their efficacy in suppressing amphetamine-induced locomotion and conditioned avoidance responding. Treatment failure occurred
despite high levels of dopamine D2receptor occupancy by the antipsychotic and was at least temporarily reversible by an additional
increase in antipsychotic dose. To explore potential mechanisms, we studied presynaptic and postsynaptic elements of the dopamine
system and observed that antipsychotic failure was accompanied by opposing changes across the synapse: tolerance to the ability of
haloperidol to increase basal dopamine and dopamine turnover on one side, and 20–40% increases in D2receptor number and 100–
chotic efficacy is linked to an increase in D2receptor number and sensitivity. These results are the first to demonstrate that “break-
inergic) treatment for years, yet remarkably little is known about
what happens to dopamine (DA) function during ongoing treat-
ment. What is known, however, is that in both humans and lab-
oratory animals withdrawal from chronic antipsychotic treat-
ment reveals a state of dopaminergic supersensitivity that is
characterized by increased vulnerability to psychosis and to the
psychomotor activating effects of dopamine agonists, respec-
tively. In humans, this has been termed “neuroleptic-induced
supersensitivity psychosis” (Chouinard et al., 1978; Chouinard
and Jones, 1980), and has been observed after withdrawal from
antipsychotic drugs such as quetiapine (Margolese et al., 2002),
ical observations, animal studies show that withdrawal from an-
et al., 1982; Meng et al., 1998), and dopamine injected into the
caudate–putamen or nucleus accumbens (Halperin et al., 1983).
Although such studies have conclusively demonstrated dopamine
Indeed, the concept of antipsychotic-induced dopamine su-
persensitivity is based almost entirely on studies examining
changes in the dopamine system when drug treatment and
dopamine antagonism have ceased, and a period of withdrawal
has elapsed (Muller and Seeman, 1978; Rupniak et al., 1983). To
our knowledge, only Clow et al. (1979, 1980) have studied anti-
psychotic-induced dopamine supersensitivity without an overt
withdrawal period. However, antipsychotics were given in the
drinking water and plasma drug levels were not monitored. Rats
consume up to 90% of their daily water intake during the dark
TheJournalofNeuroscience,March14,2007 • 27(11):2979–2986 • 2979
phase (Stellar and Hill, 1952; Fitzsimons, 1957) and metabolize
antipsychotics much faster than humans (Cheng and Paalzow,
1992; Bezchlibnyk-Butler and Jeffries, 1999); thus, the levels of
Given these considerations, key questions remain to be an-
swered by preclinical models. First, can dopamine supersensitiv-
ity be observed during ongoing treatment with therapeutically
relevant doses and modes of antipsychotic administration? Sec-
ond, can dopamine supersensitivity overcome the effects of anti-
we examined whether this was related to progressive changes in
presynaptic dopamine levels, turnover and release, and/or to
Male Sprague Dawley rats (Charles River Laboratories, Montreal, Que-
controlled colony room with a 12 h reverse light/dark cycle (lights off at
8:00 A.M.). Food and water were available ad libitum. All testing was
conducted during the dark phase of the animals’ circadian cycle and was
in compliance with the institute’s animal care committee.
Haloperidol (HAL; 0.25 mg/kg/d or 0.75 mg/kg/d for all studies except
bec, Canada) was dissolved in a 0.5% glacial acetic acid/H2O solution
(pH adjusted to ?5 with NaOH). Olanzapine (OLZ; 10 mg/kg/d) was
dissolved in a 2% glacial acetic acid/H2O solution (pH adjusted to ?5
with NaOH). Both drugs were given via an Alzet osmotic minipump
(model 2ML2; 14 d delivery; Durect, Cupertino, CA). D-Amphetamine
sulfate (AMPH; 1.5 mg/kg; US Pharmacopoeia, Rockville, MD) was dis-
solved in 0.9% saline and given subcutaneously in a volume of 1 ml/kg
Minipumps containing either vehicle (VEH; 0.5% glacial acetic acid/
H2O solution), HAL, or OLZ were implanted under 1.5% isoflurane
and hemostats were used to loosen connective tissue between the scapu-
the scapulae with the flow moderator away from the incision. The inci-
sion was closed using 9 mm surgical staples and cleaned with 70%
The purpose of experiment 1 was two-fold: (1) to confirm that the HAL
doses used would yield clinically relevant levels of D2receptor blockade
receptor blockade by HAL would change during continued treatment.
On days 2 and 13 after minipump implantation, in vivo D2receptor
occupancy was determined in animals from the VEH (n ? 4 per time
point) and HAL (n ? 7 per time point) groups. Individual animals from
was collected. The striata and cerebellum were rapidly dissected,
Packard, Montreal, Quebec, Canada) at room temperature. The next
day, 6 ml of scintillation fluid (Aquasure; Canberra Packard) was added
to the mixture and vials were left on a shaker for 24 h at room tempera-
efficiency. Striatal and cerebellar counts were expressed as disintegra-
tions per minute/milligram. The ratio of striatum minus cerebellum (an
index of specific binding)/cerebellum (an index of nonspecific binding)
was used to generate an index of the binding potential (BP) of DA D2
receptors. Percent D2occupancy by haloperidol was calculated as fol-
lows: 100 ? (BPvehicle? BPhaloperidol/BPvehicle), where BPvehicleis the
pooled D2binding potential of all the vehicle animals and BPhaloperidolis
the binding potential of a haloperidol-treated rat.
