial part of the STN. There are successive GABAergic connections from the core region to the STN (via the ventral pallidum).
Reciprocally, by its glutamatergic projections to the ventral pallidum32,
the STN controls limbic information outflow33. Therefore, via this
connection, STN may exert an opposite effect on these microcircuits,
exerting an inhibitory control on the ‘natural reinforcer circuit’ while
activating the ‘cocaine circuit’. This interpretation is consistent with the
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Figure 4 Effect of a change in dose during continuous reinforcement for
cocaine self-administration. (a,b) The mean number of injections (7 s.e.m.)
during the first 10 min (a) and the whole 60-minute session (b) of selfadministration for each dose for the sham-lesioned group (n ¼ 6) and the
STN-lesioned group (n ¼ 6). *, P o 0.05, compared with the sham-lesioned
group; #, P o 0.05 and **, P o 0.01, compared with the 250-mg dose.
(c) Representative injection patterns of representative rats from the shamlesioned and STN-lesioned groups for each dose of cocaine during a
1-hour session.
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Figure 5 Effects of bilateral STN lesions on conditioned place preference for
food or cocaine. The preference score (7 s.e.m.) is presented as the time
spent in the compartment associated with food (a) or cocaine (5 mg/kg and
10 mg/kg intraperitoneally; b) on the test day minus the time spent in this
compartment on the preconditioning day for sham-lesioned rats (n ¼ 6, 8
and 6, respectively) and STN-lesioned rats (n ¼ 6, 5 and 7, respectively).
*, P r 0.05, and **, P r 0.01, compared with the sham-lesioned group.
fact that via its connections to the various subloops of the topographically organized cortico–basal ganglia conveying motor, cognitive
and limbic information, the STN lesion differentially affects goaldirected behavior for natural or drug reward.
It is unlikely that STN lesions affect stimulus-reward association
processes. STN lesions do not disrupt the association between a
conditioned stimulus and a food reward15. In our experiments here,
the fact that STN-lesioned rats showed a place preference indicates an
intact association between the reinforcer and the environment. In
contrast, some of our results presented here indicate that the STN
can differentially modify the salience of the reward. This was indicated
by the increased willingness of STN-lesioned rats to work for food
reward with the progressive ratio schedule and by the vertical downward shift in the dose-response curve for cocaine. Such a shift in
cocaine dose-response function reflects a change in reward efficacy,
such as that seen in low-responder rats compared with high responders
to novelty34 or for rats with short versus long access to cocaine35 or
after lesioning of the sublenticular region of the extended amygdala,
which includes part of the shell of the accumbens36. In the conditioned
place-preference paradigm, the resulting preference-aversion provides
an indirect measure of the incentive salience (euphoria-dysphoria)
produced by a drug. In parallel, protocols such as the progressive ratio
schedule allow measurement of the motivational value of a reinforcer.
Our study here using these paradigms has indicated that STN lesions
increase the salience of food and decrease that of cocaine. Modulation
of the salience and the reinforcing and response-invigorating effects of
natural reward or psychostimulant drugs is often attributed to the
nucleus accumbens shell area37–40. The effects of STN lesions on the
salience of the reward could thus be explained by its connections with
the nucleus accumbens, within the limbic loop. Selective areas of this
loop including the ventral striatum and the STN are indeed involved in
the prediction and evaluation of reward, as has been found in studies of
humans using functional magnetic resonance imaging41.
The effects noted after STN lesions could also reflect a loss of the
direct prefrontal influence on the STN. The prelimbic–medial orbital
areas of the prefrontal cortex are indeed connected to the medial part of
the STN30, and this specific region is involved in different aspects of
reward processing, such as reward value, in nonhuman primates42,43.
Furthermore, imaging studies of cocaine addicts have also emphasized dysfunctions in these regions44,45. As addiction is increasingly
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regarded as a compulsive disorder generating inappropriate responses and involving the prefrontal cortex46, it might well be that the
orbitofrontal-subthalamic connection is critical in regulating of
the faulty decision-making associated with addictive behavior.
Accordingly, direct evidence of functional involvement of this corticosubthalamic connection has been found in attentional and perserverative processes in rats47. Our results have thus important clinical
implications, as clinical evidence has shown that STN inactivation
reduces obsessive-compulsive disorders48.
In conclusion, our study has demonstrated that motivation for
natural rewards and drug abuse can be oppositely modulated. This
possibility opens new perspectives for the development of specific and
efficient treatments for drug abuse. In this context, manipulations that
decrease the activity of the STN may represent a new therapeutic
strategy for drug addiction.
