O36l-9230/‘)1
Brain Reseawh Bulltrin. Vol. 26. pp. 215-218. ‘CPergamon Press pk. 1991. Printed m the U.S.A.
%3.00 i
.on
Medial Preoptic Area Affects Sleep-Wakefulness Independent of Associated Body Temperature Change in Free Moving Rats B. N. MALLICK School
of
Life Sciences,
Jawaharlal
AND M. N. ALAM
Nehru University. New Delhi 110 067, Indiu
Received
19 June 1990
MALLICK, B. N. AND M. N. ALAM. Medial preoptic area nfects sleep-wakqfilness independent of ussociated body remperuture change in free moving rats. BRAIN RES BULL 26(2) 215-218, 1991. --Sleep-wakefulness and body temperature may modu-
late each other. Though both the functions are influenced by the medial preoptic area, the mechanism of action was not clear. This study was aimed at finding out whether the tonic influence of the medial preoptic area on sleep-wakefulness was independent of or secondary to simultaneous change in body temperature. The effects of inactivation of the area by a long acting local anaesthetic, marcain. on those physiological functions were investigated during the night and the day in freely moving rats. Though medial preoptic area influenced sleep-wakefulness and body temperature simultaneously, the effect on the latter was prolonged. The results suggest that the influence on sleep-wakefulness is unlikely to be associated with simultaneously changing body temperature. However, this study fails to differentiate whether the observed effects were due to inactivation of the cell body or the fibers passing through the area. Medial preoptic area
Sleep
Wakefulness
Rectal temperature
INDEPENDENT studies have shown that the medial preoptic area (mPOA) plays an important role in the regulation of body temperature (2,5) and sleep-wakefulness (15, 16, 21). It is widely accepted that sleep-waking phenomena are closely related to body and brain temperature. Hence, it is possible that mPOA-mediated changes in the former might be secondary to changes in the latter or vice versa (17.23). Nevertheless, the possibility of their independent regulation cannot be ignored (I, 4, 13). Single neuronal activity studies indirectly support the latter view. Though the temperature-sensitive neurons (2, 5, 6) and neurons (10) related to sleep (S) and wakefulness (AW) and related EEG changes (11) are reported in the mPOA, the same neuron may not be sensitive to both the physiological parameters (18). Further, it has been reported that mPOA neurons influenced by induced phasic changes in the cortical EEG may not necessarily follow the pattern exhibited during spontaneous alteration in the EEG (12,14). Since evidence confirming either of the mechanisms was lacking, this study attempted to investigate, by reversible inactivation of the mPOA in the freely moving rats, if the tonic influence of mPOA on SAW is secondary to or independent of simultaneous change in the body temperature response.
Inactivation
Marcain
during the night and the day (Table 1). Before surgery rats were acclimatized to the recording environment and the recta1 probe for a minimum of two sessions of at least three hours each. Chemitrode assembly was implanted stereotaxically (19) towards the mPOA under IP (35 mg/kg) Nembutal (sodium pentobarbitone, Loba-Chemie Indo-Austranal Co., India) anaesthesia and aseptic conditions. Bilateral electrodes for recording EEG, EMG and EOG were implanted in three rats each. of groups 3 and 6. Recording was started after at least four days recovery. From the third recovery day rats were maintained in the recording cage with recording plug and rectal probe connected for at least five h per day for acclimatization. As saline and 0.2 p,l marcain were ineffective on S-AW (I), their effects were studied on rectal temperature (Tree) only, while the effects of 0.4 ~1 marcain (which influenced S-AW) were studied simultaneously on both S-AW and Tree in three rats each, of groups 3 and 6 (Table 1). On the day of experiment, the rat with rectal probe (inserted 6 cm into the rectum and connected to a digital thermometer, CT 802, Century, India) and recording plug was left in the recording cage (ambient temperature 26 5 1°C) for a minimum period of one hour before the recording was started. EEG, EMG and EOG were recorded continuously, while Tree was recorded every 5 min. Baseline recording was done for a minimum of 30 min followed by recording after single dose local infusion either of 0.4 ~1 saline (control) or 0.2/0.4 p,l 1% marcain (bupivacaine hydrochloride, Sarabhai Chemicals, India) bilaterally into the mPOA at the rate of 0. I kl/min. Postinjection recordings were continued
METHOD
Experiments were conducted on male Wistar rats (2.50-350 g) bred and maintained in the animal house with food and water ad lib. Rats were divided into six groups and studies were made 215
MALLICK
A
NIGHT
(MARCAIN)
AND
Ah&l
LIZI
Tree AW
o---o
40.0r
I
1
39.6
-
180 i
38.8
38.0
1
ml_ 10’ 20'30'f
-
Bregma-0.3mm
-‘lo
B
ki
e z
2 Deqma-0.8mmJ
L
40 20
0
' ' 20'30'60'50'60'70'B
or
DAY (MARCAIN)
it
+ +
40.0 I:
39.6
100
15
%d
1
VI 2z +I
Rz
+’ + +
6
ox
60
1-
38.4k ‘i?
