Cholecystokinin and morphine-induced hypothermia

European Neuropsychopharmacology 9 (1999) 219–225

Cholecystokinin and morphine-induced hypothermia Mehdi Rezayat, Neda Ravandeh, Mohammad-Reza Zarrindast* Department of Pharmacology, School of Medicine, Tehran University of Medical Sciences, P.O. Box 13145 -784, Tehran, Iran Received 21 November 1997; accepted 23 June 1998

Abstract The effects of cholecystokinin-8 sulfate (CCK-8), cholecystokinin-8 unsulfate (CCK-8U), cholecystokinin-4 (CCK-4), caerulein and morphine on mice core body temperature have been studied in the present work. Subcutaneous injection of different doses of caerulein (0.05, 0.1 and 0.5 mg / kg), CCK-8 (0.05, 0.1 and 0.25 mg / kg) and morphine (10, 20 and 30 mg / kg) induced hypothermia. CCK-8U and CCK-4 did not elicit any response. The hypothermic response induced by caerulein, a CCK-related decapeptide but not morphine was decreased by selective CCKA receptor antagonist MK-329. However, the hypothermia induced by morphine but not caerulein was reduced by opioid antagonist naloxone. When morphine plus caerulein was administered a higher hypothermia was induced. Pretreatment of animals with L-365 260, a selective CCK B receptor antagonist did not alter the hypothermia induced by the drugs. The response induced by combination of the both drugs was decreased by MK-329. Administration of CCK antagonists MK-329 and L-365 260 to mice did not exert any effect on temperature. It is concluded that the CCKA receptor mechanism may be involved in the hypothermic effect of CCK agonists or morphine, while opioid receptor mechanism is not involved in CCK receptor agonists’ response.  1999 Elsevier Science B.V. All rights reserved. Keywords: CCK agents; Morphine; Hypothermia; Mice

1. Introduction Cholecystokinin (CCK) is a major intestinal, 33 amino acid peptide, with an important role in regulating the control of pancreatic secretion and bile ejection. The peptide is also found in the central nervous system (Vanderhaeghen et al., 1975; Rehfeld, 1978; Woodruff and Hughes, 1991). CCK receptors are generally divided into two classes, cholecystokinin-A (CCKA ; alimentary) and cholecystokinin-B (CCK B ; brain) (Moran et al., 1986; Dourish and Hill, 1987). The CCKA and CCK B receptors are distributed within the central nervous system, although the distribution of CCKA receptors are limited to regions of the mid and hind brain in rodents (Hill et al., 1987b; Baber et al., 1989). CCK-octapeptide (CCK-8) and the related decapeptide, caerulein, exert a wide variety of pharmaco-

*Corresponding author.

logical effects such as antinociception, catalepsy, sedation, ptosis, anticonvulsant and depression (Zetler, 1980a,b,c, 1981a,b,c). CCK receptor antagonists potentiate morphine antinociception (Watkins et al., 1984, 1985 Katsuura and Itoh, 1985; Dourish et al., 1988, 1990) and elicit a suppressive effect on b-endorphin-induced catalepsy in rats (Itoh and Katsuura, 1981). Our previous studies showed that CCK receptor activation by CCK peptides may prevent tolerance to morphine antinociception (Rezayat et al., 1994) and may have a role in morphine dependence (Zarrindast et al., 1995). CCK agonists have been shown to produce hypothermia (Morley et al., 1981; Zetler, 1982a; Kapas et al., 1987). There is a report showing that injection of CCK-8 into the cerebral ventricle of rats induces a fever-like thermoregulatory response (Szelenyi et al., 1994). Morphine affects body temperature in a variety of animal species; it elicits a biphasic effect in rats (Lotti et

0924-977X / 99 / $ – see front matter  1999 Elsevier Science B.V. All rights reserved. PII: S0924-977X( 98 )00029-7

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M. Rezayat et al. / European Neuropsychopharmacology 9 (1999) 219 – 225

al., 1965; Oka et al., 1971; Cox et al., 1976; Kaakkola and Ahtee, 1977) and also causes hyperthermia and hypothermia in mice (Glick, 1975; Zarrindast et al., 1994). In the present study, mechanism(s) of CCK receptor activation and morphine-induced hypothermia and their interactions have been investigated.

