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Effect of peptides on the development of tolerance to buprenorphine, a mixed opiate agonist-antagonist analgesic
European Journal of Pharmacology, 79 (1982) 117-123 Elsevier Biomedical Press
117
EFFECT OF PEPTIDES ON THE DEVELOPMENT OF TOLERANCE TO BUPRENORPHINE, A MIXED OPIATE AGONIST-ANTAGONIST ANALGESIC HEMENDRA N. BHARGAVA
Department of Pharmacognosy and Pharmacology, College of PharmaQ,, University of Illinois at the Medical Center, Chicago, Illinois 60612, U.S.A. Received 23 December 1981, accepted ! February 1982
H.N. BHARGAVA, Effect of peptides on the development of tolerance to buprenorphine, a mixed opiate agonist-antagonist analgesi~, European J. Pharmacol. 79 (1982) I 17-123. The subcutaneous administration of buprenorphine to male Sprague-Dawley rats produced a dose-dependent analgesia and hyperthermia in a dose range of 0.25-2 mg/kg. The subcutaneous administration of buprenorphine (0.5 mg/kg) twice a day for 4 days resulted in the development of tolerance to its analgesic and hyperthermic actions. Daily administration of melanotropin release inhibiting factor (Pro-Leu-Gly-NH2) or cyclo (Leu-Gly) (2 mg/kg s.c.) for 4 days, inhibited the development of tolerance to buprenorphine, as evidenced by greater analgesic and hyperthermic responses to buprenorphine in peptide-treated than in vehicletreated buprenorphine-tolerant rats. The repeated injections of peptides did not alter the analgesic or the hyperthermic response to buprenorphine in non-tolerant rats. These studies suggest the possible role of hypothalamic peptides in blocking the development of tolerance to the pharmacological effects of buprenorphine, a potent analgesic agent. Mixed agonist-antagonist
Buprenorphine
MIF
Pro-Leu-Gly-NH 2
Tolerance
Analgesia
Hyperthermia
Cyclo(Leu-Gly)
1. Introduction
Buprenorphine is an oripavine derivative possessing both opiate agonist and antagonist activity. Animal studies indicate that it is a potent analgesic agent having 25-40 times greater potency than morphine after parenteral administration and 7-10 times greater potency after oral administration; it has rapid onset and long duration of action (Cowan et al., 1977a). Furthermore, buprenorphine possesses low physical dependence liability (Cowan, 1974; Cowan et al., 1977b; Jacob et al., 1979). The studies with buprenorphine in humans indicate that the subjective effects experienced by heroin addicts were similar to those of morphine (Jasinski et al., 1978). In addition to its morphine like properties, buprenorphine also has a long duration of action as an opiate antagonist and antagonizes the actions of high doses of morphine for longer than 24h (Jasinski et al., 1978). The same authors (Jasinski et al., 1978) also reported 0014-2999/82/0000-0000/$02.75 © 1982 Elsevier Biomedical Press
that patients maintained on high doses of buprenorphine experience an extremely mild withdrawal syndrome two weeks after the abrupt termination of buprenorphine. A more recent study indicates that buprenorphine suppressed the self administration of heroin in heroin-dependent men; termination of buprenorphine did not result in opiate withdrawal signs and symptoms, and the dependent subjects preferred it over methadone or naltrexone (Mello and Mendelson, 1980). The multiple administration of buprenorphine results in the development of tolerance to its analgesic action as determined by the phenylquinone-writhing test (Cowan et al., 1977b). Since the major uses of buprenorphine is as a potent analgesic agent and as a pharmaco-therapeutic agent for the treatment of heroin dependence, the development of tolerance to its analgesic action is an undesirable effect. Previous studies from these laboratories have indicated that certain hypothalamic peptide hormones, like melanotropin release inhibiting factor, a tripeptide, Pro-Leu-Gly-
118
2. Materials and methods
temperature of each rat was measured prior to and at 30, 60, 90, 120 and 180 min after the drug injection by using a rectal probe and a telethermometer (Yellow Spring Instrument Co., Yellow Springs, Ohio). The animals were not restrained. They were lightly hand handled. The change in rectal temperature for each rat at each time interval was calculated as the difference between predrug and post-drug temperatures. The data are expressed as mean increases in rectal temperature, °C -+S.E.M. Six rats were used for each dose of buprenorphine.
