- Email: [email protected]
Enhancement of ethanol-induced sedation and hypothermia by centrally administered neurotensin, β-endorphin and bombesin
Nr,,r,,~burrr,urol”~~Vol. 20. pp 305 LO 3(W Pergamon Prcns Ltd 19x1. Printed in Great Britain
Mxx-390x/x I ~~130.105-~1511~?.~x~/0
ENHANCEMENT OF ETHANOL-INDUCED SEDATION AND HYPOTHERMIA BY CENTRALLY ADMINISTERED NEUROTENSIN, B-ENDDRPHIN AND BOMBESIN
D. Luttinger, C.B. Nemeroff, G.A. Mason, G.D. Frye, G.R. Breese and A.J.
Prange, Jr.
Biological Sciences Research Center Departments of Psychiatry and Pharmacology and the Neurobiology Program University of north Carolina School of Medicine Chape? Hill, North Carolina 27514 (Accepted d Uecwnb4n 19801 SUMMARY: Intracisternal administration of three endogenous neuropeptides (neurotensin, S-endorphin, or bombesin) potentiated the duration of sedation induced by a fixed dose of ethanol (5.2 g/kg) in mice. The minimally effective dose of each peptide that enhanced ethanol-induced sedation was: neurotensin, 0.18 nmoles; 8-endorphin, 1.79 nmoles; and bombesin, 0.06 nmoles. The enhancement of ethanol-induced sedation was correlated with the potentiation of ethanol-induced hypothermia for all three peptides. None of the neuropeptides studied significantly altered blood or brain ethanol concentrations, suggesting that the observed effects were not due to differences in ethanol metabolism. Several neurotransmitters have been reported to be involved in mediating the central nervous system (CNS) effects of ethanol (EtOH) (Wallgren and Barry, 1970; Kalant, 1975). Neuropharmacological manipulation of the activity of neurotransmitter systems has been particularly useful in determining the relationship between the action of EtOH and specific neurotransmitters. For example, Carlsson and co-workers have suggested that the increase in locomotor behavior in mice observed after low doses of EtOH is mediated by increased activity of catecholaminergic neuronal systems (Carlsson et al., 1974; Strambom et al., 1977). Similarly, y-amino butyric acid containing neurons also appear to be involved in certain actions of EtOH, including EtOH induced motor impairment (HH'kkinen and Kulonen, 1976; Frye et al., 1979) and EtOH withdrawal reactions (Frye et al., 1979; Goldstein, 1973). Although neuropharmacological strategies have provided useful information about the involvement of neurotransmitters in the central actions of EtOH, our understanding of the mechanism of action of EtOH remains incomplete (Liljequist and Engel, 1979). Neuropeptides constitute a relatively new class of potential neurotransmitters. It is well established that neuropeptides administered directly into the CNS can alter behavior and responses to pharmacological agents (Prange et al., 1978). neuroFor example, after CNS administration tensin (NT), an endogenous tridecapeptide which is distributed heterogeneously in the CNS of mammals, has been reported to cause hypothermia, muscle relaxation, diminished locomotor activity, and analgesia (see Nemeroff, 1980 for review). Furthermore, NT markedly potentiates barbiturate-induced sedation (Nemeroff et al., 1977). However, the latter effect may not represent a direct CNS effect since the change was associated with a si.gnificant reduction in the rate of degradation and removal of the barbiturate (Nemeroff et al., 1977). 8-endorphin (8-E), a peptide comprised of 31 amino acids, has been found in the brain and the pituitary of a variety of mammals; CNS administration produces analgesia (Tseng et al., 1976), alterations in thermoregulation (Huidobro-Toro and Way, 1979) and catalepsy (Bloom et al., 1976). Bombesin (BOM), an endogenous tetradecapeptide heterogenously distributed in the mammalian CNS, produces, after CNS administration, hyperglycemia and alterations in thermoregulation (see Brown and Vale, 1979 for review). Since NT, 8-E and BOM have several similar central actions, it was of interest to compare and contrast the effects of central administration of these neuropeptides on EtOH-induced sedation and hypothermia. 305
Preliminary Notes
306 MATERIALS AND METHODS