METHODS
Animals. Male Wistar rats (n ¼ 20; Iffa Credo) and Long-Evans rats (n ¼ 67;
Janvier) were housed in pairs and were maintained on a 12-h light-dark
cycle. During the food experiments, they were maintained at 80–85% of their
free-feeding weight by restriction of their food to 15–17 g/rat per day. For
cocaine experiments, the rats had no food restriction. Water was provided ad
libitum, except during experimental sessions. All procedures were conducted in
accordance with the French Agriculture and Forestry Ministry decree 87-849.
Surgery. Rats were anaesthetized with xylazine (15 mg/kg intramuscularly) and
ketamine (100 mg/kg intramuscularly) and were secured in a Kopf stereotaxic
apparatus. Rats received bilateral injection of ibotenic acid (9.4 mg/ml (53 mM);
n ¼ 42, STN-lesioned) or vehicle solution (phosphate buffer; 0.1 M; n ¼ 38,
sham-lesioned) at the following coordinates49: anteroposterior, 3.8 mm (from
bregma); lateral, 7 2.4 mm; dorsoventral, 8.35 mm (from skull). The volume
injected was 0.5 ml per side infused over 3 min with a 10-ml Hamilton
microsyringe connected by Tygon tubing fitted to the 30-gauge stainless steel
injector needles and fixed on a micropump. The injectors were left in place for
3 min to allow diffusion. Twenty rats (n ¼ 10, sham-lesioned; n ¼ 10, STNlesioned) were further subjected to implantation of a catheter in the jugular
vein. Seven rats were implanted with bilateral cannulae above the STN
(dorosventral, 3 mm dorsal) so that the injector would protrude 3 mm below
the cannulae. Cannulae were fixed with dental cement, and wire stylets 10 mm
in length were inserted in the guide cannulae to prevent occlusion.
Microinfusions. Rats were tested in the progressive ratio task for a couple of
sessions and then received the vehicle injection. Rats were injected with a
solution of vehicle or muscimol (2 or 3 ng per 0.5 ml per side) over a 3-min
period as described above. One injection per week was done. Muscimol
(Sigma-Aldrich) was dissolved in 0.9% NaCl saline solution at doses that have
been shown to affect attentional processes in the STN23.
Apparatus. Food reward experiments were done in four standard operant
boxes (MedAssociates) with a retractable lever, a food magazine connected with
a food pellet dispenser and a stimulus light located above the food magazine.
The apparatus and online data collection were controlled by a computer and an
interface (MedPC.). Cocaine self-administration experiments were done in eight
boxes (Imetronics) equipped with two retractable levers above which was situated
a stimulus light. The apparatus and online data collection were controlled by a
computer and an interface (Imetronics). Two Plexiglas boxes (90 5 33 cm)
divided into two main compartments, A and B (40 35 33 cm), with a small
‘intermediate’ chamber between (10 cm in length), were used for the
conditioned place-preference test. Compartments A and B had different color
patterns on the wall and differed in material on the floor.
Behavioral protocol for the continuous and progressive ratio schedule of
reinforcement. For the food experiment, as described15, rats (n ¼ 8, shamlesioned; n ¼ 8, STN-lesioned; n ¼ 7, cannulated) were trained to press a lever
for one food pellet with a continuous reinforcement schedule (fixed ratio 1).
The session ended when 100 pellets were delivered or 30 min had elapsed. After
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acquisition of stable responding, all rats were subjected to the progressive ratio
schedule, in which the number of presses required (fixed ratio) was arithmetically increased by steps of five with three repetitions of each step. The lever
press that completed each fixed ratio within this progressive ratio schedule was
rewarded. A session ended if the rat failed to press the lever for 5 consecutive
minutes or 90 min had elapsed. The ‘fixed ratio 1’ procedure started 1 week
after surgery for 10 consecutive days and the progressive ratio schedule was
then undertaken for ten consecutive daily sessions. After ten sessions of
progressive ratio for one pellet, five sessions of progressive ratio were conducted
with two pellets as the reinforcer. For each session, the value of the last
reinforced ratio reached (the breaking point) was recorded, as well as the
number of pellets earned. The mean number of pellets earned by session over
the ten sessions for one pellet and the five sessions for two pellets was measured
for each rat.
In the cocaine experiment, for the acquisition procedure (continuous
reinforcement fixed ratio 1), 2 weeks after surgery, rats (n ¼ 10, sham-lesioned;
n ¼ 10, STN-lesioned) were trained to self-administer cocaine at a dose
classically used in this procedure (250 mg per 90-ml infusion in 3 s) during
13–15 daily 2-h sessions in the operant chambers. Pressing one lever (the active
lever) switched on the light above the lever for 3 s and delivered cocaine to the
blood stream. A 20-s ‘time-out’ was imposed during which any further presses
were recorded but had no consequence. Pressing the other lever (the inactive
lever) had no further consequence.