2.1. Animals and chemicals
2.3. Effect of peptides on tolerance to buprenorphine
Male Sprague-Dawley rats weighing 150-200 g obtained from King Animal Laboratories, Oregon, Wis., were housed 3 to a cage in a room with controlled temperature (23 -+ 1°C), humidity (65 -+ 2%) and a 12 h dark-light cycle (L 06 : 00-18 : 00 h). Rats were acclimated to the above conditions for at least four days before being used and were given food and tap water ad libitum. MIF was received as a gift from the Abbott Laboratories, N. Chicago, Illinois through the courtesy of Dr. A.O. Geiszler. Cyclo (Leu-Gly) was synthesized in these laboratories as described previously (Fischer, 1906). Buprenorphine was kindly donated by Dr. A. Cowan of Temple University, Philadelphia, Pennsylvania. The peptides and buprenorphine were dissolved in distilled deionized water and administered subcutaneously (s.c.) in volumes such that each rat received 1 m l / k g of the drug solution.
The effect of MIF and cyclo (Leu-Gly) on the development of tolerance to buprenorphine were studied. Tolerance to buprenorphine was induced by sub'cutaneous injections of buprenorphine (0.5 mg/kg) twice daily at 9 a.m. and 4.30 p.m. for 4 days. The rats were injected with the appropriate peptide (2 m g / k g s.c.) or its vehicle (water) l h before the first injection of buprenorphine. The above procedure (i.e., single injection of peptide or its vehicle and two injections of buprenorphine and its vehicle) was repeated for 4 days. On day 5, the analgesic and thermic responses to buprenorphine (0.5 m g / k g s.c.) in rats from all six treatment groups were assessed as described above. Eight rats were used in buprenorphine groups and 6 rats were used in its vehicle groups.
N H 2 (MIF) and its analog cyclo (Leu-Gly) block the tolerance to morphine in mice and rats (Bhargava et al., 1980; Bhargava, 1980, 1981a,b,e) and to human fl-endorphin in the rat (Bhargava, 198 lc,d). It was, therefore, of interest to determine whether these peptides can also block the tolerance to an agonist-antagonist type analgesic, buprenorphine. The present paper describes such studies in rats.
2.2. Measurement of buprenorphine-induced gesia and changes in body temperature
anal
The analgesic response (% analgesia) to various doses (0.25-2 mg/kg) of buprenorphine was determined using a tail-flick procedure described previously (Bhargava, 1981a). The analgesic response was determined at 30, 60, 90, 120 and 180 min after buprenorphine injection. The data are expressed as mean % analgesic response -+S.E.M. Six rats were used for each dose of buprenorphine. The rats were injected with various doses of buprenorphine (0.25-2.0 m g / k g s.c.) and the rectal
2.4. Statistics The differences in the means of various treatment groups were analyzed by the analysis of variance followed by Schaffe's S-test (Kirk, 1968). The difference was considered statistically significant when the P value was less than 0.05.
3. Results
3.1. Effect of buprenorphine on analgesia and body temperature The administration of buprenorphine produced a dose-related analgesia in the rat. As shown in
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TABLE 1 D o s e - r e s p o n s e relationship for the analgesic effect of b u p r e n o r p h i n e in the rat. D o s e of
P e r c e n t analgesia, m e a n + S.E.M. ( n - 6 )
buprenorphine
T i m e after b u p r e n o r p h i n e injection ( m i n )
m g / k g s.c. 30
60
0.25
46.7 + 17.7
0.50
94.3+
1.00 2.00
100 100
+ +
9(/
41.8 + 13.5
120
45.2 + 17.7
11.8 +
180 4.1
4.0 +
1.5
3.8 a
100
-- 0.0 a
I00
+
0.0 a
4 3 . 0 ~ 1 4 . 7 ~'
46.8+17.6 a
0.0 a 0.0 a
100 100
± +
I(X) + 100 ~
0.0 a 0.{) ~
73.5~12.4 a 82.8 + 9.4 "~b
55.0+15.9 a 59.5 ~ 15.5"
0.0 a 0.0 a
P ' < 0 . 0 5 vs. 0.25 m g / k g group. h P < 0 . 0 5 vs. 0.50 m g / k g group.
0.05) greater hyperthermic response than lower doses. Furthermore, the hyperthermic effect was evident for the entire 3 h period of observation (table 2).
table 1, buprenorphine at a dose of 0.25 m g / k g s.c., produced 46.7% analgesia at 30 min after its administration. This level of analgesia was maintained for 90 min. The analgesic response decreased rapidly and was only 11.8 and 4.0%, respectively at 120 and 180 min after buprenorphine injection. A dose of 0.5 m g / k g of buprenorphine maintained 100% analgesia for 90 min which was then reduced to half at 120 and 180 min after its administration. A higher level of analgesia was obtained with 1 and 2 m g / k g doses of buprenorphine. The administration of buprenorphine produced a hyperthermic response in the rat. Thirty min after buprenorphine (0.25 mg/kg) injection, the body temperature of rats increased by 0.84°C. The increase in body temperature was dose related since higher doses produced a significantly ( P <
3.2. Effect of peptides on development of tolerance to the analgesic action of buprenorphine Based upon the dose-response relationship studies with buprenorphine on analgesic and hyperthermic responses, a dose of 0.5 m g / k g was selected for the tolerance studies. Chronic administration of buprenorphine (0.5 m g / k g s.c.) twice a day for 4 days resulted in the development of tolerance to the analgesic effect of buprenorphine and this tolerance was inhibited by daily injections of MIF or cyclo (Leu-Gly). As can be seen from table 3, administration of buprenorphine (0.5
TABLE 2 D o s e - r e s p o n s e relationship for the h y p e r t h e r m i c effect of b u p r e n o r p h i n e in the rat. D o s e of buprenorphine
I n c r e a s e in rectal t e m p e r a t u r e ( ° C ) , m e a n + S.E.M. (n = 6) T i m e a f t e r b u p r e n o r p h i n e injection (min)
m g / k g s.c.