Adult, male Swiss-Webster mice (26-30 pros)were purchased from Flow Laboratories (Dublin, Virginia) and were group housed (6/cage) in a controlled environment animal facility (12 hrs light, 12 hrs dark) with laboratory chow (Wayne Blox Laboratory Rodent Chow) and water available ad libitum for at least one week. All experiments were conducted during the-phase of the light-dark cycle, at ambient temperatures of 24-26OC. The first series of experiments examined the effects of intracisternally (IC) administered NT, B-E or BOM on the duration of EtOH-induced sedation and the magnitude of EtOH-induced hypothermia. A hypnotic dose of EtOH, 5.2 g/kg (15% w/v), was administered by intraperitoneal (IP) injection. The EtOH caused the mice to lose the righting reflex within three minutes. Ten minutes after EtOH injection (after loss of the righting reflex) mice received an IC injection of NT, B-E, BOM or 0.9% NaCl vehicle in a volume of 10 pl, as previously described (Nemeroff et al., 1977). The range of peptide doses studied was 0.02-59.8 nmoles. Duration of sleep was defined as the time from loss of the righting reflex to its subsequent recovery (i.e., three rightings within one minute) determined by an observer ignorant of the treatment conditions. Colonic temperature was measured, using a lubricated thermistor probe (Yellow Springs Instrument Company, Yellow Springs, Ohio) inserted three cm into the rectum, at four time points, 0, 30 and 60 minutes after EtOH injection and at the time that the righting reflex returned. To determine if the three peptides altered the pharmacokinetics of EtOH, the effects of 5.98 nmoles of each peptide (equivalent to 10 ug NT, 20.7 pg B-E, and 9.7 pg BOM) or vehicle on blood and brain EtOH concentrations were determined. The injection protocol was identical to that described above. Sixty minutes after EtOH injection the mice were decapitated, trunk blood samples (20 pl) collected and brains rapidly removed. Brains were homogenized in 2.0 ml of deionized water. The tubes were capped tightly and centrifuged at 20,000 x g and the supernatants kept on ice. Twenty pl of brain supernatant or trunk blood were diluted in 180 ul of distilled water containing 0.3 mg/ml of tert-bhtanol as an internal standard. Blood and brain EtOH concentrations were determined the same day by the microanalytical gas chromatography method of Manno and Manno (1978). RESULTS The dose of EtOH used induced a mean duration of sleep of approximately 75 (S.E. + 5) minutes. EtOH administration alone reduced colonic temperature by approximately 3.5OC (S.E. 2 0.2). This drop in temperature occurred within 30 minutes of EtOH administration, with no further change in temperature being observed between 30 and 60 minutes. All three neuropeptides significantly potentiated the sedation induced by 5.2 g/kg of EtOH. Both EtOH-induced sedation and hypothermia were augmented by NT and B-E in a dose related manner (Fig. 1). BOM significantly enhanced EtOH-induced sedation but did not alter EtOH-induced hypothermia in a dose related manner (Fig. 1). However, for all three peptides studied there was an inverse, linear correlation between the duration of sedation and colonic temperature (p < 0.001). When the mice regained the righting reflex, NT treated and B-E treated mice had significantly lower colonic temperatures than vehicle-treated mice (data not shown). Mice receiving EtOH and BOM engaged in a stereotypic hindlimb scratching of the head, as has previously been reported when the peptide is administered alone. None of the other treatments resulted in this behavior. None of the mice treated with EtOH in combination with NT died; however treatment with EtOH, together with B-E or BOM, was lethal to several mice. The fact that NT did not produce death in the EtOH-treated mice was in sharp contrast to its effects in pentobarbital-treatedanimals; NT administered IC produced a dose-related mortality in mice treated with a nonlethal dose (50 mg/kg ip) of the barbiturate (NT LDsO = 3.1 nmoles = 5.23 pg) (Nemeroff et al., 1977). Sixty minutes after EtOH administration (i.e., the time when mice receiving EtOH plus vehicle begin to regain the righting reflex) there was no significant difference in blood or brain EtOH concentrations between mice injected with vehicle or one of the three neuropeptides (Table I). Furthermore, no significant differences were found in blood EtOH concentrations of mice that had regained the righting reflex by time of sacrifice and those that had not (data not shown).