Experiment 2a: acute versus chronic HAL
In experiment 2a, we assessed changes in the locomotor response to
AMPH as a function of chronic HAL exposure.
Apparatus. The locomotor response to AMPH was assessed in clear
Plexiglas cages (27 ? 48 ? 20 cm) equipped with a row of 6 photocell
beams placed 3 cm above the floor of the cage. Photocell beam breaks
were detected and recorded by a computer.
Groups and procedures. The influence of HAL or vehicle on AMPH-
induced locomotion was assessed 2 and 12 d after minipump implanta-
also examined on the fifth day of withdrawal from HAL in the animals
the locomotor activity room and left in their transport cages for 5 min.
The animals were then placed in the locomotor activity cages and left
undisturbed for 30 min during which baseline levels of locomotor activ-
ity were monitored. Animals were then injected with AMPH and a 60
min test period followed during which locomotor activity was recorded.
Experiment 2b: effects of further augmenting HAL levels
In experiment 2b, we assessed whether an acute injection of HAL could
attenuate the locomotor response to AMPH once chronic and continu-
ous HAL treatment via minipump was no longer able to do so.
Rats were implanted with minipumps containing VEH or 0.75 mg/
kg/d HAL as described above. AMPH-induced locomotion was assessed
on the 12th and 13th days after implantation. On day 12, one-half of the
animals in each group received a subcutaneous injection of either saline
or HAL (0.1 mg/kg) 30 min before the AMPH injection. On day 13,
animals previously given saline before AMPH now received HAL, and
Experiment 2c: acute versus chronic OLZ
Experiment 2c was conducted to assess changes in the effect of continu-
ous OLZ exposure on AMPH-induced locomotion over time. The dose
et al., 2003; Turrone et al., 2005). Animals were implanted with
described in experiment 2a.
In experiment 3, we monitored the ability of HAL treatment to suppress
the conditioned avoidance response to an aversive conditioned stimulus
Rats were trained and tested in six identical two-way active avoidance
shuttle boxes (64 ? 24 ? 30 cm; Med Associates, St. Albans, VT) set in
individual ventilated and sound- and light-attenuating cubicles (97 ?
a white polyvinyl chloride partition with an arch-shaped doorway (15 ?
9 cm) and a 4-cm-high barrier was fixed onto the doorway. Animals had
to jump over the barrier to cross from one compartment to the other.
Illumination was provided by a 28 V house light mounted on the back
wall of the right compartment. The shuttle boxes were equipped with a
tilting grid floor and animal location was detected by microswitches.
Scrambled foot shocks [0.6 mA; unconditioned stimulus (US)] lasting
20 s were delivered to the grid floor. The conditioned stimulus (CS) was
a 74 dB tone lasting 10 s. Stimulus presentation was controlled by
computer-run Med-Associates programs, which also recorded the be-
havioral measures described below.
2980 • J.Neurosci.,March14,2007 • 27(11):2979–2986 Samahaetal.•AntipsychoticFailureoverTime
Each CS presentation was immediately followed by foot shock. Move-
ment to the other compartment during the 10 s CS presentation was
recorded as “avoidance.” Movement to the other compartment during
presentation of the foot shock was recorded as “escape,” and failure to
move to the other compartment during presentation of the foot shock
was recorded as “escape failure.” Intertrial compartment crossings were
also recorded. Each training/testing session consisted of 30 trials (30
CS–US presentations) with an intertrial interval of 30–60 s. Rats were
first trained once a day for a total of 9 d. Only animals that reached a
training criterion of ?50% avoidance on days 4 and 5 were kept for
additional training on days 6–9 (32 of 48 rats).
Finally, only animals that had ?80% avoidance on days 8 and 9 were
of HAL was chosen because it is halfway between the 0.25 and 0.75
mg/kg/d doses used in experiment 2a. Starting on day 3 of HAL/VEH
treatment, the same animals were tested for conditioned avoidance re-
sponding (CAR) once a day for five consecutive days (i.e., until day 7 of
treatment), and then on days 10 and 12 of treatment.
Experiment 4: in vivo microdialysis
In experiment 4, we used in vivo microdialysis techniques in freely mov-
ing animals to examine changes in extracellular DA, dihydroxyphenyla-
cetic acid (DOPAC), homovanillic acid (HVA), and norepinephrine
(NE) in the nucleus accumbens during ongoing HAL treatment.
Animals were anesthetized with sodium pentobarbital (Nembutal, 65
mg/kg, i.p.; Sigma, Oakville, ON) and atropine sulfate (0.13 mg/kg, s.c.;
Sabex) to minimize bronchial secretions. Stereotaxic procedures were
tific, Montreal, Quebec, Canada) into the nucleus accumbens (antero-
mm from the site) at a 10° angle to avoid damage to the lateral ventricle.