0.25 0.50 1.00 2.00
30
60
90
120
180
0.84+0.13 1.27_+0.16 a 1.79 ± 0.22 a.b 1.78 ± 0.28 a.h
1.30+0.21 1.69+0.23 2.06 + 0. I I a 1.98 ± 0. I 8 ~'
1.18+0.13 1.30-~ 0.18 2.05 ± 0.09 a.b 2.00 + 0.15 ~.b
0.93+0.15 1.10+0.12 1.64 + 0.12 a,b 1.93 + 0.12 ~.b,¢
0. 9 0 + 0 . 2 2 0,84 ± 0.06 1.03 + 0.23 1.79 + 0. I 0 ~,bx
P < 0 . 0 5 vs. 0.25 m g / k g group. b P < 0 . 0 5 vs. 0.50 m g / k g group. " P < 0 . 0 5 vs. 1.00 m g / k g group.
120 TABL E 3 Effect of Pro-Leu-Gly-NH 2 (MIF) and cyclo (Leu-Gly) (CLG) on tolerance to the analgesic action of buprenorphine in the rat. Treatment a
Water + water MIF+water CLG+water Water+buprenorphine MIF+buprenorphine CLG+buprenorphine
n
6 6 6 8 8 8
Percent analgesia, mean -+ S.E.M. Time after buprenorphine injection (min) 30
60
90
88.8 + 8.1 86.7 -+ 9.3 83.3-+ 9.5 14.1-+ 3.5 b 32.6-+ 11.5 ~ 32.1 + 8.5 c
77.8 -+ 11.1 80.7-+ 8.4 8 0 . 2+- 10.7 1.5--+ 0.1 t, 23.4-+ 8.8 c 19.9-+ 4.3 ~
34.3 -+ 5.6 34.5+6.0 40.5-+7.7 1.5-+0.1 b 12.4-+5.3 ~ 12.1 +2.8 c
" Rats were injected with vehicle (water) or the peptide (2 m g / k g s.c.) and then injected with buprenorphine (0,5 m g / k g s.c. twice a day) or its vehicle (water). This procedure was repeated for 4 days. On day 5, analgesic response to buprenorphine (0.5 m g / k g s.c.) was determined for each treatment group. b P < 0 . 0 5 vs. the corresponding water-treated group. c P < 0 . 0 5 vs. the w a t e r + b u p r e n o r p h i n e group.
mg/kg) produced 88.8, 77.8 and 34.3% analgesia at 30, 60 and 90 min respectively after its injection in rats treated chronically with water (vehicle). This analgesic response was not modified by daily injections of either MIF or cyclo (Leu-Gly) (P = 0.15). However, the analgesic activity of buprenorphine was modified by the peptides in buprenorphine tolerant rats (P < 0.001). There was also a significant difference in the analgesic response for water and buprenorphine groups ( P < 0.0001). Further analysis indicated that the peptide vehicletreated water and buprenorphine groups differed significantly at 30, 60 and 90 rain (P < 0.0001) in response to a challenge dose of buprenorphine. Similarly, for 30, 60 and 90 min MIF-treated and CLG-treated water and buprenorphine groups differed significantly (P < 0.0001). Analyses of the data by Schaffe's S-test indicated that the treatment with peptides significantly inhibited the development of tolerance to the analgesic effect of buprenorphine. However, the response to buprenorphine in MIF and CLG groups did not differ (table 3). In contrast to 88.8% analgesia at 30 rain after buprenorphine injection in water treated rats, only 14.1% analgesia was observed in rats treated chronically with buprenorphine. The analgesia was practically absent (1.5%) at 60 and 90 min after buprenorphine injection in buprenorphine tolerant rats.
