Dopaminergic Agonists and Muscarinic Antagonists Improve ...

0022-3565/04/3092-737–744$20.00
THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 309, No. 2
Copyright © 2004 by The American Society for Pharmacology and Experimental Therapeutics 59519/1143745
JPET 309:737–744, 2004
Printed in U.S.A.
Dopaminergic Agonists and Muscarinic Antagonists Improve
Lateralization in Hemiparkinsonian Rats in a Novel Exploratory
Y-Maze
Makoto Nakagawa, Makoto Ohgoh, Yukio Nishizawa, and Hiroo Ogura
Tsukuba Research Laboratories, Eisai Co., Ltd., Ibaraki, Japan
Received September 5, 2003; accepted January 28, 2004
Parkinson’s disease (PD) is a progressive neurological disorder
affecting mainly elderly people. In this disease, the
degeneration of nigrostriatal dopaminergic neurons is evident,
as revealed by direct biochemical assay (Hornykiewicz,
1966, 1998) and more recently by positron emission tomography,
measuring the uptake of the dopamine precursor 18 F-
DOPA (Brooks et al., 1990; Takikawa et al., 1994). A deficit
in this dopaminergic input into the striatum causes overactivity
in the indirect motor pathway accompanied with underactivity
in the direct motor pathway in the basal ganglia
(Wichmann and DeLong, 1993). The imbalance of these motor
circuits leads to motor dysfunction, including akinesia
(impaired initiation of movement) and bradykinesia (reduced
amplitude and velocity of voluntary movement), which produce
the characteristic features of PD (Kish et al., 1988).
The immediate metabolic precursor of dopamine, levodopa
(L-3,4-dihydroxyphenylalanine, L-dopa), has been used in
Article, publication date, and citation information can be found at
http://jpet.aspetjournals.org.
DOI: 10.1124/jpet.103.059519.
ABSTRACT
Parkinson’s disease (PD) is characterized by the degeneration
of nigrostriatal dopaminergic neurons. Its primary clinical symptoms
are akinesia, tremor, and rigidity, which usually start from
one side, resembling the lateralization in hemiparkinsonian rats
having 6-hydroxydopamine-induced unilateral lesion of the medial
forebrain bundle. A novel exploratory Y-maze was designed
to detect the lateralization of hemiparkinsonian rats in
terms of biased turns in the maze. Dopamine agonists levodopa
(L-3,4-dihydroxyphenylalanine, 10–30 mg/kg) and apomorphine
(0.1–0.3 mg/kg), but not methamphetamine (0.5–2 mg/
kg), improved the lateralization in the rat model. However, high
doses of the dopamine agonists, 30 and 0.3 mg/kg, respectively,
caused small movements in the arms that seemed to
parallel the increase in counts per turn in the Y-maze. Interestingly,
the muscarinic antagonists trihexyphenidyl and scopolamine
improved lateralization moderately, while increasing total
turns, an index of locomotive activity. ()-5-Methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine
maleate (MK-801)
(0.3 mg/kg), an N-methyl-D-aspartate (NMDA) glutamate receptor
antagonist, increased total counts, but did not alleviate the
lateralization. The 2-adrenoceptor antagonist idazoxan (1 and
10 mg/kg) and 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(f)quinoxaline
(1 and 3 mg/kg), a non-NMDA glutamate receptor
antagonist, did not affect any of the indices. These findings
suggest that the clinical action of drugs on unbalanced movement
in PD could be predicted by measuring their effects on
lateralization of the 6-hydroxydopamine-lesioned rat model in
this exploratory Y-maze.
medications for the treatment of PD. It provides a drastic
reversal of all signs and symptoms of PD except dementia
and postural instability (Williams and Carlyle, 1979; Silver
and Ruggieri, 1998). Historically, direct dopamine agonists,
such as bromocriptine and pergolide, have also been used as
adjuncts to levodopa in patients with advanced disease who
already have motor complications (Olanow et al., 2001). However,
it has recently been demonstrated that initiation of
therapy with a direct dopamine agonist is associated with a
reduced risk for development of motor complications to levodopa
(Watts, 1997; Poewe, 1998). Until levodopa was introduced,
muscarinic antagonists had been the most effective
class of drugs for treatment of PD (Kopin, 1993; Dalvi and
Ford, 1998), but they are now prescribed in a supportive role
in the treatment of the disorder (Standaert and Young, 1995).