The cannula was cemented in place with dental acrylic applied around
the cannula and three skull screws. The hemisphere to be implanted was
icillin (Pen G, i.m.; Vetoquinol, Lavaltrie, Quebec, Canada) and Keto-
profen (2 mg/kg, s.c.) after surgery. Three to 4 d after cannulation, ani-
HAL as described above. Animals were left to recover from intracranial
implantation, amphetamine-induced DA, DOPAC, HVA, and NE re-
lease were measured in separate groups of animals according to the fol-
Four hexagonal chambers were used for microdialysis. Each chamber
bec, Canada) consisted of Plexiglas walls with wooden ceilings and
cubicles and lighting was provided on a reverse cycle by overhead lights.
Each dialysis probe consisted of a 2.5 mm length of semipermeable dial-
ysis membrane (inner diameter, 200 ?m; 13, 000 molecular weight cut-
off; Spectrum Laboratories, Rancho Dominguez, CA), closed at one end
with glue and inserted into a 21.5- to 22-mm-long 26 gauge piece of
end of a 40–50 cm piece of PE20 tubing that connected to a single chan-
nel liquid swivel (home made, Concordia University). The swivel was in
turn connected to a Harvard syringe pump (Harvard Apparatus, South
diameter, 108 ?m; inner diameter, 40 ?m; HRS Scientific) extended
of the silica tubing in polyethylene tubes. The external length of the PE
tubing was protected from chewing by steel spring casing. The probes
were inserted the day before the microdialysis testing session and were
cannula. To prevent occlusion, artificial CSF [ACSF; containing (in
mM)145 Na?, 2.7 K?, 1.3 Ca2?, 1.0 Mg2?, 150 Cl?, 0.2 ascorbate, and
2 Na2HPO4, pH, 7.4 ? 0.1) was perfused overnight at a rate of 0.3 ?l/
min. ACSF infusion rate was changed to 0.8 ?l/min during dialysate
High-performance liquid chromatography
Samples were collected every 20 min and a 10 ?l volume of dialysate was
immediately extracted from each sample and analyzed using one of two
similar high-performance liquid chromatography (HPLC) systems with
electrochemical detection. The samples were loaded onto C-18 reverse-
phase columns (15 cm ? 5 ?m, Spherisorb-ODS2; Chromatography
Sciences, St. Laurent, Quebec, Canada) through manual injection ports
(Reodyn 7125; 20 ?l loop). Reduction and oxidation currents for DA,
ford, MA) coulometric detectors (Coulochem III, with a model 5011
analytical cell). The currents for DA and NE (?280 mV) were measured
channels of the Coulochem detectors. The mobile phases (19% acetoni-
trile, 0.076 M SDS, 0.1 M EDTA, 0.058 M NaPO4, 0.027 M citric acid, pH
3.35) were circulated through each closed system at a flow rate of 1.2
ml/min by Waters (Lachine, Quebec, Canada) 515 HPLC pumps. The
peaks obtained for DA, DOPAC, HVA, and NE were integrated and
quantified by EZChrom Chromatography Data System (Scientific Soft-
ware, San Ramon, CA). The mobile phase was adjusted to allow for the
separation and quantification of target analytes in a single run.
were achieved (?10% variation in three consecutive samples), ani-
mals were injected with saline and two more samples were collected.
Animals were then given AMPH (1.5 mg/kg, s.c.) and eight additional
samples were collected.
Postmortem cannula placement verification
Rats were anesthetized with Somnotol (i.p.; MTC Pharmaceuticals,
Cambridge, Ontario, Canada) and perfused transcardially with 0.9% sa-
line (150 ml) and formaldehyde (150 ml, formalin 10% v/v; Anachemia,
Montreal, Quebec, Canada). Brains were extracted, left in formaldehyde
for 24 h, and 20 ?m coronal sections were taken on a cryostat. Sections
were stained with cresyl violet and probe tracts were verified under a
In experiment 5, we quantified changes in striatal D2binding capacity
9-hydroxynaphthoxazine) is chemically defined as (?)-4-propyl-
3,4,4a,5,6,10b-hexahydro-2H-naphtho[1,2-b][1,4]oxazin-9-ol. To pre-
of withdrawal, animals in each treatment group were killed by CO2nar-
cosis and their striata were dissected and stored at ?70°C until use. In
used to determine whether the mere presence of HAL in the tissue could
influence the density of D2
were added to these striata during processing. All striata were homoge-
nized individually in buffer (4 mg of frozen tissue/ml of buffer), using a
Teflon-glass homogenizer with the piston rotating at 500 rpm and 10 up
NaCl. The homogenate was not washed, centrifuged, or preincubated
Highstates as a function of chronic HAL treatment.
Highstates were measured using [3H](?)PHNO, a D2agonist.
Highstates. Thus, 0.75 and 1.5 nM haloperidol
Samahaetal.•AntipsychoticFailureoverTimeJ.Neurosci.,March14,2007 • 27(11):2979–2986 • 2981
these procedures (Seeman et al., 1984).