3.3. Effect of peptides on the development of tolerance to the hyperthermic response of buprenorphine Chronic administration of buprenorphine resulted in the development of tolerance to its hyperthermic effect. As shown in table4, thirty min after the buprenorphine (0.5 mg/kg) injection, an increase in body temperature by 1.76°C in nontolerant rats was noted, whereas, only 0.97°C increase was seen in rats which were given buprenorphine chronically. However, the hyperthermic effect of buprenorphine was significantly altered by chronic injections of MIF or CLG (P<0.001). The chronic water and chronic buprenorphine groups also differed significantly (P < 0.001). As seen with analgesia, the hyperthermic response to buprenorphine differed significantly in water-, MIF- or CLG-treated groups at all time intervals. Analyses of data by Schaffe's S-test indicated that daily administration of MIF or cyclo (Leu-Gly) did not alter the hyperthermic response of buprenorphine in rats which were given vehicle (water) chronically. However, daily injections of MIF or cyclo (Leu-Gly) completely blocked the development of tolerance to the hyperthermic effect of buprenorphine. The increase in body temperature after buprenorphine (0.5 mg/kg) injection in chronically vehicle injected and chronically buprenorphine injected rats treated with peptides did not differ.
121
TABLE 4 Effect of Pro-Leu-Gly-NH 2 (MIF) and cyclo (Leu-Gly) (CLG) on tolerance to the hyperthermic response of buprenorphine in the rat. Treatment a
W a t e r + water MIF+water CLG+water Water + buprenorphine M I F + buprenorphine C L G + buprenorphine
n
6 6 6 8 8 8
Increase in rectal temperature, (°C), mean ± S.E.M. Time after buprenorphine injection (min) 30
60
90
1.76 ÷ 0.24 1.64+0.25 1.55-+0.13 0.97_+0.04 b 1.65 -+ 0.10 c 1.61 ± 0.10 c
1.60 + 0.20 1.48=0.24 1.37-+0.13 0.87±0.06 b 1.63 ± 0.10 c 1.48 ± 0.17 c
1.49 ± 0.18 1.30±0.21 1.18-+0.16 0.71 -+0.06 b 1.25 ± 0.10 ~ 1.31 + 0.16 c
a Rats were injected with vehicle (water) or the peptide (2 m g / k g s.c.) and then injected with buprenorphine (0.5 m g / k g s.c. twice a day) or its vehicle (water). This procedure was repeated for 4 days. On day 5, the thermic response to buprenorphine (0.5 m g / k g s.c.) was determined for each treatment group. b P < 0 . 0 5 vs. the w a t e r + w a t e r group. c P < 0 . 0 5 vs. the w a t e r + b u p r e n o r p h i n e group.
4. Discussion
Chronic administration of buprenorphine resuited in the development of tolerance to its analgesic and hyperthermic effects in the rat. The development of tolerance to the analgesic effect of buprenorphine has been reported in the mouse also (Cowan et al., 1977b). Although Cox et al. (1976) failed to show tolerance to the hyperthermic effect of opiate agonists, like morphine, others have shown that tolerance indeed develops to the hyperthermic effects of both the endogenous and the exogenous opiates (Huidobro-Toro and Way, 1980; Bhargava, 1981e). This study demonstrates that agonist-antagonist buprenorphine produces hyperthermia and tolerance develops to it. It must be noted that the inhibition of development of tolerance to the analgesic effect of buprenorphine was only partial, whereas the effect on tolerance to the hyperthermic effect was more pronounced. It is possible that different mechanisms may be involved in the two processes. The evidence for this stems from the fact that another hypothalamic peptide, thyrotropin releasing hormone (TRH) which does not antagonize morphine induced analgesia (Martin et al., 1977) was effective in blocking the development of tolerance to the analgesic but not to the hypothermic effect of morphine in mice (Bhargava, 1981f).
The mechanism by which the peptides inhibit the development of opiate-induced tolerance is not evident at present. However, there may be several different ways that the peptides could influence the development of tolerance to opiates. It is possible that these peptides may be exerting a narcotic antagonistic effect, since narcotic antagonists, like naloxone and naltrexone can prevent narcotic tolerance and dependence development (Mushlin and Cochin, 1976; Bhargava, 1978; Huidobro-Toro and Way, 1980). Indeed, MIF has been shown to possess naloxone like activity in some tests. For instance, morphine-induced analgesia was antagonized by MIF when given 10 min prior to morphine injection; however, unlike naloxone, MIF was inactive in reversing the Straub-tail reflex, or the inhibition of electrically induced contraction of the vas deferens (Kastin et al., 1979). Kastin et al. (1980) found that a 15 mg/kg dose of morphine in mice maintained high level of analgesia for more than three hours. Administration of MIF (0.01-10 mg/kg) significantly antagonized the analgesic effect of morphine (15 mg/kg). In contrast, Chiu and Mishra (1979) were unable to observe an antagonism of morphine-induced analgesia by MIF in doses of 1.0-40 mg/kg. A study by Dickinson and Slater (1980) indicated that in rats, chronic treatment with MIF (1 mg/kg per day for 10 days) decreased slightly the analgesic
122
effect of morphine. In contrast, acute treatment with MIF (2 mg/kg) had no effect on morphine (2 or 4 mg/kg) induced analgesia; chronic treatment for 5 days with MIF ( 2 m g / k g per day) did not antagonize analgesia induced by 2 mg/kg of morphine but not that induced by 4 mg/kg of morphine. Our preliminary studies indicate that MIF (0.5-2 mg/kg) treatment antagonizes analgesia induced by 5 mg/kg of morphine but not that produced by 2.5 and 10 mg/kg (unpublished observations). These studies show that complex interactions may be taking place between MIF and opiates. Detailed studies are under way to characterize MIF as an opiate antagonist. It is known that opiates, on chronic administration, induce supersensitivity of brain dopamine receptors (Lal, 1975; Ritzmann et al., 1979; Bhargava, 1980, 1981 a) which results from chronic depression of dopaminergic transmission. The opiate induced supersensitivity of dopamine receptors is blocked by MIF and cyclo (Leu-Gly) (Ritzmann et al., 1979; Bhargava, 1980, 1981a). Buprenorphine also interacts with central dopaminergic systems as evidenced by an elevation in rat forebrain homovanillic acid concentration following buprenorphine treatment (Cowan et al., 1976). Since MIF is known to potentiate the behavioral effects of 1-dopa (Plotnikoff et al., 1971) it is possible that the inhibition of development of tolerance to buprenorphine by peptides may involve central dopaminergic systems.