Because 6-hydroxydopamine (6-OHDA) is able to produce
selective dopaminergic lesions, it has been used extensively
in rodents to reproduce PD symptoms (Ungerstedt and
Arbuthnott, 1970; Schwarting and Huston, 1996). After
unilateral lesioning with 6-OHDA at dopaminergic somata
or ascending dopaminergic fibers, dopaminergic agents
Downloaded from jpet.aspetjournals.org by guest on August 29, 2013
ABBREVIATIONS: PD, Parkinson’s disease; 6-OHDA, 6-hydroxydopamine; MFB, medial forebrain bundle; NBQX, 2,3-dihydroxy-6-nitro-7-
sulfamoylbenzo(f)quinoxaline; NMDA, N-methyl-D-aspartate; MK-801, ()-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate.
737

738 Nakagawa et al.
drastically enhance turning. Direct dopamine agonists
such as apomorphine act on up-regulated dopamine receptors
and elicit contralateral turning, whereas indirect
dopaminergic stimulants such as methamphetamine increase
ipsilateral turning (Ungerstedt and Arbuthnott,
1970; Gerlach and Riederer, 1996; Schwarting and Huston,
1996). The potency of dopaminergic agents can be easily
quantified in terms of this turning behavior of unilaterally
lesioned rats. Because drug-induced turning occurs as a
result of dopaminergic receptor supersensitivity, it is difficult
to observe behavioral/motor deficits due to dopamine
depletion within the striatum in the absence of challenge
with dopaminergic agents, and therefore this method cannot
be used for the pharmacological evaluation of nondopaminergic
agents. Moreover, the relation of the druginduced
turning to the clinical symptoms of PD is unclear.
Animals with a unilateral dopaminergic lesion must have
some impairment of contralateral motor function, which
might correspond to clinical PD symptoms, such as akinesia
and bradykinesia. If we can evaluate the degree of the motor
dysfunction and assess the degree to which a drug can improve
it, we may be able to understand better how drugs
work clinically to improve akinesia and bradykinesia. Recently,
a new evaluation method, the so-called stepping test
(Olsson et al., 1995; Kirik et al., 1998; Mukhida et al., 2001),
has been proposed. Although this method can assess akinesia
or rigidity of PD model rats very sensitively, the successful
use of such a model depends on the investigator’s technique.
In this study, we have developed a novel method named the
exploratory Y-maze, with which the lateralization of
hemiparkinsonian rats can be easily assessed in terms of
biased selection of arms in the maze. We have used this
model to examine various antiparkinsonian drugs and compounds,
to see whether they improve the lateralization in
unilaterally 6-OHDA-lesioned rats. We found, interestingly,
that not only dopaminergic agonists but also muscarinic antagonists
alleviated the bias of these rats.
Materials and Methods
Animals. Male Wistar rats (5 weeks old, weighing 130–150 g;
Charles River Japan, Inc., Kanagawa, Japan) were used. Before the
study, rats were acclimatized to quarters maintained at a room
temperature of 23 1°C with 55 5% relative humidity under a
12-h light/dark cycle for 1 week. During periods of acclimatization
and for 4 weeks after operation, solid diet (FM; Oriental Yeast
Industry, Tokyo, Japan) and water were provided ad libitum. During
experiments, the solid diet was limited to 20 g/animal/day for body
weight control (300–400 g).
Surgical Procedure for Medial Forebrain Bundle Lesion.
Animals were anesthetized with pentobarbital (Nembtal; Abbott
Laboratories, Chicago, IL) (50 mg/kg i.p.) and placed in rat stereotaxic
apparatus with the incisor bar fixed at 2.0 mm. Each animal
received a unilateral injection of 4 l of 6-OHDA HCl (Sigma-Aldrich,
St. Louis, MO) (4 mg/ml in 0.1% ascorbic acid) into the right
medial forebrain bundle (MFB) through a stainless steel cannula at
a rate of 0.4 l/min using a microliter syringe pump. The stereotaxic
coordinates, based on Paxinos and Watson (1996), were 2.5 mm
posterior from the bregma, 1.8 mm lateral to the midline, and 7.8
mm from the dura. The cannula was left for 5 min after the end of the
injection to prevent backward flow and to allow for toxin diffusion.
Thirty minutes before and immediately after operation, rats were
injected i.p. with 25 mg/kg desipramine to prevent concurrent damage
to noradrenergic pathways by 6-OHDA infusion (Roberts et al.,
1975).