307
Preliminary Notes
Figure 1. Effect of IC NT (Panel A), B-E (Panel B) and BOM (Panel C) on the duration of sleep (min) and colonic temperature ("C) sixty minutes after an IP injection of EtOH (5.2 g/kg). Duration of sleep is expressed as % control + SEM. The EtOH and vehicle-treated mice slept for 75 + 5 minutes (n = 96). Experimental values were compared with those of controls by Dunnett's test for multiple comparisons (two-tailed). Each point is the mean of 8 or more mice. represents the effects of peptide on duration of sleep. _ ___ represents the effects of peptide on EtOH induced hypothermia. * * p < 0.01 (Dunnett's test for multiple comparisons). p < 0.05, TABLE I THE EFFECT OF INTBACISTEHNALZYADMINISTERED NEUNOTENSIN, 8-ENDOHPHIN AND BOMBESIN ON BLOOD AND BRAIN ETHANOL CON~ENT~TIONS IN MICE Ethanol Concentrations N -
Blooda
N -
Brainb
EtOH + Vehicle
8
5.62 ? 0.20
7
3.09 + 0.13
EtOH + Neurotensin
8
5.88 f 0.13
8
3.08 + 0.15
EtOH + 8-Endorphin
6
6.20 + 0.24
6
3.02 + 0.18
EtOH + Bombesin
8
5.96 -I0.24
8
2.89 ?:0.12
a mg EtOH/ml b mg EtOH/g tissue Mice were treated with EtOH (5.2 g/kg/IP). Ten minutes later they were injected intracisternallywith vehicle (0.9% NaCl) or peptide (5.98 nmoles). Sixty minutes after EtOH treatment mice were decapitated and brain and blood EtOH concentrations assayed. No significant differences were found when ethanol levels were compared between EtOH-vehicle group and EtOH-peptide groups (Dunnett's test for multiple comparisons).
Preliminary Notes DISCUSSION NT potentiates pentobarbital-inducedsedation and lethality and this effect may be due, at least in part, to a decreased rate of pentobarbital metabolism (Nemeroff et al., 1977). However, the increased sedation in mice receiving EtOH plus NT, B-E or BOM in the present experiments does not appear to be due to alterations in the rate of EtOH elimination. Although IC NT prolongs both pentobarbital (Nemeroff et al., 1977) and EtOHinduced sedation and EtOH-induced hypothermia, the salient difference in the interaction of these CNS depressants and NT was that NT delays the removal of pentobarbital from blood and brain, but does not change EtOH pharmacokinetics. Thus the present findings indicate that NT can enhance responses to sedative drugs other than by an action on drug metabolism. This raises the possibility that mechanisms other than delayed drug removal may also play a role in the NT-pentobarbital interaction. The interactions described between NT and EtOH are shared by the other two peptides tested. This is not astonishing for the three peptides-NT, B-E, and BOM--have several other properties in common, as noted above. The observed effects, however, are not due to non-specific effects of peptides. Several neuropeptides [luteinizing hormone-releasinghormone (LHRH), pro-leu-gly-NH* (MIF-I) and somatostatin] have been studied and found to exert no effect on EtOH-induced sedation (Cott et al., 1976). Moreover, some peptides [thyrotropin-releasinghormone (TRH), several congeners of TRH, and substance P] decreased EtOH-induced sedation (Cott et al., 1976).. It is important to note that the enhancement of EtOHinduced sleep by NT, B-E, and BOM, is probably not due to summation of sedative effects since no dose of NT, B-E or BOM when administered alone produced loss of the righting reflex. Further study is necessary to determine if the increased duration of EtOH-induced sedation is secondary to the potentiation of EtOH-induced hypothermia.