Acknowledgements These studies were supported in part by a USPHS grant DA-02598 from the National Institute on Drug Abuse. The author thanks George Matwyshyn for providing excellent technical assistance, and Ms. Dorothy Laverne Guilty for her help in preparation of this manuscript.
References Bhargava, H.N., 1978, The effects of naltrexone on the development of physical dependence on morphine, European J. Pharmacol. 50, 193. Bhargava, H.N., 1980, Cyclo (leucylglycine) inhibits the development of morphine induced analgesic tolerance and dopamine receptor supersensitivity in rats, Life Sci. 27, 117.
Bhargava, H.N., 1981a, The effect of melanotropin release inhibiting factor and cyclo (Leu-Gly) on tolerance to morphine induced antinociception in the rat: a dose response study, Br. J. Pharmacol. 72, 707. Bhargava, H.N., 1981b, The effect of peptides on tolerance to the cataleptic and hypothermic effect of morphine in the rat, Neuropharmacology 20, 385. Bhargava, H.N., 1981c, Effects of Pro-Leu-Gly-NH 2 and cyclo (Leu-Gly) on tolerance to the pharmacological actions of human beta endorphin in the rat, Fed. Proc. 40, 304. Bhargava, H.N., 1981d, Inhibition of tolerance to the pharmacological effects of human beta endorphin by prolylleucylglycinamide and cyclo (leucylglycine) in the rat, J. Pharmacol. Exp. Ther. 218, 404. Bhargava, H.N., 1981e, Structure activity relationship studies with hypothalamic peptide hormones. Effect on opiate induced tolerance, The Pharmacologist 23, 120. Bhargava, H.N., 1981f, Dissociation of tolerance to the analgesic and hypotherrnic effects of morphine by using thyrotropin releasing hormone, Life Sci. 29, 1015. Bhargava, H.N., R. Walter and R.F. Ritzmann, 1980, Development of narcotic tolerance and physical dependence: effects of Pro-Leu-Gly-NH 2 and cyclo (Leu-Gly), Pharmacol. Biochem. Behav. 12, 73. Chiu, S. and R.K. Mishra, 1979, Antagonism of morphineinduced catalepsy by L- Propyl- L-leucylglycinamide, European J. Pharmacol. 53, 119. Cowan, A., 1974, Evaluation in non-human primates: evaluation of the physical dependence capacities of oripavinethebaine partial agonists in Patas Monkeys, Adv. Biochem. Psychopharmacol. 8, 427. Cowan, A., D.W. Dettmar and D.S. Walter, 1976, The effect of acute doses of buprenorphine on concentrations of homovanillic acid (HVA), 5-hydroxyindole-acetic acid (5HIAA) and 3-methoxy-4-hydroxy-phenylglycol (MHPG) in the rat forebrain, Br. J. Pharmacol. 58, 275P. Cowan, A., J.C. Doxey and E.J.R. Harry, 1977a, The animal pharmacology of buprenorphine, an oripavine analgesic agent, Br. J. Pharmacol. 60, 547. Cowan, A., J.W. Lewis and I.R. Macfarlane, 1977b, Agonist and antagonist properties of buprenorphine, a new antinociceptive agent, Br. J. Pharmacol. 60, 537. Cox, B., M. Ary, W. Chesarek and P. Lomax, 1976, Morphine hyperthermia in the rat: an action on the central thermostat, European J. Pharmacol. 36, 33. Dickinson, S.L. and P. Slater, 1980, Opiate receptor antagonism by L-Propyl-L-Leucyl-Glycinamide, MIF-1, Peptides 1,293. Fischer, E., 1906, Synthese von polypeptiden XV, Chem. Ber, 39, 2893. Huidobro-Toro, J.P. and E.L. Way, 1980, Rapid development of tolerance to the hyperthermic effect of fl-endorphin, European J. Pharmacol. 65, 221. Jacob, J.J.C., J.M. Michaud and E.C. Tremblay, 1979. Mixed agonist opiate and physical dependence, Br. J. Clin. Pharmacol. 7, 291 S. Jasinski, D.R., J.S. Pevnick and J.D. Griffith, 1978, Human pharmacology and abuse potential of the analgesic buprenorphine, Arch. Gen. Psychiat. 35, 501.