Lateralization in the Exploratory Y-Maze. Lateralization, a
locomotor bias, was measured in an exploratory Y-maze made of
white acrylic boards, as shown in Fig. 1. The exploratory Y-maze was
placed in a dark room. Each arm was 30 cm in length, 11 cm in width
and 12 cm in height, and was equipped with a photoelectric switch
(type E3S-BT11; Omron Co., Tokyo, Japan) (4 cm above the floor, 18
cm from the tip). At the center of the maze, the space was narrowed,
and the ceiling was lowered. Rats could go only in the direction of
Fig. 1. Schematic illustration of the exploratory
Y-maze. A, upper view of the Y-maze. The center
of the maze is narrowed (8 cm in width). Photo
beams are indicated as broken lines. B, side view
of one arm of this maze. The ceiling of the center
part is low (9.5 cm in height).

Anti-Parkinsonian Drugs Improve Bias in Exploratory Y-Maze 739
Fig. 2. Behavioral analysis of 6-OHDA-lesioned rats in an exploratory Y-maze. Four weeks after the lesions, behavior was analyzed without
administration of any drugs. The average of two behavior analyses is shown. The numbers of naive and lesioned rats were 15 and 71, respectively. ,
p 0.001 compared with naive animals.
their nose at the center, whereas the space in each arm was large
enough for them to turn there. Interruptions of the infrared beams
were recorded through a digital I/O interface (Muromachi Kikai Co.,
Ltd., Tokyo, Japan) at 0.1-s intervals by a computer (PC9801FA or
PC9801RX; NEC, Tokyo, Japan) outside the testing room. The ratio
of choice of the right arm (right%) was measured for 30 min as an
index of lateralization. Otherwise, the number of interruptions of the
beam (total count), the number of entrances from one arm to another
(total turns), and number of counts per turn (counts/turn) were
recorded. Total counts reflect small movements within one arm,
whereas total turns reflect major movement to another arm. However,
both indexes reflect the spontaneous motor activity of the
animals. One more index, the counts/turn, reflects the quality of
the movement. This index indicates the ratio of large and small
movements.
Lesion Verification. Four weeks after the operation, rats were
placed in the exploratory Y-maze, and locomotor bias was measured.
The criteria for a successful lesion and the inclusion of the animal in
subsequent drug studies were set as more than 85% average right%
and more than 80% right% in each of two consecutive measurements.
Drug Treatment. Apomorphine (Sigma-Aldrich) dissolved in
0.1% ascorbic acid and NBQX (Tocris Cookson Inc., Ballwin, MO)
dissolved in saline made slightly alkaline with 0.1 N NaOH were
administered immediately before recording of movements in the
exploratory Y-maze. Methamphetamine (Dainippon Pharmaceutical
Co. Ltd., Osaka, Japan), MK-801 (Sigma-Aldrich), scopolamine (Tokyo
Kasei Kogyo Co., Ltd., Tokyo, Japan), and N-methylscopolamine
(Sigma-Aldrich) were each dissolved in saline. Idazoxan (Sigma-
Aldrich) was suspended in 0.5% methylcellulose. Agents were administered
intraperitoneally 15 min before recording of movements
in the exploratory Y-maze. Levodopa and benserazide (Sigma-Aldrich)
(4:1) were suspended in 0.5% methylcellulose and then administered
intraperitoneally 15 and 45 min before recording, respectively.
Biochemical Analysis. At the end of the experiments, rats were
decapitated and the brain was removed. The whole striata and
frontal cortex were dissected rapidly on ice and immediately frozen
in liquid nitrogen until analysis. Determination of the dopamine and
norepinephrine contents was performed as described previously (Kagaya
et al., 1996). 3,4-Dihydroxybenzylamine was used as an internal
standard. Monoamine levels were analyzed on a 150-mm ODS
column (CA-5 ODS; Eicom, Kyoto, Japan) combined with an electron
capture detector (ECD-100; Eicom), and the assay was conducted by
the internal standard method.
Statistical Analysis. Behavioral data were analyzed by means of
one-way analysis of variance followed by Dunnett’s test or by means
of the F test followed by Student’s t test using the software package
SAS 6.12 (SAS Institute Japan, Tokyo, Japan). In all tests, differences
were considered significant when p was smaller than 0.05.
Results
Effect of MFB Lesion. The effect of MFB lesion was
assessed, using the four indices described under Materials
and Methods, 4 weeks after lesioning. The value of right% in
the naive group was almost at the chance level. As shown in
Fig. 2, right% was significantly increased to about 80% in
lesioned animals (t 11.93; p 0.0001), whereas other
indices, total counts and total turns, showed no change. The
increase of right% was highly reproducible, and did not
change during repeated trials.
Fig. 3. Correlation between lateralization and dopamine content in the
striatum (top) and norepinephrine content in the frontal cortex (bottom).
The ordinate shows dopamine or norepinephrine level as a percentage of
that in the intact side of the striatum or frontal cortex, respectively.
