Neuroscience and Behavioral Physiology, Vol. 44, No.

3, March, 2014

Effects of Short-Term Administration of Haloperidol on

the Background of Impairments to Brain Metabolism
and Neuron Activity
F. V. Kopytova and E. L. Dovedova

Translated from Rossiiskii Fiziologicheskii Zhurnal imeni I. M. Sechenova, Vol. 98, No. 6, pp. 706713,
June, 2012. Original article submitted December 16, 2011.
The effects of amphetamine and haloperidol on spike activity levels, the ability to acquire a conditioned
defensive reflex to time, and the state of components of the dopaminergic system in neurons in the sensorimotor cortex and caudate nucleus in rabbits were studied. Chronic administration of amphetamine
increased the level of neuron spike activity and the ability to acquire and reproduce conditioned reflexes
in the sensorimotor cortex, and decreased these parameters in the caudate nucleus. Biochemical studies
demonstrated hyperactivation of the dopaminergic system in the caudate nucleus, as compared with the
sensorimotor cortex. On the background of chronic amphetamine, haloperidol induced effects in both
locations which were close to those of single doses to intact animals. The neuroleptic led to suppression
of neuron activity and a decrease in the number of conditioned reactive neurons in the sensorimotor cortex, in contrast to neurons in the caudate nucleus. Short-term exposure to haloperidol produced changes
in biochemical parameters towards normalization of neurotransmitter metabolism.
Keywords: brain neurons, conditioned reflex activity, sensorimotor cortex, caudate nucleus, amphetamine, haloperidol,
dopaminergic system.

Complex studies using physiological and biochemical

methods have addressed the comparative aspects of
phenamine intoxication and its elimination by the shortterm effects of haloperidol on neurons in the synaptic and
caudate nucleus, which are directly linked not only with the
realization of motor acts, but also the organization of the
integrative mechanisms of the brain [14, 15]. A suitable
model for studying the actions of psychotropic substances
on the functioning of neurons is provided by the formation
of cellular analogs of conditioned reflexes to time which
arise during the performance of motor conditioned reflexes
in animals previously subjected to chronic phenamine
intoxication accompanied by derangements of higher nervous activity, with correction of these derangements using
the neuroleptic haloperidol.

Data from studies of dopamine metabolism are presented as a means of analyzing the brain cell activity data, as
the dopaminergic system is the primary target in the mechanism of action of these pharmacological agents [1, 12, 13].
METHODS
Physiological experiments were performed on 11 conscious non-immobilized rabbits. Neuron spike activity in
the sensorimotor cortex and caudate nucleus was recorded
extracellularly with glass microelectrodes filled with 2.5 M
KCl solution. Motor conditioned reflexes to time (CRT)
were developed to the combination of sounds (clicks, 1020
per sec) with electrocutaneous (square-wave impulses,
1 msec, frequency 4050 per sec, 46 V) stimulation of the
animals forelimb, presented at constant 30-sec intervals.
The conditioned stimulus was presented for 2 sec, and electrocutaneous stimulation started after 1.5 sec exposure to
sound only and continued for 0.5 sec. Conditioned responses were identified as activity occurring at the site of the

Neurology Scientific Center, Russian Academy of Medical

Sciences, 5 Obukha Per., 103064 Moscow, Russia;
e-mail: kopytova_FV@mail.ru.

348
0097-0549/14/4403-0348 2014 Springer Science+Business Media New York

Effects of Short-Term Administration of Haloperidol

349

Fig. 1. Frequency characteristics of spike activity of sensorimotor cortex and caudate nucleus neurons in the brains of
intact animals (a) and on the background of chronic administration of amphetamine(b) and single doses of haloperidol in phenamine intoxication (c). Columns show proportions of neurons (%) with the corresponding numbers of
spikes/sec: 00.01, 0.021, 25, and 625.