10 0
- 80 --"
5 :
- 60
,l,j’
- 40 - 20
765432101234567
PIG. 1. Reconstruction of histological sections of rat brain according to the atlas of Paxinos and Watson. The filled areas show extension of 0.4 ~1 marcain injected into the mPOA which was effective in influencing SAW and Tree. Extension of 0.4 pl saline and 0.2 pl marcain was within the same boundary. Ineffective injection sites are shown by hatched circles. Though injections were made bilaterally, it is shown on one side only. The demarcation of mPOA is shown on the opposite side according to the atlas. Abbreviations: ac-anterior commissure; OX-optic chiasma; AHy-anterior hypothalamus; 3V-third ventricle; VDBD-nu vertical limb diagonal band, dors.
until the responses returned to the baseline. Sleep-wakefulness was recorded for 100 min while the Tree for 5-7 h depending on 0.2 or 0.4 ~1 marcain infused. In addition, as a control study, Tree for five rats each from experimental (without injection) and normal (without surgery) groups was monitored for the duration corresponding to the recording period, both during the night and the day. At the end of experiments, rats were sacrificed under deep anaesthesia and the site along with spread of infused chemical (Fig. 1) were histologically identified (1). The data were analyzed statistically (3). Effects of marcain on SAW were analyzed (1). The effect of marcain on the Tree was compared with that of 1) the baseline; 2) the postsaline injection; 3) the Tree without injection; and 4) the Tree after injection of 0.210.4 ~1 marcain. The level of significance was determined by applying Wilcoxon matched-pair signed-rank test for 1) mentioned above and Mann-Whitney test for the rest.
37.&t
m
kid
PRE-INJ PERIOD
-POST-INJECTION PERIOD
10'20'30'*
I
10'20'
50'60' 70'80'91
-0
+
INJECTION PIG. 2. The figure shows the mean rectal temperature (Tree) and the mean percent time spent in wakefulness (AW) by the rats before and after 0.4 pl marcain into the mPOA bilaterally during night (A) and day (8) recording. Marcain precipitatedS during the night, while AW during the day though the Tree showed an increase during both the times and continued even when the S-AW returned to a nonsignificant level as compared to that of preinjection values. **, an increase in Tree; + +, an increase in AW; and --, a decrease in AW as compared to preinjection at the significance level of p
Brain Reseawh Bulltrin. Vol. 26. pp. 215-218. ‘CPergamon Press pk. 1991. Printed m the U.S.A.
%3.00 i
.on
Medial Preoptic Area Affects Sleep-Wakefulness Independent of Associated Body Temperature Change in Free Moving Rats B. N. MALLICK School
of
Life Sciences,
Jawaharlal
AND M. N. ALAM
Nehru University. New Delhi 110 067, Indiu
Received
19 June 1990
MALLICK, B. N. AND M. N. ALAM. Medial preoptic area nfects sleep-wakqfilness independent of ussociated body remperuture change in free moving rats. BRAIN RES BULL 26(2) 215-218, 1991. --Sleep-wakefulness and body temperature may modu-
late each other. Though both the functions are influenced by the medial preoptic area, the mechanism of action was not clear. This study was aimed at finding out whether the tonic influence of the medial preoptic area on sleep-wakefulness was independent of or secondary to simultaneous change in body temperature. The effects of inactivation of the area by a long acting local anaesthetic, marcain. on those physiological functions were investigated during the night and the day in freely moving rats. Though medial preoptic area influenced sleep-wakefulness and body temperature simultaneously, the effect on the latter was prolonged. The results suggest that the influence on sleep-wakefulness is unlikely to be associated with simultaneously changing body temperature. However, this study fails to differentiate whether the observed effects were due to inactivation of the cell body or the fibers passing through the area. Medial preoptic area
Sleep
Wakefulness
Rectal temperature
INDEPENDENT studies have shown that the medial preoptic area (mPOA) plays an important role in the regulation of body temperature (2,5) and sleep-wakefulness (15, 16, 21). It is widely accepted that sleep-waking phenomena are closely related to body and brain temperature. Hence, it is possible that mPOA-mediated changes in the former might be secondary to changes in the latter or vice versa (17.23). Nevertheless, the possibility of their independent regulation cannot be ignored (I, 4, 13). Single neuronal activity studies indirectly support the latter view. Though the temperature-sensitive neurons (2, 5, 6) and neurons (10) related to sleep (S) and wakefulness (AW) and related EEG changes (11) are reported in the mPOA, the same neuron may not be sensitive to both the physiological parameters (18). Further, it has been reported that mPOA neurons influenced by induced phasic changes in the cortical EEG may not necessarily follow the pattern exhibited during spontaneous alteration in the EEG (12,14). Since evidence confirming either of the mechanisms was lacking, this study attempted to investigate, by reversible inactivation of the mPOA in the freely moving rats, if the tonic influence of mPOA on SAW is secondary to or independent of simultaneous change in the body temperature response.