Dopamine and norepinephrine contents in the intact side were 17,755
281 and 1,613 405 ng/g tissue, respectively. The abscissa shows right%
measured in the exploratory Y-maze.

740 Nakagawa et al.
The relationship between right% and dopamine content in
the striatum or norepinephrine content in the frontal cortex
was examined. When animals showed a right% value of more
than 80%, the dopamine content in the striatum was always
less than 10% (Fig. 3, top). When the striatal dopamine
content remained above 10%, it seemed to correlate negatively
with right% values. In some cases, however, severe
depletion of striatal dopamine did not cause lateralization of
more than 80%, although almost all such animals showed
higher right% values than the chance level (50%). On the
other hand, animals having right% values of more than 85%
showed various norepinephrine levels. Thus, norepinephrine
content did not correlate straightforwardly with right% values,
in contrast to dopamine content. The values of the correlation
coefficient between striatum dopamine% and right%
and between cortex norepinephrine% and right% were 0.68
and 0.19, respectively.
Effect of Dopaminergic Drugs. Intraperitoneal administration
of the dopamine precursor levodopa with benserazide
significantly reduced right% at 10 mg/kg (t 2.52; p
0.045) without significant changes of total counts, total
turns, and counts/turn. At a higher dose, 30 mg/kg, right%
was also reduced (t 2.51; p 0.045), whereas total turns
was drastically decreased (t 2.78; p 0.024) and total
counts was increased (t 2.73; p 0.027) (Fig. 4, top). As a
result, counts/turn at 30 mg/kg was markedly increased (t
4.62; p 0.0001). At this dose, we observed that the animals
turned counterclockwise in their home cage before and after
the measurement in the Y-maze.
A short-acting dopamine agonist, apomorphine, was injected
subcutaneously. As shown in Fig. 4, right% was significantly
reduced at 0.1 mg/kg (t 2.80; p 0.022), again
without significant changes in total counts, total turns, and
counts/turn. At a higher dose of 0.3 mg/kg, right% was reduced
(t 5.20; p 0.00002), whereas total turns tended to
be decreased (t 2.06; p 0.125) and the total counts value
was increased (t 3.44; p 0.004). As with levodopa, counts/
turn was significantly increased at this dose (t 5.07; p
0.00003).
On the other hand, an indirect dopamine agonist, methamphetamine,
did not affect right%, whereas it significantly
increased total counts and total turns at 0.5 mg/kg (t 3.16,
3.01; p 0.0067, 0.0097, respectively) (Fig. 4).
Fig. 4. Effects of levodopa, apomorphine, and methamphetamine on behavior of 6-OHDA-lesioned rats. Levodopa and benserazide (4:1) were
administered intraperitoneally 15 and 45 min before recording, respectively. Apomorphine was administered subcutaneously immediately before
recording. Methamphetamine was administered intraperitoneally 15 min before recording. The number of rats per group was 20 for the control and
eight for each group tested with levodopa, apomorphine, or methamphetamine. , p 0.001; , p 0.01; , p 0.05 compared with the control value.

Anti-Parkinsonian Drugs Improve Bias in Exploratory Y-Maze 741
Effects of Anticholinergic Drugs. Trihexyphenidyl, an
anticholinergic drug commonly prescribed for treatment of
early phase PD (Olanow et al., 2001), was tested (Fig. 5).
Interestingly, 10 mg/kg trihexyphenidyl significantly decreased
right% (t 2.70; p 0.019), although it increased
both counts and turns at the same dose (t 3.14, 4.55; p
0.006, 0.00009, respectively). In marked contrast to dopaminergic
drugs, trihexyphenidyl increased total turns and consequently
counts/turn decreased at the same dose (t 4.23;
p 0.00025). To examine whether this effect was related to
the central nervous system, we compared the effect of scopolamine,
which can enter the brain, with that of N-methylscopolamine,
which hardly passes though the blood-brain barrier.
Scopolamine showed almost the same effect as
trihexyphenidyl, whereas N-methylscopolamine had only a
small effect on total counts (Fig. 5).
Effects of Other Drugs under Consideration for
Therapeutic Use in PD. We tested the non-NMDA antagonist
NBQX, the NMDA antagonist MK-801, and the 2
antagonist idazoxan. Among these drugs, only MK-801 at 0.3
mg/kg significantly increased total counts (t 2.87; p
0.017) and total turns (t 2.94; p 0.015). However, this
compound did not improve lateralization at all. Idazoxan and
NBQX did not significantly change any of the indices at 1 or
10 mg/kg, and at 1 or 3 mg/kg, respectively (Fig. 6).