The density of [3H](?)-PHNO sites sensitive to the action of guani-
lylimidodiphosphate (GN), or so-called “G-sensitive [3H](?)-PHNO”
sites, was determined by obtaining two saturation curves with [3H](?)-
PHNO, one with and one without GN. Each incubation tube (12 ? 75
mm, glass) received, in the following order, 0.5 ml of buffer (with or
without a final concentration of 200 ?M GN, and with or without a final
concentration of 10 ?M S-sulpiride to define nonspecific binding), 0.25
ml [3H](?)-PHNO (with 12 final concentrations ranging from 0.1 to 5
nM), and 0.25 ml of tissue homogenate. The tubes, containing a total
which the incubates were filtered using a 12-well cell harvester (Titertek;
Skatron, Lier, Norway) and buffer-presoaked glass fiber filter mats (No.
buffer for 15 s (7.5 ml of buffer). The filters were pushed out and placed
in scintillation minivials (Packard Instruments, Chicago, IL). The mini-
vials received 4 ml each of scintillant (Ready Solve; Beckman), and were
monitored 6 h later for tritium in a Packard 4660 scintillation spectrom-
eter at 55% efficiency. Specific binding at each concentration of
of 10 ?M S-sulpiride. The data for each saturation curve were graphed as
a Scatchard plot, yielding the Bmaxdensity (in picomoles per gram) and
the KD(in nanomolars). The Bmaxin the absence of GN minus the Bmax
in the presence of GN represented the density of G-sensitive [3H](?)-
PHNO-labeled D2receptors. The Bmaxin the absence of S-sulpiride mi-
ing capacity, which was used as an index of D2receptor number.
In experiment 1, we determined whether the HAL doses tested
were yielding clinically meaningful levels of D2blockade and
whether this was maintained during chronic treatment. The lev-
in clinical treatment: during short-term HAL treatment (day 2),
average striatal D2-receptor occupancy was 84% (?SEM 2.06)
animals treated with 0.75 mg/kg/d. With continued treatment
(day 13), average D2occupancy was similar to that seen during
short-term treatment in animals given 0.75 mg/kg/d HAL (82%,
?SEM 0.64), but was reduced in animals treated with the lower
dose, although still well within clinically relevant ranges (69%,
?SEM 2.7; unpaired t test on day 2 vs day 13; t ? 4.12; p ?
both HAL doses and this was maintained during the course of
In experiment 2a, we examined changes in the ability of HAL to
block AMPH-induced locomotion with continued treatment.
Early in treatment (day 2) (Fig. 1a), both doses of HAL inhibited
the total locomotor response to AMPH by 60% (?SEM 9). Dur-
ing chronic treatment (day 12) (Fig. 1b), HAL no longer dis-
rupted AMPH-induced locomotion. On the fifth day of with-
(?SEM 28) higher in animals treated previously with 0.75 mg/
icant. Thus, HAL initially attenuated AMPH-induced locomo-
tion but gradually lost this ability, and once HAL treatment was
terminated, a supersensitive locomotor response to AMPH
In experiment 2b, we determined whether augmenting chronic
levels of HAL by giving an acute injection of the neuroleptic (0.1
mg/kg, s.c.) could suppress AMPH-induced locomotion when
the ongoing HAL treatment (via minipump) had lost the ability
to do so. AMPH-induced locomotion in rats chronically treated
experiment 2a (Fig. 1b). An acute injection of HAL suppressed
AMPH-induced locomotion in both neuroleptic-naive rats and
in rats maintained on HAL via minipumps (paired t tests on the
locomotor response to AMPH over time. Initially, both HAL (a) and OLZ (d) reduced AMPH-
locomotion ( p ? 0.05). In addition, amphetamine-induced locomotion was increased in all
starting at 40 min; 0.25 mg/kg/d HAL, t ? ?3.402; 0.75 mg/kg/d HAL, t ? ?2.307; 10
2982 • J.Neurosci.,March14,2007 • 27(11):2979–2986Samahaetal.•AntipsychoticFailureoverTime
HAL, t ? 2.84 and 3.96, respectively; p ? 0.05). In addition, an
same extent in animals maintained on HAL or VEH treatment via
minipump (unpaired t test, t ? ?1.86; p ? 0.10). Thus, during
In experiment 2c, we determined whether these effects of HAL
could be extended to an atypical antipsychotic drug, OLZ. Early
in OLZ treatment (day 2) (Fig. 1d), AMPH-induced locomotion
was attenuated relative to control levels. During chronic treat-
ment (day 12) (Fig. 1e), OLZ no longer suppressed AMPH-
comotion was not statistically different from that seen in control
rats. Thus, as seen in Figure 1a–b with HAL, OLZ initially sup-
pressed the locomotor activating effects of AMPH, but progres-
sively lost this ability during treatment.
In experiment 3, we examined whether the loss of efficacy of
antipsychotics could be seen in a nonpharmacological model
with very strong predictive validity for antipsychotic action: the
CAR test. As seen in Figure 2, HAL initially suppressed CAR
relative to predrug (day 0) and control levels. The disruptive
Experiment 4: in vivo microdialysis of DA, DOPAC, HVA,
In experiment 4, we examined the possibility that behavioral su-
persensitivity to AMPH during chronic HAL treatment was
caused by presynaptic changes in DA by assessing extracellular
levels of extracellular DA and DOPAC (Fig. 3a, inset, b), but did
not significantly change HVA (Fig. 3c) or NE levels (data not
shown). With continued treatment, HAL no longer enhanced
those seen in controls (Fig. 3d, inset, e,f) (NE, data not shown).