Acknowledgements This work was supported by grants from NIMH (MH 22536, MH 32316, MH 33127), NICHHD (HD-03110) and from the North Carolina Alcoholism Research Authority (7908). We thank the National Institute of Mental Health for providing the human p-endorphin. Bombesin and neurotensin were purchased from Bachem (Torrance, California).
REFERENCES Bloom, F.E., Segal, D., Ling, N. and Guillemin, R. (1976) Endorphins: Profound behavioral effects in rats suggest new etiological factors in mental -illness. Science 194: 630. Brown, M. and Vale, W. (1979) Bombesin-a putative mammalian neurogastrointestinal peptide. Trends in Neuroscience 2: 95. -Carlsson, A., Engel, J., StrGmbom, U., Svensson, T.H. and Waldeck, B. (1974) Suppression by dopamine agonists of the ethanol-induced stimulation of locomotor activity and brain dopamine synthesis. Neuyn-Schmiedeberg'sArch. Pharmacol. 283: 117. Cott, J.M., Breese, G.R., Cooper, B.R., Barlow, T.S. and Prange, A.J., Jr. (1976) Investigations into the mechanism of reduction of ethanol sleep by ihyroiropin-releasinghormone (TRH). 2. Pharmacol. &. Ther. 196: 596.Frye, G., Breese, G., Mailman, R., Vogel, R. and Mueller, R. (1979) Similarities in the central actions of GABA-mimetic drugs and ethanol. Brain Res. Bull. 4: 706. -Goldstein, D.B. (1973) Alcohol withdrawal reactions in mice: Effects of drugs that modify neurotransmission. 2. Pharmacol. m. Ther. 186: 1. Hlkkinen, H.M. and Kulonen, E. (1976) Ethanol intoxication and GABA (yaminobutyric acid). 2. Neurochem. 27: 631.
Preliminary Notes Huidobro-Tore, J.P. and Way, E.L. (1979) sponse of B-endorphin in mice. _ _Pharm. J. Kalant, H. (1975) Proc. 34: 1930. Liljequist, stration
Direct
effects
of
Manno, B.R. microanalysis II. Analytical
and Manno, J.E. (1978) of alcohols in blood Toxicology 2: 257.
Studies on the hyperthermic %. Ther. 211: 50.
ethanol
S. and Engel, J. (1979) on central neurotransmitter
309
on the
The effect mechanisms. A simple and urine
of
nervous
chronic Medical
approach to by a direct
re-
system.
ethanol Biology
Fed.
admini57: 199.
gas chromatographic injection technique.
Nemeroff, C.B., Bissette, G., Prange, A.J., Jr., Loosen, P.T., Barlow, T.S. and Lipton, M.A. (1977) Neurotensin: Central nervous system effects of a hypothalamic peptide. Brain Res. 128: 485. -Nemeroff, C.B. Biol. Psychiat.
(1980) Neurotensin: 15: 283.
Perchance
an endogenous
neuroleptic?
Prange, A.J., Jr., Nemeroff, C.B., Lipton, M.A., Breese, G.R. and Wilson, I.C. (1978) Peptides and the central nervous system. In Handbook of Psychopharmacology, (Iversen, L.L., Iversen, S.D. and Snyder Eds.) Vol. 13, p. 1. StrSmbom, U., Svenson, T.H. and Carlsson, A. (1973) Antagonism of ethanol’s central stimulation in mice by small doses of catecholamine receptor agonists. Psychopharmacol. 51: 293. Tseng, L.F., Loh, H.H. and Li, C.H. (1976) by intravenous injection. Nature 263: 239. Wallgren, (Elsevier,
H. and Barry, New York).
H.,
III
(1970)
g-endorphin
Actions ~---~*
of
as a potent
Alcohol
~01s.