Discussion
In PD patients, 70 to 80% of striatal dopamine content and
50% of dopaminergic neurons have been already lost when
symptoms occur (Dunnett and Björklund, 1999). The symptoms
are asymmetric, with one side more affected than the
other. A comparison of [ 123 I]2--carbomethoxy-3--(4-iodophenyl)-tropane
binding in the striatum contralaterally and
ipsilaterally to the affected body side in patients with
hemiparkinsonism revealed a significant difference (Brücke
et al., 1997), suggesting that a bias of dopaminergic degeneration
causes hemiparkinsonism. When dopaminergic neurons
in experimental animals are damaged unilaterally with
6-OHDA, the animals also show a movement bias. Measurement
of turning behavior has been the most frequent approach
to quantitate the bias and to assess the effects of
dopaminergic drugs. In this study, we could clearly and
quantitatively detect the locomotor bias (lateralization) in
unilaterally 6-OHDA-lesioned rats by using a newly developed
exploratory Y-maze method, in the absence of any drug.
Furthermore, muscarinic antagonists trihexyphenidyl and
scopolamine improved the lateralization in this rat PD
model, like the dopamine agonists levodopa and apomorphine.
In our study, when unilaterally MFB-lesioned rats showed
lateralization, the dopamine content in the striatum was
always less than 10% of that in the nonlesioned side. This is
very similar to the clinical findings in PD, as described above.
However, we found that dopamine depletion did not cause
lateralization in some cases. Rats that were completely depleted
of dopamine and norepinephrine did not always show
complete lateralization, although norepinephrine was still
found to be present in the frontal cortex of animals that
showed right% values of over 85% in the maze (Fig. 3). It is
difficult to explain this finding, but one possibility is that the
lesion causing complete depletion of dopamine and norepinephrine
may have been too severe, damaging other surrounding
neurons. Such damage to other neuronal circuits
might differently affect lateralization in such cases.
Dopaminergic drugs improve PD symptoms, although their
side effects, such as dyskinesia, require caution in clinical
Fig. 5. Effects of trihexyphenidyl and scopolamine on the behavior of 6-OHDA-lesioned rats. Trihexyphenidyl, scopolamine, or N-methylscopolamine
(NMS) was administered intraperitoneally 15 min before recording. The number of rats per group was 20 for the control and 12 for drug-treated
animals. , p 0.01; , p 0.05 compared with the control value.

742 Nakagawa et al.
use. Levodopa and apomorphine reduced the right% value of
hemiparkinsonian rats, suggesting that dopaminergic drugs
are effective to ameliorate the motor imbalance caused by
unilateral dopaminergic lesioning. High dosages of these
drugs increase counts, decrease turns, and result in an increase
in counts/turn (Fig. 4). Before and after we recorded
lateralization in the Y-maze, counterclockwise turning of rats
treated with a high dose of levodopa was frequently observed
in the home cage. This observation suggests that high doses
of dopamine agonists decrease large movements and increase
small movements simultaneously. In 6-OHDA-lesioned rats,
postsynaptic dopamine receptors on the lesioned side are
supersensitive, so that levodopa and apomorphine act on the
lesioned side more efficiently than on the normal side, and
lateralization is improved. When a high dose of dopamine
agonists is applied, dopamine receptors are excessively activated,
in particular in the lesioned side, and so this behavior
is observed, in addition to amelioration of bias. Although
measurement of turning behavior can easily be used to quantify
the potency of dopaminergic drugs, turning is increased
in a manner that depends on the dose of dopamine agonist,
which is not the case with our Y-maze. A major advantage of
our system is that we can assess movement both qualitatively
and quantitatively under each experimental condition.
Regarding the significance of an increase of counts/turn, this
index may reflect certain forms of dyskinesia observed in the
clinical situation. Marsden et al. (1981) reported that, at
least during the initial period of treatment, levodopa-induced
dyskinesias occurred at the same time as the greatest degree
of clinical improvement, and were predominantly choreic in
nature. We think that the phenomena seen with high doses of
levodopa and apomorphine may reflect such clinical symptoms
seen when the effect of the drug is most apparent.
Indirect dopamine agonists, such as methamphetamine,
did not improve lateralization, but increased locomotion.
Methamphetamine increased total counts and total turns
without changing counts/turn. Because nerve terminals of
dopamine neurons on the lesioned side in the striatum of
hemiparkinsonian rats were already lost, methamphetamine
could not enhance the release of dopamine on the lesioned
side, although it did so on the normal side.
