Gen. Pharmac. Vol. 23, No. 3, pp. 337-341, 1992 Printed in Great Britain. All rights reserved
0306-3623/92 $5.00 + 0.00 Copyright © 1992 Pergamon Press Ltd
REM SLEEP DEPRIVATION DECREASES APOMORPHINE-INDUCED STIMULATION OF LOCOMOTOR ACTIVITY BUT NOT STEREOTYPED BEHAVIOR IN MICE WATARU ASAKURA, KINZO MATSUMOTO,HIROYUKI OHTA and HIROSHI WATANABE* Section of Pharmacology, Research Institute for Wakan-Yaku (Oriental Medicines), Toyama Medical and Pharmaceutical University, Toyama 930-01, Japan (Received 7 October 1991)
Abstract--1. Effects of rapid eye movement (REM) sleep deprivation on central dopaminergic system were investigated by testing the behavioral responses to apomorphine and brain dopamine metabolism in mice. 2. REM sleep deprivation for 48 hr significantly suppressed apomorphine. HC1 (3.0 and 6.0 mg/kg, i.p.)stimulated spontaneous locomotor activity without affecting the intensity of stereotyped behavior. 3. Neither the latency of nociceptive response in a hot-plate test nor the duration of pentobarbitalinduced sleep was changed by REM sleep deprivation. 4. Dopamine turnover in the striatum and the nucleus accumbens of REM sleep-deprived mice was significantly higher than that of control animals. 5. These results suggest that REM sleep deprivation may decrease the function of postsynaptic dopamine receptor in the mesolimbic but not nigrostriatal dopaminergic system.
INTRODUCTION
MATERIALS AND METHODS
Deprivation of rapid eye movement (REM) sleep causes several behavioral changes in animals and humans (Vogel, 1975). It enhances dopamine (DA) agonists-induced aggressive behavior (Ferguson and Dement, 1969; Tufik et al., 1978; Tufik, 1981; Mogilnicka, 1981; Zelger and Carlini, 1982), episodic excitation (Trotta, 1984), stereotyped behavior (Tufik et al., 1978; Troncone et al., 1988) and locomotor activity (Arriaga et al., 1988) in rats. Those R E M sleep deprivation-induced changes in the responses to D A agonists have been explained to be due to either supersensitivity of postsynaptic D A receptor (Tufik et al., 1978; Tufik, 1981) or subsensitivity of presynaptic D A receptor (Serra et al., 1981; Tufik et al., 1987) in the rat brain. On the other hand, R E M sleep deprivation has been shown to affect specifcally the function of several DAergic systems. For example, R E M sleep deprivation increases D~ receptor density in mesolimbic area but not striatum in rats (Demontis et al., 1990). However, there are few behavioral evidences that R E M sleep deprivation changes the function of mesolimbic DAergic system. To clarify whether R E M sleep deprivation differently affects the function of nigrostriatal and mesolimbic DAergic system, we investigated the effects of R E M sleep deprivation on apomorphineinduced increase in locomotor activity and stereotyped behavior in mice, and on D A metabolism in the striatum and the mesolimbic area of the mouse brain.
Animals Male ddY mice (4 weeks old) were obtained from Nippon SLC (Shizuoka, Japan) and were housed [20-30 mice per cage (35 x 30 × 16cm)] for at least a week before the experiments. Housing conditions were thermostatically maintained at 25 + I°C, with a 12 hr light/dark cycle (light on at 07:30 a.m.). Food and water were given ad libitum. R E M sleep deprivation REM sleep was deprived using small pedestal (platform) method described by Morden et al. (1967) with slight modification for mice. Briefly, the animal was individually placed in REM sleep deprivation chamber (20 × 15 × 21 cm) at the light period and housed for 48 hr with free access to food and water. The chamber was consisted of a pedestal (1.8 cm dia, 4.5 cm height) surrounded by water (3.5 cm depth). Control mice were socially isolated for 48 hr in the plexiglas cage (25 x 18 × 12cm) with wood chip. After REM sleep deprivation or isolation, each mouse was individually placed in the new plexiglas cage. Behavioral experiments Locomotor activity was measured using animal movement analyzing systems (Scanet SV-10, MATYS, Toyama, Japan). The system consisted of rectangular enclosure (40× 38cm) of which side walls (12cm height) were equipped with 144 pairs of photosensors. Each pair of photosensors was scanned every 0.1 sec to detect animal movements. Locomotor activity was calculated from the scanning data and accumulated every 5min. One hundred minutes after the end of REM sleep deprivation, each mouse was individually placed in a transparent plexiglas cage (25 x 18 × 24 cm) which was fixed at the center of the apparatus, and the locomotor activity was measured. After 80 rain habituation, apomorphine was injected (i.p.). Measurement of stereotyped behavior was carried out in a cage (24 × 12 × 15 cm) of which side walls were made of
*To whom all correspondence should be addressed.