HVA levels (Figs. 3b,c,e,f) at both time points tested and in both
treatment groups. Early in treatment (day 2), HAL had no statis-
tically significant effect on the neurochemical response to
levels of DA and DOPAC (a, inset, b; 2-way ANOVAs on ?120 to ?20 min, main effects of
similar to those of controls with chronic treatment (d, inset, e). Basal HVA levels were not
significantly affected by HAL on either testing day (c, f). An injection of saline did not alter
a–f, Extracellular DA, DOPAC, and HVA in the nucleus accumbens during short-
Samahaetal.•AntipsychoticFailureoverTimeJ.Neurosci.,March14,2007 • 27(11):2979–2986 • 2983
AMPH, but chronic HAL treatment attenuated the AMPH-
induced decrease in DOPAC and HVA (Figs. 3e,f). Thus HAL-
a reduction in the suppressive effects of AMPH on DA turnover,
but not by significant changes in extracellular levels of DA.
Given the results of experiment 4 showing that chronic HAL
treatment did not significantly change AMPH-induced extracel-
addition of 0.75 or 1.5 nM HAL to striatal tissue did not alter D2
receptor or D2
mg/kg/d HAL did not change D2receptor Bmaxat any time point
tested (Fig. 4a). Exposure to 0.75 mg/kg/d HAL increased D2
these animals was also greater after neuroleptic withdrawal (day
21) than during acute treatment (day 2). No other comparisons
Antipsychotic treatment also influenced the density of D2re-
DA. Relative to control levels, treatment with 0.75 mg/kg/d HAL
elevated striatal D2
HighBmax(data not shown). Treatment with 0.25
Highsites by 137–188% (?SEM 20–47) from
day 2 onward, and D2
104% (?SEM 32) only during chronic treatment (day 12) and
levels went back to normal after neuroleptic withdrawal. Thus,
ioral response to AMPH in both HAL treated groups.
Highlevels did not change over time (Fig.
Previous work has shown that withdrawal from chronic antipsy-
chotic treatment leads to a supersensitive psychomotor response
to dopamine agonists (Gianutsos et al., 1974; Sayers et al., 1975;
Smith and Davis, 1975, 1976; Clow et al., 1979; Montanaro et al.,
1982; Rebec et al., 1982; Meng et al., 1998). We show here that
drawal, but develops early during antipsychotic exposure and
significantly undermines the efficacy of ongoing treatment. The
loss of efficacy was seen with typical or atypical antipsychotics in
two widely used tests of antipsychotic-like effects in animals and
receptor blockade. Thus, the effects were not likely caused by
pharmacokinetic or peripheral factors, but by compensatory
neurobiological changes in response to ongoing treatment.
is an increase in dopamine availability over time, which would
surmount the antidopaminergic effects of the antipsychotic.
However, in vivo microdialysis measurements revealed that
amphetamine-induced increases in extracellular dopamine were
unchanged during haloperidol treatment. This suggests that the
amphetamine-induced locomotion was not attributable to
changes in dopamine availability. These data extend previous
work showing that although withdrawal from haloperidol pro-
duces a supersensitive behavioral response to amphetamine
(Smith and Davis, 1975; Rebec et al., 1982; Meng et al., 1998), it
does not change the dopamine response to amphetamine
al., 1992). However, microdialysis measurements are a function
of release and uptake and the lack of change in dopamine over-
flow in our study could be reflecting increased release and in-
creased uptake or, alternatively, decreased release and decreased
uptake. Thus, despite our microdialysis findings, presynaptic
changes could still be involved in the development of dopamine
supersensitivity and loss of antipsychotic treatment efficacy. Im-
oral treatment with a dose of haloperidol very similar to the one
tested here (0.7 mg/kg/d) did not alter the sensitivity of in vivo
dopamine overflow to either an agonist or an antagonist at pre-
synaptic autoreceptors (sulpiride and quinpirole, respectively).
This suggests that the gradual loss of efficacy and behavioral su-
persensitivity that develops during antipsychotic treatment is
likely caused by postsynaptic changes.
Although haloperidol did not change the dopaminergic re-
sponse to amphetamine over time, it did attenuate the
of explanations can be considered. First, long-term exposure to
haloperidol itself reduced dopamine metabolite levels and this
could have limited the extent to which amphetamine could sup-
press these (i.e., metabolite levels could already have been near
“floor” levels). Second, amphetamine decreases dopamine me-
(a) and D2
Highstates (b) over time. a, Treatment with 0.25 mg/kg/d HAL did not change D2
2984 • J.Neurosci.,March14,2007 • 27(11):2979–2986Samahaetal.•AntipsychoticFailureoverTime
mally breaks down intraneuronal dopamine into DOPAC and
HVA (Zetterstrom et al., 1986). However, it is unclear how hal-
operidol could influence this response given that amphetamine-
induced dopamine overflow was not affected by the neuroleptic.