3. Results

2. Materials and methods

3.1. Effect of morphine and CCK agonists on mice core body temperature ( Table 1)

2.1. Animals Male albino mice (weight range 20–30 g) were used in all experiments. They remained in groups of ten in their cage under conditional temperature (24628C) with free access to food and water. Animals were deprived of food for 18 h before experiments but water was available all the time. On the day of experiment, mice were housed individually in experimental cages and allow to rest for 1 h before drug injection. The core body temperature was measured with a rectal thermistor probe (Light Labs, UK) inserted to a depth of 2 cm for period of 2 h. The data are shown as the change in body temperature from basal values. Basal values are those taken immediately before the CCK agonist administration (time 0) after saline or antagonist injection. When saline, vehicle, morphine, CCK receptor agonists or antagonists were used alone, basal values are those taken immediately before the administration of saline, vehicle or each drug (time 0). The experimental protocol was approved by the Research and Ethics Committee of the School of Pharmacy, Tehran University of Medical Sciences (No. P-50 / 94).

2.2. Statistical analysis Comparison between groups were made with analysis of variance (ANOVA) following Newman–Keul’s test. A difference with P,0.05 between experimental groups was considered statistically significant.

2.3. Drugs The following drugs were used: sulfated cholecystokinin-8 (CCK-8), unsulfated cholecystokinin-8 (CCK-8U) (Sigma, USA), caeruletide (caerulein diethylammonium hydrate), and cholecystokinin-4 (caeruletide tetrapeptide; CCK-4; Farmitalia, Italy), morphine sulphate (MacFarlan Smith, UK) and naloxone HCl (Sigma). MK329 (1-methyl-3-(2 indoloyl)amino-5-phenyl-3H-1,4-benzodiazepin-2-one) and L-365 260 [3R(1)-N-(2,3-dihydro1-methyl- 2 -oxo- 5 -phenyl- 1H -1, 4-benzodiazepin-3-yl)-N(3-methylphenyl)urea] (Merck Sharp and Dohme, UK). All the drugs were dissolved in saline except MK-329 and L-365 260 which were dissolved in dimethylsulfoxide and water (40 and 60% respectively). The doses of peptides and antagonists used were based on published studies

using these drugs (Zetler, 1982a, 1985; Faris et al., 1983; Hill et al., 1987a; O’Neill et al., 1989; Dourish et al., 1990; Rezayat et al., 1994; Zarrindast et al., 1994).

Repeated measure two-way ANOVA with time (Factor A) as covariant indicates differences between animals which were injected subcutaneously (s.c.) with different doses of drugs (Factor B). Different doses of caerulein (0.05, 0.1 and 0.5 mg / kg) [Factor time: F(5,48)51.74, P.0.05; Factor dose: F(3,144)5119.1, P,0.0001 and Factor time3dose: F(15,144)51.44, P.0.05] and CCK-8 (0.05, 0.1 and 0.25 mg / kg) [Factor time: F(5,48)51.7, P.0.05; Factor dose: F(5,144)535.7, P,0.0001 and Factor time3dose: F(15,144)50.83, P.0.05] decreased body temperature as compared with saline control. The time was not implicated in the drugs’ response and no time3dose interaction was found. Maximal hypothermic effect of caerulein was induced by 0.1 mg / kg, 60 min and maximal response of CCK-8 was induced by 0.25 mg / kg of the drug, 30 min after the drug injection. However, there was a significant difference between different doses (0.1, 0.25 and 0.5 mg / kg) of CCK-4 on body temperature [Factor time: F(5,48)57.0, P,0.0001; Factor dose: F(5,144)59.6, P,0.0001 and Factor time3 dose: F(15,144)50.88, P.0.05] and time was implicated in drug effect, there was no interaction for dose and time. Further analysis showed no response for the drug as compared with the saline control group. CCK-8U (0.05, 0.1 and 0.25 mg / kg) [Factor time: F(5,48)52.2, P.0.05; Factor dose: F(5,144)529.8, P, 0.0001 and Factor time3dose: F(15,144)50.84, P.0.05] did not change body temperature of animals as compared with the saline control group. There was a significant difference between different doses of morphine but no interaction was shown for dose3time (10, 20 and 30 mg / kg) [Factor time: F(5,48)5 2.2, P.0.05; Factor dose: F(3,144)587.2, P,0.0001 and Factor A3B: F(15,144)50.84, P.0.05]. Further analysis indicated that morphine decreased the animal’s body temperature as compared with the saline control group. The maximum response was obtained by 30 mg / kg of the drug.