analgesic
1 and 2
Mxx-390x/x I ~~130.105-~1511~?.~x~/0
ENHANCEMENT OF ETHANOL-INDUCED SEDATION AND HYPOTHERMIA BY CENTRALLY ADMINISTERED NEUROTENSIN, B-ENDDRPHIN AND BOMBESIN
D. Luttinger, C.B. Nemeroff, G.A. Mason, G.D. Frye, G.R. Breese and A.J.
Prange, Jr.
Biological Sciences Research Center Departments of Psychiatry and Pharmacology and the Neurobiology Program University of north Carolina School of Medicine Chape? Hill, North Carolina 27514 (Accepted d Uecwnb4n 19801 SUMMARY: Intracisternal administration of three endogenous neuropeptides (neurotensin, S-endorphin, or bombesin) potentiated the duration of sedation induced by a fixed dose of ethanol (5.2 g/kg) in mice. The minimally effective dose of each peptide that enhanced ethanol-induced sedation was: neurotensin, 0.18 nmoles; 8-endorphin, 1.79 nmoles; and bombesin, 0.06 nmoles. The enhancement of ethanol-induced sedation was correlated with the potentiation of ethanol-induced hypothermia for all three peptides. None of the neuropeptides studied significantly altered blood or brain ethanol concentrations, suggesting that the observed effects were not due to differences in ethanol metabolism. Several neurotransmitters have been reported to be involved in mediating the central nervous system (CNS) effects of ethanol (EtOH) (Wallgren and Barry, 1970; Kalant, 1975). Neuropharmacological manipulation of the activity of neurotransmitter systems has been particularly useful in determining the relationship between the action of EtOH and specific neurotransmitters. For example, Carlsson and co-workers have suggested that the increase in locomotor behavior in mice observed after low doses of EtOH is mediated by increased activity of catecholaminergic neuronal systems (Carlsson et al., 1974; Strambom et al., 1977). Similarly, y-amino butyric acid containing neurons also appear to be involved in certain actions of EtOH, including EtOH induced motor impairment (HH'kkinen and Kulonen, 1976; Frye et al., 1979) and EtOH withdrawal reactions (Frye et al., 1979; Goldstein, 1973). Although neuropharmacological strategies have provided useful information about the involvement of neurotransmitters in the central actions of EtOH, our understanding of the mechanism of action of EtOH remains incomplete (Liljequist and Engel, 1979). Neuropeptides constitute a relatively new class of potential neurotransmitters. It is well established that neuropeptides administered directly into the CNS can alter behavior and responses to pharmacological agents (Prange et al., 1978). neuroFor example, after CNS administration tensin (NT), an endogenous tridecapeptide which is distributed heterogeneously in the CNS of mammals, has been reported to cause hypothermia, muscle relaxation, diminished locomotor activity, and analgesia (see Nemeroff, 1980 for review). Furthermore, NT markedly potentiates barbiturate-induced sedation (Nemeroff et al., 1977). However, the latter effect may not represent a direct CNS effect since the change was associated with a si.gnificant reduction in the rate of degradation and removal of the barbiturate (Nemeroff et al., 1977). 8-endorphin (8-E), a peptide comprised of 31 amino acids, has been found in the brain and the pituitary of a variety of mammals; CNS administration produces analgesia (Tseng et al., 1976), alterations in thermoregulation (Huidobro-Toro and Way, 1979) and catalepsy (Bloom et al., 1976). Bombesin (BOM), an endogenous tetradecapeptide heterogenously distributed in the mammalian CNS, produces, after CNS administration, hyperglycemia and alterations in thermoregulation (see Brown and Vale, 1979 for review). Since NT, 8-E and BOM have several similar central actions, it was of interest to compare and contrast the effects of central administration of these neuropeptides on EtOH-induced sedation and hypothermia. 305
Preliminary Notes
306 MATERIALS AND METHODS