Inactivation
Marcain
during the night and the day (Table 1). Before surgery rats were acclimatized to the recording environment and the recta1 probe for a minimum of two sessions of at least three hours each. Chemitrode assembly was implanted stereotaxically (19) towards the mPOA under IP (35 mg/kg) Nembutal (sodium pentobarbitone, Loba-Chemie Indo-Austranal Co., India) anaesthesia and aseptic conditions. Bilateral electrodes for recording EEG, EMG and EOG were implanted in three rats each. of groups 3 and 6. Recording was started after at least four days recovery. From the third recovery day rats were maintained in the recording cage with recording plug and rectal probe connected for at least five h per day for acclimatization. As saline and 0.2 p,l marcain were ineffective on S-AW (I), their effects were studied on rectal temperature (Tree) only, while the effects of 0.4 ~1 marcain (which influenced S-AW) were studied simultaneously on both S-AW and Tree in three rats each, of groups 3 and 6 (Table 1). On the day of experiment, the rat with rectal probe (inserted 6 cm into the rectum and connected to a digital thermometer, CT 802, Century, India) and recording plug was left in the recording cage (ambient temperature 26 5 1°C) for a minimum period of one hour before the recording was started. EEG, EMG and EOG were recorded continuously, while Tree was recorded every 5 min. Baseline recording was done for a minimum of 30 min followed by recording after single dose local infusion either of 0.4 ~1 saline (control) or 0.2/0.4 p,l 1% marcain (bupivacaine hydrochloride, Sarabhai Chemicals, India) bilaterally into the mPOA at the rate of 0. I kl/min. Postinjection recordings were continued
METHOD
Experiments were conducted on male Wistar rats (2.50-350 g) bred and maintained in the animal house with food and water ad lib. Rats were divided into six groups and studies were made 215
MALLICK
A
NIGHT
(MARCAIN)
AND
Ah&l
LIZI
Tree AW
o---o
40.0r
I
1
39.6
-
180 i
38.8
38.0
1
ml_ 10’ 20'30'f
-
Bregma-0.3mm
-‘lo
B
ki
e z
2 Deqma-0.8mmJ
L
40 20
0
' ' 20'30'60'50'60'70'B
or
DAY (MARCAIN)
it
+ +
40.0 I:
39.6
100
15
%d
1
VI 2z +I
Rz
+’ + +
6
ox
60
1-
38.4k ‘i?
10 0
- 80 --"
5 :
- 60
,l,j’
- 40 - 20
765432101234567
PIG. 1. Reconstruction of histological sections of rat brain according to the atlas of Paxinos and Watson. The filled areas show extension of 0.4 ~1 marcain injected into the mPOA which was effective in influencing SAW and Tree. Extension of 0.4 pl saline and 0.2 pl marcain was within the same boundary. Ineffective injection sites are shown by hatched circles. Though injections were made bilaterally, it is shown on one side only. The demarcation of mPOA is shown on the opposite side according to the atlas. Abbreviations: ac-anterior commissure; OX-optic chiasma; AHy-anterior hypothalamus; 3V-third ventricle; VDBD-nu vertical limb diagonal band, dors.
until the responses returned to the baseline. Sleep-wakefulness was recorded for 100 min while the Tree for 5-7 h depending on 0.2 or 0.4 ~1 marcain infused. In addition, as a control study, Tree for five rats each from experimental (without injection) and normal (without surgery) groups was monitored for the duration corresponding to the recording period, both during the night and the day. At the end of experiments, rats were sacrificed under deep anaesthesia and the site along with spread of infused chemical (Fig. 1) were histologically identified (1). The data were analyzed statistically (3). Effects of marcain on SAW were analyzed (1). The effect of marcain on the Tree was compared with that of 1) the baseline; 2) the postsaline injection; 3) the Tree without injection; and 4) the Tree after injection of 0.210.4 ~1 marcain. The level of significance was determined by applying Wilcoxon matched-pair signed-rank test for 1) mentioned above and Mann-Whitney test for the rest.
37.&t
m
kid
PRE-INJ PERIOD
-POST-INJECTION PERIOD
10'20'30'*
I
10'20'
50'60' 70'80'91
-0
+
INJECTION PIG. 2. The figure shows the mean rectal temperature (Tree) and the mean percent time spent in wakefulness (AW) by the rats before and after 0.4 pl marcain into the mPOA bilaterally during night (A) and day (8) recording. Marcain precipitatedS during the night, while AW during the day though the Tree showed an increase during both the times and continued even when the S-AW returned to a nonsignificant level as compared to that of preinjection values. **, an increase in Tree; + +, an increase in AW; and --, a decrease in AW as compared to preinjection at the significance level of p