3.2. Effects of CCK antagonists on morphine- and caerulein-induced hypothermia ( Table 2) Two-way ANOVA indicates a significant difference between animals which were treated with caerulein and those which were administered with caerulein plus different doses of the CCKA receptor antagonist MK-329 (0.25

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221

Table 1 Effects of morphine and CCK analogues on mice body temperature T initial

36.860.5 39.060.1 38.360.2 38.560.2 36.160.6 37.460.6 38.360.2 39.460.2 38.860.1 38.060.3 35.960.9 38.360.2 38.760.2 38.960.1 38.460.2 38.160.2

Treatment

Change in body temperature (8C) at different times after drug injection (min)

(mg / kg)

15

30

45

60

90

120

Saline 10 ml / kg Caerulein 0.05 Caerulein 0.1 Caerulein 0.5 CCK-8 0.05 CCK-8 0.1 CCK-8 0.25 CCK-4 0.1 CCK-4 0.25 CCK-4 0.5 CCK-8U 0.05 CCK-8U 0.1 CCK-8U 0.25 Morphine 10 Morphine 20 Morphine 30

1.060.3 22.460.4 c 23.060.3 c 22.960.3 c 0.760.4 21.360.5 22.260.7 b 20.560.2 0.060.0 0.560.1 1.160.5 20.260.3 0.160.2 21.260.2 a 21.460.3 b 22.260.6 c

1.060.4 22.660.3 c 23.160.3 c 23.160.7 c 1.560.7 21.160.6 25.260.2 c 20.260.6 0.160.1 0.560.2 2.561.0 20.160.3 0.460.2 21.360.4 a 21.660.4 b 22.560.4 c

1.060.5 22.460.5 c 23.160.7 c 22.760.5 c 2.660.5 20.460.6 24.960.4 c 0.660.4 0.260.1 0.860.2 2.861.1 20.360.3 0.360.2 21.360.4 a 21.760.5 b 21.960.3 b

1.260.5 21.660.2 b 23.660.6 c 22.660.6 c 2.460.6 0.361.0 23.660.5 c 0.460.3 0.060.0 0.460.3 2.960.9 21.060.5 a 0.260.2 21.260.6 a 21.560.4 b 21.860.6 c

1.160.6 21.260.3 a 23.260.5 c 21.860.5 b 1.660.6 0.160.3 22.460.4 b 20.760.1 20.260.2 20.160.5 2.960.7 21.860.5 0.060.0 20.860.4 21.360.5 a 21.160.4 a

20.160.7 21.060.2 22.260.6 a 1.260.5 1.160.6 21.060.8 21.960.3 21.560.5 20.360.2 20.860.5 1.260.9 21.160.5 20.460.4 20.660.4 21.060.3 20.360.2

Animals were injected (s.c.) with different doses of morphine or CCK agonists and temperature was recorded 15, 30, 45, 60, 90 and 120 min after drugs injection. Each point is the mean6SEM of nine mice. a P,0.05, b P,0.01, c P,0.001 different from saline control group.

and 0.5 mg / kg) [Factor time: F(3,32)57.55, P,0.0001; Factor dose: F(3,96)512.3, P,0.0001; Factor time3dose: F(9,96)50.69, P.0.05] or CCK B receptor antagonist L365 260 (0.25 and 0.5 mg / kg) [Factor time: F(3,32)5 12.2, P,0.0001; Factor dose: F(3,96)58.3, P,0.0001; Factor time3dose: F(9,96)50.97, P.0.05]. However, ANOVA showed that time was implicated in the drugs’ response, there was no dose3time interaction. Further analysis showed that when MK-329 was administered 5 min prior to caerulein, there was a reduction in caeruleininduced hypothermia. The CCK B receptor antagonist L365 260 (0.125, 0.25 and 0.5 mg / kg) did not affect caerulein response.

ANOVA also showed that there was no difference between hypothermia induced by morphine (30 mg / kg) alone and those of morphine plus CCK receptor antagonists MK-329 or L-365 260 [Factor time: F(3,32)534.2, P,0.0001; Factor dose: F(4,128)52.2, P.0.05; Factor time3dose: F(12,128)52.8, P.0.05].