337
338
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Fig. 1. Time-course of apomorphine-induced stimulation of locomotor activity in control and REM sleepdeprived mice. Control ((3) and REM sleep-deprived ( 0 ) mice were housed for 48 hr in a socially isolated cage and a REM sleep deprivation chamber which consists of a small pedestal surrounding water, respectively. One hundred minutes after the end of isolation or REM sleep deprivation, each mouse was individually placed in the plexiglas cage, habituated for 80 min, and given apomorphine. HCI (A, 0.75; B, 1.5; C, 3.0; D, 6.0 mg/kg, i.p.). Locomotor activity was measured every 5 min. The arrow indicates apomorphine injection. Each point represents the mean value of locomotor activity for 5 min obtained from 20 mice. wire mesh (mesh size 1 cm, wire dia 2 mm). Each mouse was individually placed in the cage 100min after the end of REM sleep deprivation, habituated for 80 min, and given apomorphine (i.p.). Rating of stereotyped behavior was
(xl03 ) 16"->
0 / 0
A C
0 d
~0
0 0.
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carried out for 30 sec in a 5 min interval over a 60 min period. Stereotyped behavior was scored as described by Randall and Randall (1986): 0, no drug effect; 1, decreased locomotor activity; 2, intermittent sniffing; 3, continuous sniffing; 4, upright, intermittent sniffing; 5, upright, continuous sniffing; 6, extreme upright position, continuous oral behavior. Behavioral changes were recorded by a video camera for later analysis. The duration of pentobarbital-induced sleep and the latency of nociceptive response in a hot-plate test were measured as described by Matsumoto et al. (1991b) with minor modifications. Briefly, 60 min after the end of REM sleep deprivation, pentobarbital.Na (64mg/kg, i.p.) was administered and sleeping time was measured. In the hotplate test, 10 min after the end of REM sleep deprivation, mice were placed on the hot-plate maintained at 51-53°C. The latency of nociceptive response such as licking or flicking of hind limb or jumping was measured. Catecholamine assay
3J0
610
Apomorphine HCl(mg/kg) Fig. 2. Effect of REM sleep deprivation on locomotor activity stimulated by apomorphine in mice. Open and solid circles represent the data obtained from control and REM sleep-deprived mice, respectively. Each point represents the mean value of locomotor activity for 60 min after apomorphine administration obtained from 20-30 mice. *P < 0.05, **P < 0.01, compared with control mice (Mann-Whitney U-test).
Mice were decapitated 180min after the end of REM sleep deprivation. The brains were rapidly removed and washed in ice-cold saline. The striatum and the nucleus accumbens were dissected on an ice-cold glass plate and frozen in liquid nitrogen. After weighing, each tissue was homogenized in 1 ml ice-cold perchloric acid solution (0.25 M) containing 0.3 mg cysteine and 40 ng 3,4-dihydroxybenzylamine as an internal standard. After centrifugation (10,000g, 4°C for 10min), the supernatant and 30mg acid-washed alumina were added to 1 ml Tris-HC1 buffer (1.5 M, pH 8.6) and shaken (250 rpm) at 4°C for l0 min. The
339
REM deprivation and DAergic system
Table 1. Effect of REM sleep deprivation on the duration of pentobarbital-induced sleep and the latency of nociceptive response in the hot-plate test in mice
30-
O o)
20
o 10
8 O3 I
.75
I
f
I
1.5
3.0
6.0
Apomorphlne. HCI (rng/kg)
Fig. 3. Effect of REM sleep deprivation on apomorphine-induced stereotyped behavior in mice. Open and solid circles represent the data obtained from control and REM sleepdeprived mice, respectively. Each point represents the mean value of total score of stereotyped behavior for 60 min after apomorphine administration obtained from 15 mice. alumina was washed 3 times with distilled water. Catecholamines adsorbed to the alumina were eluated with 0.3 ml perchloric acid solution (0.25M) containing cysteine (0. l mg/ml) and determined with a high performance liquid chromatography with electrochemical detector (HPLCECD).
Drugs Pentobarbital. Na (Toyko Kasei) and apomorphine-HC1 (Sigma) were dissolved in saline and that containing 0.03% ascorbic acid, respectively, just before the experiments. Test drugs were injected i.p. at a volume of 10 ml/kg.