Third, amphetamine also suppresses dopamine metabolite levels
by inhibiting monoamine oxidase type A (Green, 1971; Zetter-
strom et al., 1986). Although we did not measure monoamine
oxidase levels in this study, haloperidol treatment does not ap-
pear to alter these (Van Der Krogt et al., 1982). As such, the
precise mechanisms by which long-term haloperidol treatment
remain to be identified.
There is evidence that postsynaptic processes contribute to
antipsychotic-induced behavioral supersensitivity. For example,
chronic haloperidol potentiates the locomotor activating effects
of intra-accumbens and intracaudate–putamen infusions of do-
pamine (Halperin et al., 1983). Changes in dopamine signaling
et al., 1983; Joyce, 2001). Both haloperidol doses we tested led to
a supersensitive locomotor response to amphetamine, but only
the higher dose increased striatal D2receptor number. Indeed,
behavioral sensitivity to dopamine is not always predicted by
In contrast, animal models of dopamine supersensitivity are sys-
tematically linked to elevations in D2
the lower haloperidol dose tested, the time course of changes in
Both the locomotor response to amphetamine and D2
were increased during antipsychotic treatment, and both re-
turned to control levels after treatment cessation. At the higher
haloperidol dose, behavioral dopamine supersensitivity was also
well predicted by changes in D2
exception. Short-term (2 d) treatment with this dose elevated
ing amphetamine-induced locomotion. This suggests two, non-
mutually exclusive possibilities. First, different neurobiological
mechanisms might underlie the dopamine supersensitivity in-
duced by low versus higher doses of antipsychotic. Second, ele-
induce dopamine supersensitivity and loss of antipsychotic effi-
supersensitivity and functional tolerance to antipsychotics are
due in part to changes in striatal dopamine receptor function,
although the assays that we have used do not allow us to deter-
mine whether this is occurring at D2receptors on local cells
and/or on cells projecting to the striatum. In support of a role of
altered dopamine receptor function was the finding that further
augmenting haloperidol levels by an acute injection of the drug
haloperidol treatment had lost the ability to do so. Thus, addi-
tional haloperidol (presumably leading to additional D2block-
ade) can overcome whatever mechanism is responsible for the
expression of dopamine supersensitivity. It remains unknown,
over time, or lose to heightened breakthrough supersensitivity.
This is reminiscent of evidence from the clinic, where increasing
dose can, at least provisionally, reduce psychotic symptoms in
patients that have developed therapeutic tolerance.
In contrast to most preclinical work where antipsychotics are
Highreceptors (Seeman et
Highlevels predicted behavioral supersensitivity to dopamine.
Highlevels, with the following
Highlevels might be necessary although not sufficient to
via minipump. This is an important issue when one considers
that the terminal half-life of haloperidol is 24 h in humans
(Bezchlibnyk-Butler and Jeffries, 1999) and 1.5 h in rats (Cheng
and Paalzow, 1992). Thus, 24 h after a single dose of haloperidol,
D2occupancy levels remain high in humans (Farde et al., 1989;
Seeman, 2002), but fall to well below clinically relevant levels in
rats (Kapur et al., 2000a). Our findings demonstrate that contin-
uous antipsychotic treatment and D2receptor blockade induces
neuroadaptations that lead to antipsychotic failure. It is possible,
therefore, that much of what is currently known about the neu-
apply if antipsychotics are given continuously rather than inter-
mittently. Indeed, merely changing the mode of antipsychotic
administration (i.e., intermittent vs continuous infusion) can
have markedly different effects (Turrone et al., 2003, 2005). An-
imal studies that use a mode of drug administration that more
closely mimics clinical antipsychotic treatment (i.e., relatively
effects of these drugs in humans (Kapur et al., 2003).
Although in the present study with rats haloperidol and olan-
zapine lost efficacy over time, not all treated patients develop
therapeutic tolerance. However, a notable proportion of initially
stabilized patients relapse during treatment and this cannot al-
ways be explained by nonadherence to treatment. For example,
even when medication is guaranteed by depot injection, the av-
erage relapse rate at 1–2 years is still 18–55% (Carpenter et al.,
1999; Schooler, 2003; De Graeve et al., 2005). Although relapse
during continued treatment can be attributed to a number of
reasons, our results suggest that an antipsychotic-induced in-
crease in dopamine sensitivity might predispose certain individ-
uals to psychotic relapse. This provides a discrete hypothesis that
can be tested in patients.
ing: (1) initially, antipsychotics block D2receptors, and increase
receptors and D2
ance of agonist (i.e., endogenous dopamine-related) drive and
receptor blockade allows the antipsychotic to exert a net anti-
dopaminergic effect. (2) With continued treatment, there is a
decrease in turnover on the presynaptic side, D2blockade by
antipsychotics is maintained, but both D2receptor numbers and
mine drive is potentiated and can more readily oppose the anti-
dopaminergic effects of the antipsychotic. This might explain
why antipsychotics so often fail. The challenge now is to identify
tipsychotic treatment tolerance develop. At the same time, the
biological (i.e., increases in D2receptors and D2
behavioral (loss of efficacy in behavioral models) markers iden-
overcome or prevent antipsychotic treatment failure.