For the ‘progressive ratio schedule of reinforcement’ part of the cocaine
experiment, when the session started, a ‘free’ injection was delivered to fill the
catheter. Pressing of the active lever after increasing ‘ratios’ (the number of lever
presses required for a reward) was required for receipt of a 250-mg injection of
cocaine. The ratios followed the modified equation of Roberts50 (number of
lever presses required: 1, 3, 6, 10, 15, 20, 25, 32, 40, 50, 62, 77, 95, 118, 145, 178,
219 and so on). The session ended when the rat failed to complete a ratio
within the hour after an injection. The rats were submitted to this schedule for
ten consecutive daily sessions. For each session, the last ratio reached was
recorded, as well as the number of reinforcers obtained and lever presses as well
as locomotor activity.
For the dose-response part of the cocaine experiment, after the progressive
ratio schedule, the rats were subjected to a ‘fixed ratio 1’ schedule for four
additional sessions. On the fifth and sixth sessions, after 1 h of testing at the
250-mg dose, the dose was decreased every hour (80 and 10 mg on the fifth day
and 30 and 0 mg on the sixth day). The number of injections obtained during
the adaptation phase (first 10 min after the dose switch) was analyzed, as well as
the total number of injections during the entire 1-h session.
Behavioral protocol for the conditioned place-preference experiment. For
preconditioning, 44 rats (sham-lesioned, n ¼ 20; STN-lesioned, n ¼ 24) were
placed for 10 min in the place-preference apparatus. Time spent in each
compartment was measured in seconds. Rats were then subdivided into six
groups and tested for food or cocaine (5 or 10 mg/kg) place preference.
For conditioned place-preference experiment using food, two groups (n ¼ 6,
sham-lesioned; n ¼ 8, STN-lesioned) were subjected to food restriction (15 g of
standard lab chow per rat). For conditioning, on days 1, 3, 5 and 7, rats had
access to sucrose pellets (4.5 g total; 100 pellets, 45 mg each) for a 30-min
period, with half of the rats in compartment A and the other half in
compartment B. On days 2, 4, 6 and 8, rats were placed for 30 min in the
opposite compartment without food. For testing, on day 9, all rats were placed
in the middle chamber from which both compartments were accessible. The
time spent in each compartment was measured for 10 min.
For the conditioned place-preference experiment using cocaine, a similar
procedure was followed with four other groups (n ¼ 8 and n ¼ 6, shamlesioned, and n ¼ 8 and n ¼ 8, STN-lesioned, for the 5- and 10-mg/kg doses,
respectively) injection of cocaine (5 10 mg/kg or 10 mg/kg intraperitioneally)
replaced the sucrose pellets, and on alternate days, rats received an injection
of vehicle.
Statistical analysis. Results are expressed as means for each variable (i.e.,
number of pellets or injections, last ratio reached in the progressive ratio
procedures, score of place preference and so on) in the various groups of rats.
For each variable, the data were submitted to mixed-design analyses of variance
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(Statview; SAS) with group (sham-lesioned versus STN-lesioned) as the
between-subject factor and session or drug as the within-subject factor. When
significant effects were found, post-hoc comparisons were made using simple
main effects analysis. The nonparametric Mann-Whitney test was used for the
conditioned place-preference experiment for group effects and the nonparametric sign test was used for conditioning effect.
Histology. At the completion of testing, all rats were deeply sedated by chloral
anesthesia (400 mg/kg intraperitoneally) and were perfused with a 4%
paraformaldehyde solution (Sigma). Brains were removed and were kept
in a 10% sucrose solution and frozen to be further cut with a cryostat. Frontal
sections of the STN 30–40 mm in thickness were cut and were stained with
cresyl violet for assessment of the extent and location of the lesions and
injection sites.
ACKNOWLEDGMENTS
This study was supported by the Centre National de la Recherche Scientifique,
a European Community 5th PCRDT program funding (QLK6-1999-02173),
and by the Fondation France Parkinson and Conseil Régional d’Aquitaine. The
authors thank P.V. Piazza for critical reading and corrections of the manuscript;
B. J. Everitt, T.W. Robbins, S.H. Ahmed and L. Vanderschuren for discussion of
the results and manuscript; and Y. Darbaky and R. LeCozannet for help.
COMPETING INTERESTS STATEMENT
The authors declare that they have no competing financial interests.
Received 16 February; accepted 2 March 2005
Published online at http://www.nature.com/natureneuroscience/
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