Statistics The significant differences ( P
0306-3623/92 $5.00 + 0.00 Copyright © 1992 Pergamon Press Ltd
REM SLEEP DEPRIVATION DECREASES APOMORPHINE-INDUCED STIMULATION OF LOCOMOTOR ACTIVITY BUT NOT STEREOTYPED BEHAVIOR IN MICE WATARU ASAKURA, KINZO MATSUMOTO,HIROYUKI OHTA and HIROSHI WATANABE* Section of Pharmacology, Research Institute for Wakan-Yaku (Oriental Medicines), Toyama Medical and Pharmaceutical University, Toyama 930-01, Japan (Received 7 October 1991)
Abstract--1. Effects of rapid eye movement (REM) sleep deprivation on central dopaminergic system were investigated by testing the behavioral responses to apomorphine and brain dopamine metabolism in mice. 2. REM sleep deprivation for 48 hr significantly suppressed apomorphine. HC1 (3.0 and 6.0 mg/kg, i.p.)stimulated spontaneous locomotor activity without affecting the intensity of stereotyped behavior. 3. Neither the latency of nociceptive response in a hot-plate test nor the duration of pentobarbitalinduced sleep was changed by REM sleep deprivation. 4. Dopamine turnover in the striatum and the nucleus accumbens of REM sleep-deprived mice was significantly higher than that of control animals. 5. These results suggest that REM sleep deprivation may decrease the function of postsynaptic dopamine receptor in the mesolimbic but not nigrostriatal dopaminergic system.
INTRODUCTION
MATERIALS AND METHODS
Deprivation of rapid eye movement (REM) sleep causes several behavioral changes in animals and humans (Vogel, 1975). It enhances dopamine (DA) agonists-induced aggressive behavior (Ferguson and Dement, 1969; Tufik et al., 1978; Tufik, 1981; Mogilnicka, 1981; Zelger and Carlini, 1982), episodic excitation (Trotta, 1984), stereotyped behavior (Tufik et al., 1978; Troncone et al., 1988) and locomotor activity (Arriaga et al., 1988) in rats. Those R E M sleep deprivation-induced changes in the responses to D A agonists have been explained to be due to either supersensitivity of postsynaptic D A receptor (Tufik et al., 1978; Tufik, 1981) or subsensitivity of presynaptic D A receptor (Serra et al., 1981; Tufik et al., 1987) in the rat brain. On the other hand, R E M sleep deprivation has been shown to affect specifcally the function of several DAergic systems. For example, R E M sleep deprivation increases D~ receptor density in mesolimbic area but not striatum in rats (Demontis et al., 1990). However, there are few behavioral evidences that R E M sleep deprivation changes the function of mesolimbic DAergic system. To clarify whether R E M sleep deprivation differently affects the function of nigrostriatal and mesolimbic DAergic system, we investigated the effects of R E M sleep deprivation on apomorphineinduced increase in locomotor activity and stereotyped behavior in mice, and on D A metabolism in the striatum and the mesolimbic area of the mouse brain.
Animals Male ddY mice (4 weeks old) were obtained from Nippon SLC (Shizuoka, Japan) and were housed [20-30 mice per cage (35 x 30 × 16cm)] for at least a week before the experiments. Housing conditions were thermostatically maintained at 25 + I°C, with a 12 hr light/dark cycle (light on at 07:30 a.m.). Food and water were given ad libitum. R E M sleep deprivation REM sleep was deprived using small pedestal (platform) method described by Morden et al. (1967) with slight modification for mice. Briefly, the animal was individually placed in REM sleep deprivation chamber (20 × 15 × 21 cm) at the light period and housed for 48 hr with free access to food and water. The chamber was consisted of a pedestal (1.8 cm dia, 4.5 cm height) surrounded by water (3.5 cm depth). Control mice were socially isolated for 48 hr in the plexiglas cage (25 x 18 × 12cm) with wood chip. After REM sleep deprivation or isolation, each mouse was individually placed in the new plexiglas cage. Behavioral experiments Locomotor activity was measured using animal movement analyzing systems (Scanet SV-10, MATYS, Toyama, Japan). The system consisted of rectangular enclosure (40× 38cm) of which side walls (12cm height) were equipped with 144 pairs of photosensors. Each pair of photosensors was scanned every 0.1 sec to detect animal movements. Locomotor activity was calculated from the scanning data and accumulated every 5min. One hundred minutes after the end of REM sleep deprivation, each mouse was individually placed in a transparent plexiglas cage (25 x 18 × 24 cm) which was fixed at the center of the apparatus, and the locomotor activity was measured. After 80 rain habituation, apomorphine was injected (i.p.). Measurement of stereotyped behavior was carried out in a cage (24 × 12 × 15 cm) of which side walls were made of
*To whom all correspondence should be addressed.
337
338
WATARU
ASAKURA
(x 102 )
et
al.