combinations when they were missed at the specified 30sec time intervals (15 sequential misses). Response significance was assessed using the WilcoxonMannWhitney U
test (p < 0.5) [4]. The responses of each recorded neuron to
control presentations of stimuli of combined modalities at
different stages of development of the CRT and on treatment with amphetamine and haloperidol were analyzed.
Four series of experiments were performed: 1) studies
of the CRT of neurons in the sensorimotor cortex and caudate nucleus in control animals; 2) creation of chronic phenamine intoxication (10 mg/kg i.p. for 68 days) followed
by recording of spike activity from neurons in the cortex
and caudate nucleus; 3) administration of single doses
(0.20.4 mg/kg i.v.) of haloperidol on the background of
phenamine intoxication for subsequent analysis of the
antipsychotic action of the neuroleptic in cellular analogs
(CRT); 4) single doses of haloperidol at the same dose, in
control animals for analysis of the intrinsic psychotropic
effects of the neuroleptic. During physiological experiments, the activity of 90 neurons in the sensorimotor cortex
and 82 neurons in the striatum were recorded in 11 animals.
The microelectrode insertion coordinates in the striatum were defined using the Fifkova and Marshal atlas. After
experiments, the position of the recording microelectrode
was identified by electrolytic labeling on brain slices
stained by the Nissl method.
Biochemical experiments were performed on 30
Wistar rats (male, body weight 180240 g). The animals
were divided into three equal groups. One group received
daily i.p. amphetamine solution (2.5 mg/kg) for 21 days, the
second received single doses of haloperidol (0.5 mg/kg),
and the third group received physiological saline. At 60 min

after administration of single doses, animals were decapitated under light ether anesthesia. The brain was extracted
in the cold and the sensorimotor cortex and caudate nucleus were harvested. Dopamine (DA) and the end product of
transmitter metabolism homovanillic acid (HVA) were
assayed fluorimetrically in brain homogenates. DA fluorescence was measured at 330/420 nm and HVA fluorescence
at 330/410 nm [7]. Biogenic amine contents were expressed
as ng/mg tissue. Differential centrifugation was used to isolate subcellular fractions: fraction 1 was pelleted at 10000 g
(20 min) and was used for assay of monoamine oxidase
(MAO), expressed as D450/mg protein in samples at 60 min
[3]. Fraction 2 was pelleted at 20000 g (15 min) and was
used for assay of tyrosine hydroxylase (TyrHd) in terms of
the oxidation of 6-methyltetrahydropterin, the cofactor of
the hydroxylation reaction, and was expressed as D335/mg
protein in samples at 60 min [11]. Enzyme activities were
measured spectrophotometrically. Protein contents were
estimated using the Lowry method. Data were analyzed statistically using Statistica 6.0 to run analysis of variance
(ANOVA), with identification of differences between
groups using a posteriori tests (Fishers post hoc LSD test).
RESULTS
Analysis of spontaneous activity, which is a specific
measure of the functional state of structures, demonstrated
a wide range of neuron discharge frequencies in the sensorimotor cortex and caudate nucleus in normal conditions
and in conditions of various pharmacological actions.
Neuron spike activity in the study structures recorded
in control animals differed in terms of frequency spectra.
Cortical neurons had a higher level of discharge activity,

Effects of Short-Term Administration of Haloperidol

349

Fig. 1. Frequency characteristics of spike activity of sensorimotor cortex and caudate nucleus neurons in the brains of
intact animals (a) and on the background of chronic administration of amphetamine(b) and single doses of haloperidol in phenamine intoxication (c). Columns show proportions of neurons (%) with the corresponding numbers of
spikes/sec: 00.01, 0.021, 25, and 625.