3.3. Effects of caerulein plus morphine in the presence or absence of CCK antagonist The effects of caerulein plus morphine in the presence or absence of CCK antagonist are shown in Fig. 1. Twoway ANOVA indicates a significant difference between the

Table 2 Effects of CCK antagonists on hypothermia induced by morphine or caerulein (CLN) T initial

Treatment

T at time of CLN or Mor injection

(mg / kg) 38.160.3 37.460.5 38.460.2 38.760.3 38.060.2 38.360.1 37.360.4 38.160.3 38.860.2 37.460.7 38.060.4 38.560.5

Vehicle MK 0.125 MK 0.25 MK 0.5 L 0.125 L 0.25 L 0.5 Vehicle MK 0.25 MK 0.5 L 0.25 L 0.5

1CLN 1CLN 1CLN 1CLN 1CLN 1CLN 1CLN 1Mor 1Mor 1Mor 1Mor 1Mor

37.960.25 37.760.4 38.560.2 38.960.3 37.860.2 38.360.1 37.660.2 38.060.4 38.660.2 38.060.5 38.160.4 38.360.4

Change in body temperature (8C) at different times after CLN or MOR injection (min) 15

30

45

60

22.360.2 21.760.4 20.860.3 b 20.860.4 b 22.860.2 22.360.2 21.760.3 23.560.3 24.060.2 24.560.6 22.360.4 23.160.5

22.660.3 21.260.4 a 21.160.3 a 21.460.5 a 22.760.3 22.360.3 21.360.3 26.060.5 24.560.3 24.960.6 24.360.5 24.660.6

22.160.3 21.060.3 20.660.3 a 20.760.3 a 22.060.3 21.960.3 21.060.3 26.060.4 24.360.3 24.960.6 25.860.6 25.760.7

21.660.3 20.960.3 0.160.3 b 20.260.2 a 21.460.4 21.960.3 20.760.2 25.860.4 24.160.3 24.960.8 25.960.7 25.860.6

Animals were treated (s.c.) either with caerulein (CLN; 0.1 mg / kg) or morphine (Mor; 30 mg / kg) in presence or absence of vehicle (10 ml / kg) or CCK antagonists. CCK receptor antagonists MK-329 (MK; 0.25 and 0.5 mg / kg) or L-365 260 ( L; 0.25 and 0.5 mg / kg) were administered 5 min before CLN and 35 min prior to morphine administration. Temperature was recorded 15–60 min after CLN or morphine injection. Each point is the mean6SEM of nine animals. a P,0.05, b P,0.01 different from control groups.

M. Rezayat et al. / European Neuropsychopharmacology 9 (1999) 219 – 225

222

3.4. Effects of naloxone on hypothermia induced by CCK agonists or morphine ( Table 3) Two-way ANOVA indicates a significant difference between hypothermia induced by morphine (recorded 15– 60 min after morphine administration; Factor time) in presence or absence of naloxone (Factor B) [Factor time: F(3,32)51.38, P.0.5; Factor dose: F(2,32)576.9, P, 0.0001; Factor A3B: F(6,64)50.64, P.0.05]. Further analysis showed that naloxone (1.5 and 3 mg / kg) decreased the hypothermia induced by morphine (30 mg / kg). The opioid antagonist did not alter hypothermia induced by caerulein (0.1 mg / kg) [Factor time: F(3,32)56.1, P,0.01; Factor dose: F(2,32)50.27, P.0.05; Factor time3dose: F(6,64)50.73, P.0.05]. Our previous work (Zarrindast et al., 1994) showed that naloxone alone has no effect on mice body temperature.

Fig. 1. Mice were treated subcutaneously with morphine (d; 30 mg / kg), caerulein (j; 0.1 mg / kg), morphine plus caerulein (.; caerulein 30 min before morphine) or MK-329 plus morphine plus caerulein (,). MK-329 (0.25 mg / kg) was adminstered 5 min before caerulein adminstration. Each point is the mean6SEM of nine animals. ** P,0.01, *** P,0.001 different from caerulein plus morphine treated animals.

hypothermia induced by caerulein (0.1 mg / kg), morphine (30 mg / kg), caerulein plus morphine and caerulein plus morphine in the presence of CCKA receptor antagonist MK-329 (0.25 mg / kg), recorded 15–60 min after drug injection [Factor time: F(4,40)51.9, P.0.05; Factor dose: F(3,120)566.4, P,0.0001; Factor time3dose: F(12,120)50.5, P.0.05]. Further analysis showed that when caerulein administered 30 min prior to morphine induced a higher hypothermia as compared with either drugs, and MK-329 when injected 5 min before caerulein reduced hypothermia induced by morphine plus caerulein. The CCK B receptor antagonist L-365 260 (0.125, 0.25 and 0.5 mg / kg) did not alter the hypothermic response induced by caerulein plus morphine (data not shown).