123 Kastin, A.J., C. Nissen, J.E. Zadina, A.V. Schally and R.H. Ehrensing, 1980, Naloxone like actions of MIF-I do not require the presence of the pituitary, Pharmacol. Bioehem. Behav. 13, 907. Kastin, A.J., R.D. Olson, R.H. Ehrensing, M.C. Berzas, A.V. Schally and D.H. Coy, 1979, MIF-I's differential actions as an opiate antagonist, Pharmacol. Biochem. Behav. 1 I, 72 I. Kirk, R.E., 1968, in: Experimental Design: Procedures of Behavioral Sciences, (Brooks/Cole Publishing Co., Los Angeles) p. 245. Lal, H., 1975, Narcotic dependence, narcotic action and dopamine receptors, Life Sci. 17, 483. Martin, B.R., W.L. Dewey, T. Chau-Pham and A.J. Prange, Jr.,
1977, Interactions of thyrotropin releasing hormone and morphine sulfate in rodents, Life Sci. 20, 715. Mello, N.K. and J.H. Mendelson, 1980, Buprenorphine suppresses heroin use by heroin addicts, Science 207, 657. Mushlin, B.E. and J. Cochin, 1976, Tolerance to morphine in the rat: its prevention by naloxone, Life Sci. 18, 797. Plotnikoff, N.P., A.J. Kastin, M.S. Anderson and A.V. Schally, 1971, Dopa potentiation by hypothalamic factor MSH release inhibiting hormone (MIF), Life Sci. 10, 1279. Ritzmann, R.F., R. Walter, H.N. Bhargava and L.B. Flexner, 1979, Blockage of narcotic-induced dopamine receptor supersensitivity by cyclo (Leu-Gly), Proc. Natl. Acad. Sci. U.S.A. 76, 5997.
117
EFFECT OF PEPTIDES ON THE DEVELOPMENT OF TOLERANCE TO BUPRENORPHINE, A MIXED OPIATE AGONIST-ANTAGONIST ANALGESIC HEMENDRA N. BHARGAVA
Department of Pharmacognosy and Pharmacology, College of PharmaQ,, University of Illinois at the Medical Center, Chicago, Illinois 60612, U.S.A. Received 23 December 1981, accepted ! February 1982
H.N. BHARGAVA, Effect of peptides on the development of tolerance to buprenorphine, a mixed opiate agonist-antagonist analgesi~, European J. Pharmacol. 79 (1982) I 17-123. The subcutaneous administration of buprenorphine to male Sprague-Dawley rats produced a dose-dependent analgesia and hyperthermia in a dose range of 0.25-2 mg/kg. The subcutaneous administration of buprenorphine (0.5 mg/kg) twice a day for 4 days resulted in the development of tolerance to its analgesic and hyperthermic actions. Daily administration of melanotropin release inhibiting factor (Pro-Leu-Gly-NH2) or cyclo (Leu-Gly) (2 mg/kg s.c.) for 4 days, inhibited the development of tolerance to buprenorphine, as evidenced by greater analgesic and hyperthermic responses to buprenorphine in peptide-treated than in vehicletreated buprenorphine-tolerant rats. The repeated injections of peptides did not alter the analgesic or the hyperthermic response to buprenorphine in non-tolerant rats. These studies suggest the possible role of hypothalamic peptides in blocking the development of tolerance to the pharmacological effects of buprenorphine, a potent analgesic agent. Mixed agonist-antagonist
Buprenorphine
MIF
Pro-Leu-Gly-NH 2
Tolerance
Analgesia
Hyperthermia
Cyclo(Leu-Gly)
1. Introduction
Buprenorphine is an oripavine derivative possessing both opiate agonist and antagonist activity. Animal studies indicate that it is a potent analgesic agent having 25-40 times greater potency than morphine after parenteral administration and 7-10 times greater potency after oral administration; it has rapid onset and long duration of action (Cowan et al., 1977a). Furthermore, buprenorphine possesses low physical dependence liability (Cowan, 1974; Cowan et al., 1977b; Jacob et al., 1979). The studies with buprenorphine in humans indicate that the subjective effects experienced by heroin addicts were similar to those of morphine (Jasinski et al., 1978). In addition to its morphine like properties, buprenorphine also has a long duration of action as an opiate antagonist and antagonizes the actions of high doses of morphine for longer than 24h (Jasinski et al., 1978). The same authors (Jasinski et al., 1978) also reported 0014-2999/82/0000-0000/$02.75 © 1982 Elsevier Biomedical Press