combinations when they were missed at the specified 30sec time intervals (15 sequential misses). Response significance was assessed using the WilcoxonMannWhitney U
test (p < 0.5) [4]. The responses of each recorded neuron to
control presentations of stimuli of combined modalities at
different stages of development of the CRT and on treatment with amphetamine and haloperidol were analyzed.
Four series of experiments were performed: 1) studies
of the CRT of neurons in the sensorimotor cortex and caudate nucleus in control animals; 2) creation of chronic phenamine intoxication (10 mg/kg i.p. for 68 days) followed
by recording of spike activity from neurons in the cortex
and caudate nucleus; 3) administration of single doses
(0.20.4 mg/kg i.v.) of haloperidol on the background of
phenamine intoxication for subsequent analysis of the
antipsychotic action of the neuroleptic in cellular analogs
(CRT); 4) single doses of haloperidol at the same dose, in
control animals for analysis of the intrinsic psychotropic
effects of the neuroleptic. During physiological experiments, the activity of 90 neurons in the sensorimotor cortex
and 82 neurons in the striatum were recorded in 11 animals.
The microelectrode insertion coordinates in the striatum were defined using the Fifkova and Marshal atlas. After
experiments, the position of the recording microelectrode
was identified by electrolytic labeling on brain slices
stained by the Nissl method.
Biochemical experiments were performed on 30
Wistar rats (male, body weight 180240 g). The animals
were divided into three equal groups. One group received
daily i.p. amphetamine solution (2.5 mg/kg) for 21 days, the
second received single doses of haloperidol (0.5 mg/kg),
and the third group received physiological saline. At 60 min

after administration of single doses, animals were decapitated under light ether anesthesia. The brain was extracted
in the cold and the sensorimotor cortex and caudate nucleus were harvested. Dopamine (DA) and the end product of
transmitter metabolism homovanillic acid (HVA) were
assayed fluorimetrically in brain homogenates. DA fluorescence was measured at 330/420 nm and HVA fluorescence
at 330/410 nm [7]. Biogenic amine contents were expressed
as ng/mg tissue. Differential centrifugation was used to isolate subcellular fractions: fraction 1 was pelleted at 10000 g
(20 min) and was used for assay of monoamine oxidase
(MAO), expressed as D450/mg protein in samples at 60 min
[3]. Fraction 2 was pelleted at 20000 g (15 min) and was
used for assay of tyrosine hydroxylase (TyrHd) in terms of
the oxidation of 6-methyltetrahydropterin, the cofactor of
the hydroxylation reaction, and was expressed as D335/mg
protein in samples at 60 min [11]. Enzyme activities were
measured spectrophotometrically. Protein contents were
estimated using the Lowry method. Data were analyzed statistically using Statistica 6.0 to run analysis of variance
(ANOVA), with identification of differences between
groups using a posteriori tests (Fishers post hoc LSD test).
RESULTS
Analysis of spontaneous activity, which is a specific
measure of the functional state of structures, demonstrated
a wide range of neuron discharge frequencies in the sensorimotor cortex and caudate nucleus in normal conditions
and in conditions of various pharmacological actions.
Neuron spike activity in the study structures recorded
in control animals differed in terms of frequency spectra.
Cortical neurons had a higher level of discharge activity,

350

Fig. 2. Effects of amphetamine and haloperidol on the ability of sensorimotor cortex and caudate nucleus neurons to acquire conditioned reactions
to time. Columns show proportions of neurons producing conditioned
responses (%) in intact animals (a), in chronic administration of
amphetamine (b), after haloperidol on the background of amphetamine (c),
and after single doses of haloperidol (d).

and the proportion of neurons with frequencies of 225

spikes/sec was greater than 70%, while only 28% were lowfrequency cells (less than 1 spike/sec). Caudate nucleus
neurons were characterized by a low level of baseline discharge activity, with long interspike intervals. About 70% of
cells had frequencies of less than 1 spike/sec (Fig. 1, a). Phenamine intoxication led to motor restlessness on the part of
the animals, with decreased thresholds of forelimb motor
reactions to electrocutaneous stimulation by factors of 35
compared with the control level before amphetamine.
Rabbits which had acquired motor CRT 1520 min
after single doses of amphetamine (i.v., 0.510 mg/kg)
showed stereotypical flexor movements of the forelimbs,
which persisted for 4050 min. Neuron activity in the cortex and striatum was recorded during development of the
CRT, (Fig. 2, a). Phenamine intoxication did not induce
marked changes in cortical neuron activity. As combinations were presented, the number of conditioned-responding neurons increased from the control level of 51% to 63%
(Fig. 2, b), and the reproduction of trace processes also
became more intense and prolonged.
Chronic amphetamine produced significant suppression
of neuron spike activity in the caudate nucleus (on average,
from 1.5 to 0.7 spikes/sec) (Fig. 1, b). However, about 53%
of spontaneously active striatum cells retained the ability to
acquire the CRT and reproduced 60% of trace responses
(Fig. 2). Conditioned responses to 16 misses were insignif-