Highreceptors are normal. At this stage the bal-
Highsites are elevated. At this later stage, endogenous dopa-
Asper H, Baggiolini M, Burki HR, Lauener H, Ruch W, Stille G (1973) Tol-
erance phenomena with neuroleptics catalepsy, apomorphine stereotyp-
ies and striatal dopamine metabolism in the rat after single and repeated
Bezchlibnyk-ButlerKZ,JeffriesJJ (1999) Clinicalhandbookofpsychotropic
drugs. Toronto: Hogrefe and Huber.
Burt DR, Creese I, Snyder SH (1977) Antischizophrenic drugs: chronic
treatment elevates dopamine receptor binding in brain. Science
Carpenter Jr WT, Buchanan RW, Kirkpatrick B, Lann HD, Breier AF, Sum-
Samahaetal.•AntipsychoticFailureoverTimeJ.Neurosci.,March14,2007 • 27(11):2979–2986 • 2985
merfeltAT (1999) Comparativeeffectivenessoffluphenazinedecanoate
injections every 2 weeks versus every 6 weeks. Am J Psychiatry
Cheng YF, Paalzow LK (1992) Linear pharmacokinetics of haloperidol in
the rat. Biopharm Drug Dispos 13:69–76.
Chesi AJ, Feasey-Truger KJ, Alzheimer C, ten Bruggencate G (1995) Dopa-
mine autoreceptor sensitivity is unchanged in rat nucleus accumbens
after chronic haloperidol treatment: an in vivo and in vitro voltammetric
study. Eur J Neurosci 7:2450–2457.
Chouinard G, Jones BD (1980) Neuroleptic-induced supersensitivity psy-
chosis: clinical and pharmacologic characteristics. Am J Psychiatry
Chouinard G, Jones BD, Annable L (1978) Neuroleptic-induced supersen-
sitivity psychosis. Am J Psychiatry 135:1409–1410.
Clow A, Jenner P, Marsden CD (1979) Changes in dopamine-mediated be-
haviour during one year’s neuroleptic administration. Eur J Pharmacol
ClowA,TheodorouA,JennerP,MarsdenCD (1980) Changesinratstriatal
dopamine turnover and receptor activity during one years neuroleptic
administration. Eur J Pharmacol 63:135–144.
Compton DR, Johnson KM (1989) Effects of acute and chronic clozapine
and haloperidol on in vitro release of acetylcholine and dopamine from
striatum and nucleus accumbens. J Pharmacol Exp Ther 248:521–530.
De Graeve D, Smet A, Mehnert A, Caleo S, Miadi-Fargier H, Mosqueda GJ,
LecompteD,PeuskensJ (2005) Long-actingrisperidonecomparedwith
oral olanzapine and haloperidol depot in schizophrenia: a Belgian cost-
effectiveness analysis. Pharmacoeconomics 23 [Suppl 1]:35–47.
Ekblom B, Eriksson K, Lindstrom LH (1984) Supersensitivity psychosis in
schizophrenic patients after sudden clozapine withdrawal. Psychophar-
macology (Berl) 83:293–294.
Farde L, Wiesel FA, Halldin C, Sedvall G (1988) Central D2-dopamine re-
ceptor occupancy in schizophrenic patients treated with antipsychotic
drugs. Arch Gen Psychiatry 45:71–76.
FardeL,WieselFA,NordstromAL,SedvallG (1989) D1-andD2-dopamine
receptor occupancy during treatment with conventional and atypical
neuroleptics. Psychopharmacology (Berl) 99 [Suppl]:S28–S31.
Fitzsimons JT (1957) Normal drinking in rats. J Physiology 138:39P.
Fleminger S, Rupniak NM, Hall MD, Jenner P, Marsden CD (1983)
roleptic treatment correlates with increased D-2 receptors, but not with
increases in D-1 receptors. Biochem Pharmacol 32:2921–2927.
FloresG,BarbeauD,QuirionR,SrivastavaLK (1996) Decreasedbindingof
dopamine D3 receptors in limbic subregions after neonatal bilateral le-
sion of rat hippocampus. J Neurosci 16:2020–2026.
GianutsosG,DrawbaughRB,HynesMD,LalH (1974) Behavioralevidence
for dopaminergic supersensitivity after chronic haloperidol. Life Sci
Green AL (1971) Inhibition of rat and mouse monoamine oxidase by (?)-
amphetamine. Proceedings of the biochemical society 121:37P–38P.
Halperin R, Guerin Jr JJ, Davis KL (1983) Chronic administration of three
neuroleptics: effects of behavioral supersensitivity mediated by two dif-
ferent brain regions in the rat. Life Sci 33:585–592.
Ichikawa J, Meltzer HY (1992) The effect of chronic atypical antipsychotic
drugs and haloperidol on amphetamine-induced dopamine release in
vivo. Brain Res 574:98–104.
Joyce JN (2001) D2 but not D3 receptors are elevated after 9 or 11 months
chronic haloperidol treatment: influence of withdrawal period. Synapse
Kahne GJ (1989) Rebound psychoses following the discontinuation of a
high potency neuroleptic. Can J Psychiatry 34:227–229.