that patients maintained on high doses of buprenorphine experience an extremely mild withdrawal syndrome two weeks after the abrupt termination of buprenorphine. A more recent study indicates that buprenorphine suppressed the self administration of heroin in heroin-dependent men; termination of buprenorphine did not result in opiate withdrawal signs and symptoms, and the dependent subjects preferred it over methadone or naltrexone (Mello and Mendelson, 1980). The multiple administration of buprenorphine results in the development of tolerance to its analgesic action as determined by the phenylquinone-writhing test (Cowan et al., 1977b). Since the major uses of buprenorphine is as a potent analgesic agent and as a pharmaco-therapeutic agent for the treatment of heroin dependence, the development of tolerance to its analgesic action is an undesirable effect. Previous studies from these laboratories have indicated that certain hypothalamic peptide hormones, like melanotropin release inhibiting factor, a tripeptide, Pro-Leu-Gly-
118
2. Materials and methods
temperature of each rat was measured prior to and at 30, 60, 90, 120 and 180 min after the drug injection by using a rectal probe and a telethermometer (Yellow Spring Instrument Co., Yellow Springs, Ohio). The animals were not restrained. They were lightly hand handled. The change in rectal temperature for each rat at each time interval was calculated as the difference between predrug and post-drug temperatures. The data are expressed as mean increases in rectal temperature, °C -+S.E.M. Six rats were used for each dose of buprenorphine.
2.1. Animals and chemicals
2.3. Effect of peptides on tolerance to buprenorphine
Male Sprague-Dawley rats weighing 150-200 g obtained from King Animal Laboratories, Oregon, Wis., were housed 3 to a cage in a room with controlled temperature (23 -+ 1°C), humidity (65 -+ 2%) and a 12 h dark-light cycle (L 06 : 00-18 : 00 h). Rats were acclimated to the above conditions for at least four days before being used and were given food and tap water ad libitum. MIF was received as a gift from the Abbott Laboratories, N. Chicago, Illinois through the courtesy of Dr. A.O. Geiszler. Cyclo (Leu-Gly) was synthesized in these laboratories as described previously (Fischer, 1906). Buprenorphine was kindly donated by Dr. A. Cowan of Temple University, Philadelphia, Pennsylvania. The peptides and buprenorphine were dissolved in distilled deionized water and administered subcutaneously (s.c.) in volumes such that each rat received 1 m l / k g of the drug solution.
The effect of MIF and cyclo (Leu-Gly) on the development of tolerance to buprenorphine were studied. Tolerance to buprenorphine was induced by sub'cutaneous injections of buprenorphine (0.5 mg/kg) twice daily at 9 a.m. and 4.30 p.m. for 4 days. The rats were injected with the appropriate peptide (2 m g / k g s.c.) or its vehicle (water) l h before the first injection of buprenorphine. The above procedure (i.e., single injection of peptide or its vehicle and two injections of buprenorphine and its vehicle) was repeated for 4 days. On day 5, the analgesic and thermic responses to buprenorphine (0.5 m g / k g s.c.) in rats from all six treatment groups were assessed as described above. Eight rats were used in buprenorphine groups and 6 rats were used in its vehicle groups.
N H 2 (MIF) and its analog cyclo (Leu-Gly) block the tolerance to morphine in mice and rats (Bhargava et al., 1980; Bhargava, 1980, 1981a,b,e) and to human fl-endorphin in the rat (Bhargava, 198 lc,d). It was, therefore, of interest to determine whether these peptides can also block the tolerance to an agonist-antagonist type analgesic, buprenorphine. The present paper describes such studies in rats.
2.2. Measurement of buprenorphine-induced gesia and changes in body temperature
anal
The analgesic response (% analgesia) to various doses (0.25-2 mg/kg) of buprenorphine was determined using a tail-flick procedure described previously (Bhargava, 1981a). The analgesic response was determined at 30, 60, 90, 120 and 180 min after buprenorphine injection. The data are expressed as mean % analgesic response -+S.E.M. Six rats were used for each dose of buprenorphine. The rats were injected with various doses of buprenorphine (0.25-2.0 m g / k g s.c.) and the rectal
2.4. Statistics The differences in the means of various treatment groups were analyzed by the analysis of variance followed by Schaffe's S-test (Kirk, 1968). The difference was considered statistically significant when the P value was less than 0.05.