Kopytova and Dovedova

icant, though their intensity to subsequent misses increased
sharply and persisted through the following 15 misses.
Biochemical experiments addressed the state of components of the DA system in the sensorimotor cortex and
caudate nucleus in terms of measures of the activity of the
enzymes synthesizing (TyrHd) and catabolizing (MAO)
neurotransmitters, as well as the concentrations of DA and
the end product of its metabolism (HVA) in normal concentrations and on exposure to pharmacological agents in vivo.
In conditions of prolonged amphetamine administration, the sensorimotor cortex and caudate nucleus showed
significant activation of neurotransmitter system study
parameters (Fig. 3).
More marked increases in enzyme activity (TyrHd
158%; MAO 158%), as well as in DA levels (160%) and
HVA levels (240%) were seen in the caudate nucleus than in
the cortex (TyrHd 110%; MAO 117%; DA 130%; HVA
220%) (see Table 1).
These data provide evidence of clear hyperfunction of
the neurotransmitter system in terms of the activities of the
enzymes synthesizing(TyrHd) and catabolizing (MAO)
neurotransmitters and in terms of DA and its metabolic end
product (HVA) contents.
Administration of haloperidol on the background of
chronic amphetamine led to decreases in both the frequency
of cortical spike discharges, on average from 5 to 2.6 spikes
per sec, and the reproduction of trace processes, with the CRT
level decreasing from 51% to 40% (Fig. 1, c; Fig. 2, a, c).
In contrast to this, the spontaneous activity of caudate
nucleus neurons increased in conditions of haloperidol on the
background of phenamine intoxication, on average from 1.5
to 4.1 spikes/sec, and the number of initially active cells able
to reproduce trace reactions increased to 76% from 60%, as
after single doses of haloperidol (Fig. 1; Fig. 2, a, c, d).
Administration of single doses of haloperidol to control
animals had no effect on their behavior, though the neuroleptic produced a marked increase in the level of spontaneous neuron activity in the caudate nucleus (from 1.5 to 2.8
spikes/sec) and some reduction in the sensorimotor cortex
(from 5 to 3.4 spikes/sec) (Fig. 1). As compared with intact
animals, there was an increase in the number of neurons in
the striatum reproducing the CRT from 60% to 78%, while
there was a decrease from 50% to 35% in the sensorimotor
cortex (Fig. 2).
In conditions of short-term administration of haloperidol, there was some increase in the activity of the components of the dopaminergic system in both of the structures
studied as compared with control levels (see Table 1).
It follows from the data presented in Table 1 that the
effects of haloperidol were different in the brain structures
studied. In the sensorimotor cortex, increases in the components of the DA system on exposure to haloperidol were
minor as compared with controls: TyrHd 110%; MAO
114%; DA 106%. In the caudate nucleus, these changes
were greater, especially in relation to the enzyme systems:

Effects of Short-Term Administration of Haloperidol

351

Fig. 3. Effects of amphetamine and haloperidol on the activities of components of the dopaminergic system in the sensorimotor cortex and
caudate nucleus in rat brain. 1) Tyrosine hydroxylase; 2) monoamine oxidase; 3) dopamine; 4) homovanillic acid.