Adult, male Swiss-Webster mice (26-30 pros)were purchased from Flow Laboratories (Dublin, Virginia) and were group housed (6/cage) in a controlled environment animal facility (12 hrs light, 12 hrs dark) with laboratory chow (Wayne Blox Laboratory Rodent Chow) and water available ad libitum for at least one week. All experiments were conducted during the-phase of the light-dark cycle, at ambient temperatures of 24-26OC. The first series of experiments examined the effects of intracisternally (IC) administered NT, B-E or BOM on the duration of EtOH-induced sedation and the magnitude of EtOH-induced hypothermia. A hypnotic dose of EtOH, 5.2 g/kg (15% w/v), was administered by intraperitoneal (IP) injection. The EtOH caused the mice to lose the righting reflex within three minutes. Ten minutes after EtOH injection (after loss of the righting reflex) mice received an IC injection of NT, B-E, BOM or 0.9% NaCl vehicle in a volume of 10 pl, as previously described (Nemeroff et al., 1977). The range of peptide doses studied was 0.02-59.8 nmoles. Duration of sleep was defined as the time from loss of the righting reflex to its subsequent recovery (i.e., three rightings within one minute) determined by an observer ignorant of the treatment conditions. Colonic temperature was measured, using a lubricated thermistor probe (Yellow Springs Instrument Company, Yellow Springs, Ohio) inserted three cm into the rectum, at four time points, 0, 30 and 60 minutes after EtOH injection and at the time that the righting reflex returned. To determine if the three peptides altered the pharmacokinetics of EtOH, the effects of 5.98 nmoles of each peptide (equivalent to 10 ug NT, 20.7 pg B-E, and 9.7 pg BOM) or vehicle on blood and brain EtOH concentrations were determined. The injection protocol was identical to that described above. Sixty minutes after EtOH injection the mice were decapitated, trunk blood samples (20 pl) collected and brains rapidly removed. Brains were homogenized in 2.0 ml of deionized water. The tubes were capped tightly and centrifuged at 20,000 x g and the supernatants kept on ice. Twenty pl of brain supernatant or trunk blood were diluted in 180 ul of distilled water containing 0.3 mg/ml of tert-bhtanol as an internal standard. Blood and brain EtOH concentrations were determined the same day by the microanalytical gas chromatography method of Manno and Manno (1978). RESULTS The dose of EtOH used induced a mean duration of sleep of approximately 75 (S.E. + 5) minutes. EtOH administration alone reduced colonic temperature by approximately 3.5OC (S.E. 2 0.2). This drop in temperature occurred within 30 minutes of EtOH administration, with no further change in temperature being observed between 30 and 60 minutes. All three neuropeptides significantly potentiated the sedation induced by 5.2 g/kg of EtOH. Both EtOH-induced sedation and hypothermia were augmented by NT and B-E in a dose related manner (Fig. 1). BOM significantly enhanced EtOH-induced sedation but did not alter EtOH-induced hypothermia in a dose related manner (Fig. 1). However, for all three peptides studied there was an inverse, linear correlation between the duration of sedation and colonic temperature (p < 0.001). When the mice regained the righting reflex, NT treated and B-E treated mice had significantly lower colonic temperatures than vehicle-treated mice (data not shown). Mice receiving EtOH and BOM engaged in a stereotypic hindlimb scratching of the head, as has previously been reported when the peptide is administered alone. None of the other treatments resulted in this behavior. None of the mice treated with EtOH in combination with NT died; however treatment with EtOH, together with B-E or BOM, was lethal to several mice. The fact that NT did not produce death in the EtOH-treated mice was in sharp contrast to its effects in pentobarbital-treatedanimals; NT administered IC produced a dose-related mortality in mice treated with a nonlethal dose (50 mg/kg ip) of the barbiturate (NT LDsO = 3.1 nmoles = 5.23 pg) (Nemeroff et al., 1977). Sixty minutes after EtOH administration (i.e., the time when mice receiving EtOH plus vehicle begin to regain the righting reflex) there was no significant difference in blood or brain EtOH concentrations between mice injected with vehicle or one of the three neuropeptides (Table I). Furthermore, no significant differences were found in blood EtOH concentrations of mice that had regained the righting reflex by time of sacrifice and those that had not (data not shown).