(x 102
A ""> ~c 20-
4~ 0
0~0
20
10
C ~
0 LI U v 0 U'
-15
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15
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45
60
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I
I
0
-15
Time (min)
(x 102 )
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1
60
(x 102
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15 30 Time (min)
D
o
20-
/ \o/O..o...o
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, /
10
.J I
-16
0
15 30 Time (min)
45
60
-15
0
I
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16 30 Time (min)
I
I
45
60
Fig. 1. Time-course of apomorphine-induced stimulation of locomotor activity in control and REM sleepdeprived mice. Control ((3) and REM sleep-deprived ( 0 ) mice were housed for 48 hr in a socially isolated cage and a REM sleep deprivation chamber which consists of a small pedestal surrounding water, respectively. One hundred minutes after the end of isolation or REM sleep deprivation, each mouse was individually placed in the plexiglas cage, habituated for 80 min, and given apomorphine. HCI (A, 0.75; B, 1.5; C, 3.0; D, 6.0 mg/kg, i.p.). Locomotor activity was measured every 5 min. The arrow indicates apomorphine injection. Each point represents the mean value of locomotor activity for 5 min obtained from 20 mice. wire mesh (mesh size 1 cm, wire dia 2 mm). Each mouse was individually placed in the cage 100min after the end of REM sleep deprivation, habituated for 80 min, and given apomorphine (i.p.). Rating of stereotyped behavior was
(xl03 ) 16"->
0 / 0
A C
0 d
~0
0 0.
I''~-te i 0 0.75
1.15
carried out for 30 sec in a 5 min interval over a 60 min period. Stereotyped behavior was scored as described by Randall and Randall (1986): 0, no drug effect; 1, decreased locomotor activity; 2, intermittent sniffing; 3, continuous sniffing; 4, upright, intermittent sniffing; 5, upright, continuous sniffing; 6, extreme upright position, continuous oral behavior. Behavioral changes were recorded by a video camera for later analysis. The duration of pentobarbital-induced sleep and the latency of nociceptive response in a hot-plate test were measured as described by Matsumoto et al. (1991b) with minor modifications. Briefly, 60 min after the end of REM sleep deprivation, pentobarbital.Na (64mg/kg, i.p.) was administered and sleeping time was measured. In the hotplate test, 10 min after the end of REM sleep deprivation, mice were placed on the hot-plate maintained at 51-53°C. The latency of nociceptive response such as licking or flicking of hind limb or jumping was measured. Catecholamine assay
3J0
610
Apomorphine HCl(mg/kg) Fig. 2. Effect of REM sleep deprivation on locomotor activity stimulated by apomorphine in mice. Open and solid circles represent the data obtained from control and REM sleep-deprived mice, respectively. Each point represents the mean value of locomotor activity for 60 min after apomorphine administration obtained from 20-30 mice. *P < 0.05, **P < 0.01, compared with control mice (Mann-Whitney U-test).
Mice were decapitated 180min after the end of REM sleep deprivation. The brains were rapidly removed and washed in ice-cold saline. The striatum and the nucleus accumbens were dissected on an ice-cold glass plate and frozen in liquid nitrogen. After weighing, each tissue was homogenized in 1 ml ice-cold perchloric acid solution (0.25 M) containing 0.3 mg cysteine and 40 ng 3,4-dihydroxybenzylamine as an internal standard. After centrifugation (10,000g, 4°C for 10min), the supernatant and 30mg acid-washed alumina were added to 1 ml Tris-HC1 buffer (1.5 M, pH 8.6) and shaken (250 rpm) at 4°C for l0 min. The
339
REM deprivation and DAergic system
Table 1. Effect of REM sleep deprivation on the duration of pentobarbital-induced sleep and the latency of nociceptive response in the hot-plate test in mice
30-
O o)
20
o 10
8 O3 I
.75
I
f
I
1.5
3.0
6.0
Apomorphlne. HCI (rng/kg)
Fig. 3. Effect of REM sleep deprivation on apomorphine-induced stereotyped behavior in mice. Open and solid circles represent the data obtained from control and REM sleepdeprived mice, respectively. Each point represents the mean value of total score of stereotyped behavior for 60 min after apomorphine administration obtained from 15 mice. alumina was washed 3 times with distilled water. Catecholamines adsorbed to the alumina were eluated with 0.3 ml perchloric acid solution (0.25M) containing cysteine (0. l mg/ml) and determined with a high performance liquid chromatography with electrochemical detector (HPLCECD).
Drugs Pentobarbital. Na (Toyko Kasei) and apomorphine-HC1 (Sigma) were dissolved in saline and that containing 0.03% ascorbic acid, respectively, just before the experiments. Test drugs were injected i.p. at a volume of 10 ml/kg.
Statistics The significant differences ( P