Interestingly, anticholinergics improved lateralization of
hemiparkinsonian rats (Fig. 5). Anticholinergics are typically
used in younger PD patients (i.e., 70 years of age or less), in
Fig. 6. Effects of NBQX, MK-801, and idazoxane on the behavior of 6-OHDA-lesioned rats. NBQX was administered immediately before recording. The
other compounds were administrated intraperitoneally 15 min before recording. The number of rats per group was six in the idazoxane study and eight
in the MK-801 and NBQX studies. , p 0.05 compared with control value.

Anti-Parkinsonian Drugs Improve Bias in Exploratory Y-Maze 743
whom tremor is a dominant clinical feature and cognitive
function is preserved (Olanow and Koller, 1998). Because
N-methylscopolamine does not pass through the blood-brain
barrier and did not affect the right% value, the effects are
likely to be mediated through the central cholinergic system.
It has been reported that muscarinic antagonists potentiated
rotation induced by a D1 agonist (Morelli et al., 1993) and
also that muscarinic agonists act functionally like dopamine
antagonists (Fink-Jensen et al., 1998), so muscarinic antagonists
may act functionally like dopamine agonists and improve
lateralization. Domino and Lisong (1998) reported that
trihexyphenidyl enhanced contraversive turning in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced
hemiparkinsonian
monkeys. Muscarinic antagonists were also able to
antagonize reserpine-induced rigidity in rats (Goldstein et
al., 1975). Furthermore, anticholinergics were able to alleviate
akinesia and movement initiation deficits in the
6-OHDA-lesioned rat model (Schallert et al., 1978, 1979).
These results are consistent with our finding that anticholinergics
work in a PD model. In the case of anticholinergics,
counts/turn was decreased at high dosage, which is opposite
to the observation with a dopaminergic agonist. Whereas a
high dosage of scopolamine improved lateralization, scopolamine
at the same dosage is also known to induce amnesia
(Bammer, 1982; Ogura et al., 2000). Although scopolamine
improved lateralization in this study, this effect may not be
useful in the clinical situation, because of the other effects
of anticholinergics (Molchan et al., 1992), such as eliciting
amnesia.
-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid,
NMDA or 2-adrenergic antagonists do not improve the lateralization
in unilaterally lesioned rats. A noncompetitive
-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid antagonist
was reported to improve levodopa-induced dyskinesia
in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine monkeys
(Konitsiotis, 2000). Anti-parkinsonian actions of NMDA
glutamate antagonists have also been reported in rodents
and nonhuman primates (Steece-Collier et al., 2000). In a
clinical study, an NMDA antagonist improved levodopa-induced
dyskinesia (Verhagen Metman et al., 1998), and 2-
adrenergic antagonists such as idazoxan and efaroxan improved
symptoms of PD, particularly rigidity (Brefel-
Courbon, 1998). Based on these study findings, we evaluated
the effects of NBQX, MK-801, and idazoxan on the lateralization
induced by unilateral lesion of the nigro-striatal dopaminergic
system in rats. Much to our surprise, these compounds
failed to improve imbalance of movement when they
were administered alone. Although it is also important to test
them in combination with levodopa, their use in monotherapy
is unlikely to improve imbalance of motor function.
We have established a novel exploratory Y-maze test that
easily detects lateralization in hemiparkinsonian rats. Our
findings indicate that this method can detect anti-parkinsonian
effects of nondopaminergic agents without the use of
dopaminergic agents in combination. Furthermore, we can
characterize the behavior using four indices. A key feature of
the model is that we can distinguish lateralization (indicated
as right%) from an increase of locomotion (detected as total
counts in our system). Some medicines currently used for PD,
e.g., levodopa and trihexyphenidyl, were found to improve
lateralization in our system, whereas others were ineffective.
The new pharmacological evaluation method that we have
developed may therefore be useful in characterizing some
aspects of the clinical efficacy of PD drugs. The method
should contribute to the discovery of new anti-parkinsonian
medicines and should be useful in studies of the mechanisms
underlying the regulation of the dopaminergic system in the
basal ganglia.
References
Bammer G (1982) Pharmacological investigations of neurotransmitter involvement
in passive avoidance responding: a review and some new results. Neurosci Biobehav
Rev 6:247–296.
Brefel-Courbon C, Thalamas C, Saint Paul HP, Senard J-M, Montastruc J-L, and
Rascol O (1998) 2-Adrenoceptor antagonists: a new approach to Parkinson’s
disease? CNS Drugs 10:189–207.