307
Preliminary Notes
Figure 1. Effect of IC NT (Panel A), B-E (Panel B) and BOM (Panel C) on the duration of sleep (min) and colonic temperature ("C) sixty minutes after an IP injection of EtOH (5.2 g/kg). Duration of sleep is expressed as % control + SEM. The EtOH and vehicle-treated mice slept for 75 + 5 minutes (n = 96). Experimental values were compared with those of controls by Dunnett's test for multiple comparisons (two-tailed). Each point is the mean of 8 or more mice. represents the effects of peptide on duration of sleep. _ ___ represents the effects of peptide on EtOH induced hypothermia. * * p < 0.01 (Dunnett's test for multiple comparisons). p < 0.05, TABLE I THE EFFECT OF INTBACISTEHNALZYADMINISTERED NEUNOTENSIN, 8-ENDOHPHIN AND BOMBESIN ON BLOOD AND BRAIN ETHANOL CON~ENT~TIONS IN MICE Ethanol Concentrations N -
Blooda
N -
Brainb
EtOH + Vehicle
8
5.62 ? 0.20
7
3.09 + 0.13
EtOH + Neurotensin
8
5.88 f 0.13
8
3.08 + 0.15
EtOH + 8-Endorphin
6
6.20 + 0.24
6
3.02 + 0.18
EtOH + Bombesin
8
5.96 -I0.24
8
2.89 ?:0.12
a mg EtOH/ml b mg EtOH/g tissue Mice were treated with EtOH (5.2 g/kg/IP). Ten minutes later they were injected intracisternallywith vehicle (0.9% NaCl) or peptide (5.98 nmoles). Sixty minutes after EtOH treatment mice were decapitated and brain and blood EtOH concentrations assayed. No significant differences were found when ethanol levels were compared between EtOH-vehicle group and EtOH-peptide groups (Dunnett's test for multiple comparisons).
Preliminary Notes DISCUSSION NT potentiates pentobarbital-inducedsedation and lethality and this effect may be due, at least in part, to a decreased rate of pentobarbital metabolism (Nemeroff et al., 1977). However, the increased sedation in mice receiving EtOH plus NT, B-E or BOM in the present experiments does not appear to be due to alterations in the rate of EtOH elimination. Although IC NT prolongs both pentobarbital (Nemeroff et al., 1977) and EtOHinduced sedation and EtOH-induced hypothermia, the salient difference in the interaction of these CNS depressants and NT was that NT delays the removal of pentobarbital from blood and brain, but does not change EtOH pharmacokinetics. Thus the present findings indicate that NT can enhance responses to sedative drugs other than by an action on drug metabolism. This raises the possibility that mechanisms other than delayed drug removal may also play a role in the NT-pentobarbital interaction. The interactions described between NT and EtOH are shared by the other two peptides tested. This is not astonishing for the three peptides-NT, B-E, and BOM--have several other properties in common, as noted above. The observed effects, however, are not due to non-specific effects of peptides. Several neuropeptides [luteinizing hormone-releasinghormone (LHRH), pro-leu-gly-NH* (MIF-I) and somatostatin] have been studied and found to exert no effect on EtOH-induced sedation (Cott et al., 1976). Moreover, some peptides [thyrotropin-releasinghormone (TRH), several congeners of TRH, and substance P] decreased EtOH-induced sedation (Cott et al., 1976).. It is important to note that the enhancement of EtOHinduced sleep by NT, B-E, and BOM, is probably not due to summation of sedative effects since no dose of NT, B-E or BOM when administered alone produced loss of the righting reflex. Further study is necessary to determine if the increased duration of EtOH-induced sedation is secondary to the potentiation of EtOH-induced hypothermia.