Kapur S, Wadenberg ML, Remington G (2000a) Are animal studies of anti-
psychotics appropriately dosed? Lessons from the bedside to the bench.
Can J Psychiatry 45:241–246.
Kapur S, Zipursky R, Jones C, Remington G, Houle S (2000b) Relationship
between dopamine D(2) occupancy, clinical response, and side effects: a
double-blind PET study of first-episode schizophrenia. Am J Psychiatry
Kapur S, VanderSpek SC, Brownlee BA, Nobrega JN (2003) Antipsychotic
dosing in preclinical models is often unrepresentative of the clinical con-
LlorcaPM,VaivaG,LanconC (2001) Supersensitivitypsychosisinpatients
MargoleseHC,ChouinardG,BeauclairL,BelangerMC (2002) Therapeutic
tolerance and rebound psychosis during quetiapine maintenance mono-
Meng ZH, Feldpaush DL, Merchant KM (1998) Clozapine and haloperidol
block the induction of behavioral sensitization to amphetamine and as-
sociated genomic responses in rats. Brain Res Mol Brain Res 61:39–50.
Montanaro N, Dall’Olio R, Gandolfi O, Vaccheri A (1982) Differential en-
Muller P, Seeman P (1977) Brain neurotransmitter receptors after long-term
haloperidol: dopamine, acetylcholine, serotonin, alpha-noradrenergic and
Muller P, Seeman P (1978) Dopaminergic supersensitivity after neurolep-
tics: time-course and specificity. Psychopharmacology (Berl) 60:1–11.
Pierce RC, Rowlett JK, Bardo MT, Rebec GV (1991) Chronic ascorbate po-
tentiates the effects of chronic haloperidol on behavioral supersensitivity
but not D2 dopamine receptor binding. Neuroscience 45:373–378.
Rebec GV, Peirson EE, McPherson FA, Brugge K (1982) Differential sensi-
tivity to amphetamine following long-term treatment with clozapine or
haloperidol. Psychopharmacology (Berl) 77:360–366.
RupniakMN,JennerP,MarsdenCD (1983) Theeffectofchronicneurolep-
tic administration on cerebral dopamine receptor function. Life Sci
SayersAC,BurkiHR,RuchW,AsperH (1975) Neuroleptic-inducedhyper-
sensitivity of striatal dopamine receptors in the rat as a model of tardive
dyskinesias. Effects of clozapine, haloperidol, loxapine and chlorproma-
zine. Psychopharmacologia 41:97–104.
Schooler NR (2003) Relapse and rehospitalization: comparing oral and de-
pot antipsychotics. J Clin Psychiatry 64 [Suppl 16]:14–17.
SeeRE,ChapmanMA,MurrayCE,AravagiriM (1992) Regionaldifferences
in chronic neuroleptic effects on extracellular dopamine activity. Brain
Res Bull 29:473–478.
Seeman P (1980) Brain dopamine receptors. Pharmacol Rev 32:229–313.
Seeman P (2002) Atypical antipsychotics: mechanism of action. Can J Psy-
Seeman P, Ulpian C, Wreggett KA, Wells JW (1984) Dopamine receptor
parameters detected by [3H]spiperone depend on tissue concentration:
analysis and examples. J Neurochem 43:221–235.
Seeman P, Weinshenker D, Quirion R, Srivastava LK, Bhardwaj SK, Grandy
DK, Premont RT, Sotnikova TD, Boksa P, El-Ghundi M, O’Dowd BF,
George SR, Perreault ML, Mannisto PT, Robinson S, Palmiter RD, Tall-
erico T (2005) Dopamine supersensitivity correlates with D2High
states, implying many paths to psychosis. Proc Natl Acad Sci USA
Smith RC, Davis JM (1975) Behavioral supersensitivity to apomorphine
pharmacol Commun 1:285–293.
Smith RC, Davis JM (1976) Behavioral evidence for supersensitivity after
chronic administration of haloperidol, clozapine, and thioridazine. Life
Stellar E, Hill JH (1952) The rats rate of drinking as a function of water
deprivation. J Comp Physiol Psychol 45:96–102.
Controlled, double-blind investigation of the clozapine discontinuation
symptoms with conversion to either olanzapine or placebo. The Collabora-
TurroneP,RemingtonG,KapurS,NobregaJN (2003) Differentialeffectsof
within-day continuous vs. transient dopamine D2 receptor occupancy in
the development of vacuous chewing movements (VCMs) in rats. Neu-
Turrone P, Remington G, Kapur S, Nobrega JN (2005) Continuous but not
rats. Biol Psychiatry 57:406–411.
Van Der Krogt JA, Van Valkenburg CF, Belfroid RD (1982) Rat brain
monoamine oxidase activity is not affected by repeated administration of
haloperidol. J Pharm Pharmacol 34:529–531.
Zetterstrom T, Sharp T, Ungerstedt U (1986) Further evaluation of the
mechanism by which amphetamine reduces striatal dopamine metabo-
lism: a brain dialysis study. Eur J Pharmacol 132:1–9.
2986 • J.Neurosci.,March14,2007 • 27(11):2979–2986Samahaetal.•AntipsychoticFailureoverTime