TABLE 1. Characteristics of the Dopaminergic System in Normal Conditions and on Exposure to

Amphetamine and Haloperidol in Vivo in the Rat Brain

Notes. Data are shown as M m, where M is the mean and m is mean square deviation, *p < 0.05
statistically significant differences from control (ANOVA, a posteriori Fishers test). For units see
Methods section.

TyrHd 152%; MAO 146%. At the same time, the DA

content was close to normal 91% (Fig. 3).
This result appears to reflect increased utilization of
neurotransmitter on interaction with the receptor apparatus,
as the main element in the mechanism of action of haloperidol is its action on D2 receptors [12, 21].
DISCUSSION
Comparative analysis of the actions of amphetamine
and haloperidol in different structures identified the characteristics of neuron reactions and neurotransmitter metabolism in the study areas of the brain.
In phenamine intoxication, the intensity of the reproduction of traces of previous combinations by sensorimotor

cortex neurons showed marked increases as compared with

normal, especially in the later (6th9th) misses. The spike
activity of most caudate nucleus neurons was suppressed on
exposure to amphetamine, and this effect has been reported
by other authors [16, 17]. These changes in the cortex were
relatively small, and appeared to occur as a result of weakening of the effect of the striatum on the cortex [15, 17, 22].
However, some initially active caudate nucleus cells
were able to acquire the CRT, and trace responses reached
a maximum in the first six misses as compared with the
level of responses to combinations.
We have previously seen this effect in putamen neurons [8] this is a structure with similar morphology and
histology to the caudate nucleus [15, 18]. Our studies noted

352
increased activity in dopamine synthesis and catabolism,
which was particularly apparent in biochemical processes
in the caudate nucleus.
During exposure to amphetamine, the effect of
haloperidol in the brain structures studied here produced
some shift in the level and dynamics of the intensity of conditioned neuron reactions towards normalization. The opposite direction of the effect of haloperidol as compared with
that of amphetamine was also apparent in measures of the
dopaminergic system, which was particularly clear in relation to DA and HVA contents, as shown in Fig. 3. This tendency to changes in components of the DA system appears
also to occur on the background of chronic amphetamine, as
supported by our physiological experiments.
Published data show that the action of haloperidol has
phasic characteristics over time and short-term haloperidol
in rats produces positive effects at 60120 min on neurotransmitter metabolic activity, as shown in our studies, and
this appears to be compensatory in nature [5].
Medical treatment (administration of amphetamine,
haloperidol) produces changes not only to dopamine metabolism, but also to the metabolism of other biogenic amines,
particularly acetylcholine, and the interaction between neurotransmitter systems is impaired; along with DA, this may
be involved in regulating membrane and synapse properties,
transforming the excitation arising in the structure. In the
present experiments, hyperactivation of the DA system
probably suggests weakening of the ACh system, leading to
impairment of the animals motor activity (production of
stereotypical movements); conversely, motor functions may
be activated by normalization of DA metabolism in conditions of short-term administration of the neuroleptic.
A number of experimental studies have demonstrated a
reciprocal interaction between the dopaminergic and acetylcholine systems [16, 19, 20]. Thus, prolonged administration of haloperidol in experimental parkinsonism leads to
depletion of DA, with activation of acetylcholine metabolism: transmitter synthesis increases(choline acetyltransferase activity) and acetylcholinesterase activity is suppressed [2, 10]. In the conditions of our experiments,
hyperactivation of the DA system may suggest weakening
of the ACh system, leading to impairment of the animals
motor activity (occurrence of stereotypical movements)
and, conversely, motor functions may be activated by normalization of DA metabolism in conditions of short-term
administration of the neuroleptic.
Thus, our data are supported at the biochemical level
in the mechanisms of neurotransmitter metabolism, especially the dopamine system, which is the target for the
action of the pharmacological agents used here. Our results
can be regarded as an experimental neurophysiological and
biochemical model appropriate for the features of the structural-functional organization of the brain and for subsequent pharmacological analysis of the mechanisms of
action of psychotropic substances and antipsychotic effects.

Kopytova and Dovedova

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