Brooks DJ, Ibanez V, Sawle GV, Quinn N, Lees AJ, Mathias CJ, Bannister R,
Marsden CD, and Frackowiak RS (1990) Differing patterns of striatal 18 F-dopa
uptake in Parkinson’s disease, multiple system atrophy and progressive supranuclear
palsy. Ann Neurol 28:547–555.
Brücke T, Asenbaum S, Pirker W, Djamshidian S, Wenger S, Wöber Ch, Müller Ch,
and Podreka I (1997) Measurement of the dopaminergic degeneration in Parkinson’s
disease with [ 123 I]-CIT and SPECT. Correlation with clinical findings and
comparison with multiple system atrophy and progressive supranuclear palsy.
J Neural Transm Suppl 50:9–24.
Dalvi A and Ford B (1998) Antiparkinsonian agents: clinically significant drug
interactions and adverse effects and their management. CNS Drugs 9:291–310.
Domino EF and Lisong NI (1998) Trihexyphenidyl interactions with the dopamine
D1-selective receptor agonist SKF-82958 and the D2-selective receptor agonist
N-0923 in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced hemiparkinsonian
monkeys. J Pharmacol Exp Ther 284:307–311.
Dunnett SB and Björklund A (1999) Prospects for new restorative and neuroprotective
treatments in Parkinson’s disease. Nature (Lond) 399:A32–A39.
Fink-Jensen A, Kristensen P, Shannon HE, Calligaro DO, Delapp NW, Whitesitt C,
Ward JS, Thomsen C, Rasmusseen T, Sheardown MJ, et al. (1998) Muscarinic
agonists exhibit functional dopamine antagonism in unilaterally 6-OHDA lesioned
rats. Neuroreport 9:3481–3486.
Gerlach M and Riederer P (1996) Animal models of Parkinson’s disease: an empirical
comparison with the phenomenology of the disease in man. J Neural Transm
103:987–1041.
Goldstein JM, Barnett A, and Malick JB (1975) The evaluation of anti-parkinson
drugs on reserpine-induced rigidity in rats. Eur J Pharmacol 33:183–188.
Hornykiewicz O (1966) Dopamine (3-hydroxytyramine) and brain function. Pharmacol
Rev 18:925–964.
Hornykiewicz O (1998) Biochemical aspects of Parkinson’s disease. Neurology 51
(Suppl 2):S2–S9.
Kagaya T, Kajiwara A, Nagato S, Akasaka K, and Kubota A (1996) E2011, a novel,
selective and reversible inhibitor of monoamine oxidase type A. J Pharmacol Exp
Ther 278:243–251.
Kirik D, Rosenblad C, and Björklund A (1998) Characterization of behavioral and
neurodegenerative changes following partial lesions of the nigrostriatal dopamine
system induced by intrastriatal 6-hydroxydopamine in the rat. Exp Neurol 152:
259–277.
Kish SJ, Shannak K, and Hornykiewicz O (1988) Uneven pattern of dopamine loss in
the striatum of patients with idiopathic Parkinson’s disease: pathophysiologic and
clinical implications. N Engl J Med 318:876–880.
Konitsiotis S, Blanchet PJ, Verhagen L, Lamers E, and Chase TN (2000) AMPA
receptor blockade improves levodopa-induced dyskinesia in MPTP monkeys. Neurology
54:1589–1595.
Kopin IJ (1993) The pharmacology of Parkinson’s disease therapy: an update. Annu
Rev Pharmacol Toxicol 32:467–495.
Marsden CD, Parkes JD, and Quinn N (1981) Fluctuations of disability in Parkinson’s
disease - clinical aspects, in Movement Disorders (Marsden CD and Fahn S
eds) pp 96–122, Butterworth Scientific, London.
Molchan SE, Martinez RA, Hill JL, Weingartner HJ, Thompson K, Vitiello B, and
Sunderland T (1992) Increased cognitive sensitivity to scopolamine with age and a
perspective on the scopolamine model. Brain Res Rev 17:215–226.
Morelli M, Fenu S, Cozzolino A, Pinna A, Carta A, and Di Chiara G (1993) Blockade
of muscarinic receptors potentiates D1 dependent turning behavior and c-fos
expression in 6-hydroxydopamine-lesioned rats but does not influence D2 mediated
responses. Neuroscience 53:673–678.
Mukhida K, Baker KA, Sadi D, and Mendez I (2001) Enhancement of sensorimotor
behavioral recovery in hemiparkinsonian rats with intrastriatal, intranigral, and
intrasubthalamic nucleus dopaminergic transplants. J Neurosci 21:3521–3530.