3.5. Effects of CCK antagonists on mice body temperature ( Table 4) However, there was a significant difference between animals which received single administration of different doses of CCKA receptor antagonist MK-329 (0.125, 0.25 and 0.5 mg / kg) [Factor time: F(3,32)50.19, P.0.05; Factor dose: F(3,96)510.1, P,0.0001; Factor time3dose: F(9,96)50.77, P.0.05] or L-365 260 (0.125, 0.25 and 0.5 mg / kg) [Factor time: F(3,32)50.07, P.0.05; Factor dose: F(3,96)59.4, P,0.0001; Factor time3dose: F(9,96)5 0.48, P.0.05], further analysis indicated that the drugs did not alter the animals’ body temperature measured at different times.

4. Discussion In the present study, administration of CCK-8 and caerulein which are effective on CCK receptors (Bock, 1991; Woodruff and Hughes, 1991; Holladay and Lin, 1992; Slaninova et al., 1991), induced hypothermia in

Table 3 Effects of naloxone on hypothermia induced by caerulein (CLN) or morphine T initial

Treatment

T at time of CLN or Mor injection

(mg / kg)

37.660.5 38.960.2 38.560.2 37.460.9 37.660.4 38.660.2

Saline Nal 1.5 Nal 3 Saline Nal 1.5 Nal 3

Change in body temperature (8C) at different times after CLN or MOR injection (min)

1CLN 1CLN 1CLN 1Mor 1Mor 1Mor

37.760.5 39.060.2 38.360.2 37.561.0 37.860.4 38.660.2

15

30

45

60

23.160.2 23.860.4 23.560.3 23.260.5 20.660.1 c 20.360.2 c

23.760.2 24.060.3 24.260.5 23.660.6 21.360.3 b 20.360.2 c

23.460.2 23.260.3 23.360.4 23.960.6 21.560.4 c 20.460.2 c

22.960.3 23.060.4 22.860.4 23.160.6 21.660.3 b 20.460.2 c

Animals were treated (s.c.) with CLN (0.1 mg / kg) or morphine (Mor; 30 mg / kg) either with saline or naloxone. Saline (10 ml / kg) or naloxone (Nal; 1.5 and 3 mg / kg) were administered intraperitoneally 5 min before CLN or morphine. Each point is the mean6SEM of nine mice. b P,0.01, c P,0.001 different from control group.

M. Rezayat et al. / European Neuropsychopharmacology 9 (1999) 219 – 225

223

Table 4 Effects of CCK antagonists on mice body temperature T initial

38.260.2 37.660.5 38.660.1 37.560.4 38.860.2 37.960.3 38.860.2

Treatment

Change in body temperature (8C) at different times after drug injection (min)

(mg / kg)

15

30

45

60

Vehicle 10 ml MK 0.125 MK 0.25 MK 0.5 L 0.125 L 0.25 L 0.5

0.360.2 20.460.4 20.660.2 0.160.2 20.660.3 20.560.2 20.160.1

0.060.0 20.860.6 20.960.3 0.660.4 21.160.5 20.760.4 20.360.2

0.060.0 21.060.7 21.060.3 0.360.2 21.460.6 20.760.3 21.360.5

20.160.2 20.960.5 21.060.4 0.760.3 21.260.6 21.260.3 21.360.3

Mice were administered (s.c.) vehicle, CCK receptor antagonist, MK-329 (MK) or CCK B receptor antagonist L-365 260 ( L). Temperature was recorded 15–60 after the drugs injections. Each point is the mean6SEM of nine animals.