Acknowledgements This work was supported by grants from NIMH (MH 22536, MH 32316, MH 33127), NICHHD (HD-03110) and from the North Carolina Alcoholism Research Authority (7908). We thank the National Institute of Mental Health for providing the human p-endorphin. Bombesin and neurotensin were purchased from Bachem (Torrance, California).
REFERENCES Bloom, F.E., Segal, D., Ling, N. and Guillemin, R. (1976) Endorphins: Profound behavioral effects in rats suggest new etiological factors in mental -illness. Science 194: 630. Brown, M. and Vale, W. (1979) Bombesin-a putative mammalian neurogastrointestinal peptide. Trends in Neuroscience 2: 95. -Carlsson, A., Engel, J., StrGmbom, U., Svensson, T.H. and Waldeck, B. (1974) Suppression by dopamine agonists of the ethanol-induced stimulation of locomotor activity and brain dopamine synthesis. Neuyn-Schmiedeberg'sArch. Pharmacol. 283: 117. Cott, J.M., Breese, G.R., Cooper, B.R., Barlow, T.S. and Prange, A.J., Jr. (1976) Investigations into the mechanism of reduction of ethanol sleep by ihyroiropin-releasinghormone (TRH). 2. Pharmacol. &. Ther. 196: 596.Frye, G., Breese, G., Mailman, R., Vogel, R. and Mueller, R. (1979) Similarities in the central actions of GABA-mimetic drugs and ethanol. Brain Res. Bull. 4: 706. -Goldstein, D.B. (1973) Alcohol withdrawal reactions in mice: Effects of drugs that modify neurotransmission. 2. Pharmacol. m. Ther. 186: 1. Hlkkinen, H.M. and Kulonen, E. (1976) Ethanol intoxication and GABA (yaminobutyric acid). 2. Neurochem. 27: 631.
Preliminary Notes Huidobro-Tore, J.P. and Way, E.L. (1979) sponse of B-endorphin in mice. _ _Pharm. J. Kalant, H. (1975) Proc. 34: 1930. Liljequist, stration
Direct
effects
of
Manno, B.R. microanalysis II. Analytical
and Manno, J.E. (1978) of alcohols in blood Toxicology 2: 257.
Studies on the hyperthermic %. Ther. 211: 50.
ethanol
S. and Engel, J. (1979) on central neurotransmitter
309
on the
The effect mechanisms. A simple and urine
of
nervous
chronic Medical
approach to by a direct
re-
system.
ethanol Biology
Fed.
admini57: 199.
gas chromatographic injection technique.
Nemeroff, C.B., Bissette, G., Prange, A.J., Jr., Loosen, P.T., Barlow, T.S. and Lipton, M.A. (1977) Neurotensin: Central nervous system effects of a hypothalamic peptide. Brain Res. 128: 485. -Nemeroff, C.B. Biol. Psychiat.
(1980) Neurotensin: 15: 283.
Perchance
an endogenous
neuroleptic?
Prange, A.J., Jr., Nemeroff, C.B., Lipton, M.A., Breese, G.R. and Wilson, I.C. (1978) Peptides and the central nervous system. In Handbook of Psychopharmacology, (Iversen, L.L., Iversen, S.D. and Snyder Eds.) Vol. 13, p. 1. StrSmbom, U., Svenson, T.H. and Carlsson, A. (1973) Antagonism of ethanol’s central stimulation in mice by small doses of catecholamine receptor agonists. Psychopharmacol. 51: 293. Tseng, L.F., Loh, H.H. and Li, C.H. (1976) by intravenous injection. Nature 263: 239. Wallgren, (Elsevier,
H. and Barry, New York).
H.,
III
(1970)
g-endorphin
Actions ~---~*
of
as a potent
Alcohol
~01s.
analgesic
1 and 2