Ogura H, Kosasa T, Kuriya Y, and Yamanishi Y (2000) Donepezil, a centrally acting
acetylcholinesterase inhibitor, alleviates learning deficits in hypocholinergic models
in rats. Methods Find Exp Clin Pharmacol 22:89–95.
Olanow CW and Koller WC (1998) An algorithm (decision tree) for the management
of Parkinson’s disease: treatment guidelines. Neurology 50 (Suppl 3):S1–S57.
Olanow CW, Watts RL, and Koller WC (2001) An algorithm (decision tree) for the
management of Parkinson’s disease (2001): treatment guidelines. Neurology 56
(Suppl 5):S1–S88.
Olsson M, Nikkhah G, Bentlage C, and Björklund A (1995) Forelimb akinesia in the
rat Parkinson model: differential effects of dopamine agonists and nigral transplants
as assessed by a new stepping test. J Neurosci 15:3863–3875.
Paxinos G and Watson C (1996) The Rat Brain in Stereotaxic Coordinates, 3rd ed,
Academic Press, San Diego.
Poewe W (1998) Should treatment of Parkinson’s disease be started with a dopamine
agonist? Neurology 51 (Suppl 2):S21–S24.

744 Nakagawa et al.
Roberts DCs, Zis AP, and Fibiger HC (1975) Ascending catecholamine pathways and
amphetamine-induced locomotor activity: importance of dopamine and apparent
non-involvement of norepinephrine. Brain Res 93:441–454.
Schallert T, De Ryck M, Whishaw IQ, Ramirez VD, and Teitelbaum P (1979)
Excessive bracing reactions and their control by atropine and L-dopa in an animal
analog of parkinsonism. Exp Neurol 64:33–43.
Schallert T, Whishaw IQ, Ramirez VD, and Teitelbaum P (1978) Compulsive, abnormal
walking caused by anticholinergics in akinetic, 6-hydroxydopaminetreated
rats. Science (Wash DC) 199:1461–1463.
Schwarting RK and Huston JP (1996) The unilateral 6-hydroxydopamine lesion
model in behavioral brain research. Analysis of functional deficits, recovery and
treatments. Prog Neurobiol (Oxford) 50:275–331.
Silver DE and Ruggieri S (1998) Initiating therapy for Parkinson’s disease. Neurology
50 (Suppl 6):S18–S22.
Standaert DG and Young AB (1995) Treatment of central nervous system degenerative
disorders, in Goodman & Gilman’s The Pharmacological Basis of Therapeutics,
9th ed. (Hardman JG, Molinoff PB, Ruddon RW, and Goodman Gilman A eds)
pp 503–519, McGraw-Hill Companies, New York.
Steece-Collier K, Chambers LK, Jaw-Tsai SS, Menniti FS, and Greenamyre JT
(2000) Antiparkinsonian actions of CP-101,606, an antagonist of NR2B subunitcontaining
N-methyl-D-aspartate receptors. Exp Neurol 163:239–243.
Takikawa S, Dhawan V, Chaly T, Robeson W, Dahl R, Zanzi I, Mandel F, Spetsieris
P, and Eidelberg D (1994) Input functions for 6-[fluorine-18]fluorodopa quantitation
in parkinsonism: comparative studies and clinical correlations. J Nucl Med
35:955–963.
Ungerstedt U and Arbuthnott GW (1970) Quantitative recording of rotational behavior
in rats after 6-hydroxydopamine lesions of the nigrostriatal dopamine
system. Brain Res 24:485–493.
Verhagen Metman L, Del Dotto P, Natté R, van den Munckhof P, and Chase TN
(1998) Dextromethorphan improves levodopa-induced dyskinesias in Parkinson’s
disease. Neurology 51:203–206.
Watts RL (1997) The role of dopamine agonists in early Parkinson’s disease. Neurology
49 (Suppl 1):S34–S48.
Wichmann T and DeLong MR (1993) Pathophysiology of Parkinsonian motor abnormalities,
in Advances in Neurology, Vol 60: Parkinson’s Disease: from Basic Research
to Treatment (Narabayashi H, Nagatsu T, Yanagisawa N, and Mizuno Y
eds) pp 53–61 Raven Press, New York.
Williams BO and Carlyle D (1979) Levodopa/benserazide (‘Madopar’) combination
therapy in elderly patients with parkinsonism. Curr Med Res Opin 6:1–7.
Address correspondence to: Dr. Makoto Nakagawa, Tsukuba Research
Laboratories, Eisai Co., Ltd., 5-1-3 Tokodai, Tsukuba-shi, Ibaraki 300-2635,
Japan. E-mail: m-nakagawa@hhc.eisai.co.jp