mice. The results are consistent with other studies in this respect (Morley et al., 1981; Zetler, 1982a; Kapas et al., 1987). Our results show that caerulein is more potent than CCK-8. Others have also found a different pharmacological profile between these two substances (Jurna and Zetler, 1981; Van Ree et al., 1983; Hill et al., 1987a). CCK-8 and caerulein both have high affinity for CCKA and CCK B receptors (Dourish and Hill, 1987; Bock, 1991; Slaninova et al., 1991; Woodruff and Hughes, 1991; Holladay and Lin, 1992). Therefore it could be concluded that both CCK receptors are involved in the hypothermic response of the peptides. However, the present data showed that the CCK B receptor agonists; unsulfated CCK8 and CCK-4 (Dourish and Hill, 1987) did not alter the animals’ body temperature, and therefore, the hypothermic response of CCK and caerulein cannot be related to CCK B receptors. Other results even indicate that CCK B receptor activation may induce hyperthermia (Szelenyi et al., 1994). Measurement of the CCK-8 contents show a heterogenous regional distribution of CCK-8 within the brain with the highest levels being found in the amygdala, cerebral cortex, olfactory lobes and hippocampus (Crawley, 1985; Dockray, 1980; Dockray et al., 1985; Larssen and Rehfeld, 1979; Rehfeld and Hansen, 1984). Intracerebral and systemic administration of CCK-related peptides induced behavioural changes (Jurna and Zetler, 1980, 1981; Itoh and Katsuura, 1981) indicating a central effect of the CCK peptides (Zetler, 1980a, 1982b). It has been shown that intraventricular (Morley et al., 1981) and intrahypothalamic (Shian and Lin, 1985) injection of CCK-8 induces hypothermia in rats, which indicates that the response induced is due to a central action of the agent. Other reports also suggest that the peptides pass the blood brain barrier (Jurna and Zetler, 1981). To evaluate the receptor subtype involved in the hypothermia obtained in the present work, CCKA and CCK B receptor antagonists were challenged against caerulein. The CCKA receptor antagonist MK-329 (devazepide; L364 718) but not the CCK B receptor L-365 260 (Woodruff and Hughes, 1991) antagonised the hypothermia induced by caerulein, therefore involvement of the CCKA receptor

sites is more likely. This hypothesis is supported by other investigators (Szelenyi et al., 1994) who showed that CCK type A receptors may be involved in the hypothermia induced by CCK agents. Although, our results suggest that the CCK receptor mechanism is implicated in thermoregulation, single administration of neither the CCKA receptor antagonist MK-329 nor the CCK B receptor antagonist L-365 260 elicited any response. Accumulating evidence supports the hypothesis that there is a physiological antagonism between CCK and opioids (Faris et al., 1983; Faris, 1985). CCK has been suggested to have opioid and antiopioid effects. The opioid modulating properties of CCK have been shown to be elicited through the activation of CCK receptors; opioidlike and antiopioid effects of CCK to be mediated through CCKA or CCK B receptors respectively [for review see Cessellin (1995)]. Also, the finding that levels of CCK in the brain decrease in response to systemically administered morphine (Lamers et al., 1980) indicates a functional interaction between CCK and opioid systems. Morphine has also been shown to alter the body temperature of animals (Lotti et al., 1965; Oka et al., 1971; Cox et al., 1976; Kaakkola and Ahtee, 1977; Glick, 1975; Zarrindast et al., 1994). Therefore, the interaction of CCK and morphine in hypothermia induced by these drugs was studied. The present data are consistent with our previous study (Zarrindast et al., 1994) in which morphine induced hypothermia. The opioid receptor antagonist naloxone only attenuated the morphine-induced hypothermic effect and not that of caerulein, indicating that at least the opioid receptor mechanism is not involved in the hypothermia induced by the CCK agonist. However, contrary to our results, Zetler (1982a) found that naloxone (0.5 and 2 mg / kg) attenuated the hypothermic effect of caerulein. Combination of morphine with caerulein induced a higher hypothermic response. The CCKA receptor antagonist MK-329 but not the CCK B receptor antagonist L365 260 decreased hypothermia induced by the combination of both drugs. Since opioid-like effects of CCK seem to result from the stimulation of CCKA receptor sites (Cessellin, 1995), therefore it seems that the potentiation

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M. Rezayat et al. / European Neuropsychopharmacology 9 (1999) 219 – 225

response induced by coadministration of both drugs, is at least partly via CCKA receptors. A functional relationship between CCK and dopamine in different regions of brain has been demonstrated. There appears to be a complex interaction between dopamine and the CCK system in the nucleus accumbens (Vaccarino and Rankin, 1989). CCK-8 is colocalized with dopamine in a subpopulation of A10 dopamine-containing neurones ¨ (Hokfelt et al., 1980). Moreover, the CCK peptides may be released with dopamine. It is also possible that CCK-8 influences the release of dopamine or CCK-8 itself by a presynaptic action. The involvement of dopaminergic mechanism(s) in thermoregulation has been shown previously (Zarrindast and Tabatabai, 1992). The dopaminergic system also has been shown to mediate morphine hypothermia (Zarrindast and Tabatabai, 1992), therefore, the involvement of dopaminergic mechanism(s) in the CCKinduced hypothermia and / or potentiated response induced by both the drugs seems possible. To clarify the exact mechanism(s) involved, more studies are required.

Acknowledgements The authors would like to thank Merck Sharp and Dohme, Farmitalia for gifts of drugs and Dr. Mahmoud Ghazi Khansari for assistance in the preparation of the illustrations and computer programs used for this study.

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