3. Results
3.1. Effect of buprenorphine on analgesia and body temperature The administration of buprenorphine produced a dose-related analgesia in the rat. As shown in
119
TABLE 1 D o s e - r e s p o n s e relationship for the analgesic effect of b u p r e n o r p h i n e in the rat. D o s e of
P e r c e n t analgesia, m e a n + S.E.M. ( n - 6 )
buprenorphine
T i m e after b u p r e n o r p h i n e injection ( m i n )
m g / k g s.c. 30
60
0.25
46.7 + 17.7
0.50
94.3+
1.00 2.00
100 100
+ +
9(/
41.8 + 13.5
120
45.2 + 17.7
11.8 +
180 4.1
4.0 +
1.5
3.8 a
100
-- 0.0 a
I00
+
0.0 a
4 3 . 0 ~ 1 4 . 7 ~'
46.8+17.6 a
0.0 a 0.0 a
100 100
± +
I(X) + 100 ~
0.0 a 0.{) ~
73.5~12.4 a 82.8 + 9.4 "~b
55.0+15.9 a 59.5 ~ 15.5"
0.0 a 0.0 a
P ' < 0 . 0 5 vs. 0.25 m g / k g group. h P < 0 . 0 5 vs. 0.50 m g / k g group.
0.05) greater hyperthermic response than lower doses. Furthermore, the hyperthermic effect was evident for the entire 3 h period of observation (table 2).
table 1, buprenorphine at a dose of 0.25 m g / k g s.c., produced 46.7% analgesia at 30 min after its administration. This level of analgesia was maintained for 90 min. The analgesic response decreased rapidly and was only 11.8 and 4.0%, respectively at 120 and 180 min after buprenorphine injection. A dose of 0.5 m g / k g of buprenorphine maintained 100% analgesia for 90 min which was then reduced to half at 120 and 180 min after its administration. A higher level of analgesia was obtained with 1 and 2 m g / k g doses of buprenorphine. The administration of buprenorphine produced a hyperthermic response in the rat. Thirty min after buprenorphine (0.25 mg/kg) injection, the body temperature of rats increased by 0.84°C. The increase in body temperature was dose related since higher doses produced a significantly ( P <
3.2. Effect of peptides on development of tolerance to the analgesic action of buprenorphine Based upon the dose-response relationship studies with buprenorphine on analgesic and hyperthermic responses, a dose of 0.5 m g / k g was selected for the tolerance studies. Chronic administration of buprenorphine (0.5 m g / k g s.c.) twice a day for 4 days resulted in the development of tolerance to the analgesic effect of buprenorphine and this tolerance was inhibited by daily injections of MIF or cyclo (Leu-Gly). As can be seen from table 3, administration of buprenorphine (0.5
TABLE 2 D o s e - r e s p o n s e relationship for the h y p e r t h e r m i c effect of b u p r e n o r p h i n e in the rat. D o s e of buprenorphine
I n c r e a s e in rectal t e m p e r a t u r e ( ° C ) , m e a n + S.E.M. (n = 6) T i m e a f t e r b u p r e n o r p h i n e injection (min)
m g / k g s.c.
0.25 0.50 1.00 2.00
30
60
90
120
180
0.84+0.13 1.27_+0.16 a 1.79 ± 0.22 a.b 1.78 ± 0.28 a.h
1.30+0.21 1.69+0.23 2.06 + 0. I I a 1.98 ± 0. I 8 ~'
1.18+0.13 1.30-~ 0.18 2.05 ± 0.09 a.b 2.00 + 0.15 ~.b
0.93+0.15 1.10+0.12 1.64 + 0.12 a,b 1.93 + 0.12 ~.b,¢
0. 9 0 + 0 . 2 2 0,84 ± 0.06 1.03 + 0.23 1.79 + 0. I 0 ~,bx
P < 0 . 0 5 vs. 0.25 m g / k g group. b P < 0 . 0 5 vs. 0.50 m g / k g group. " P < 0 . 0 5 vs. 1.00 m g / k g group.
120 TABL E 3 Effect of Pro-Leu-Gly-NH 2 (MIF) and cyclo (Leu-Gly) (CLG) on tolerance to the analgesic action of buprenorphine in the rat. Treatment a
Water + water MIF+water CLG+water Water+buprenorphine MIF+buprenorphine CLG+buprenorphine
n
6 6 6 8 8 8
Percent analgesia, mean -+ S.E.M. Time after buprenorphine injection (min) 30
60
90
88.8 + 8.1 86.7 -+ 9.3 83.3-+ 9.5 14.1-+ 3.5 b 32.6-+ 11.5 ~ 32.1 + 8.5 c
77.8 -+ 11.1 80.7-+ 8.4 8 0 . 2+- 10.7 1.5--+ 0.1 t, 23.4-+ 8.8 c 19.9-+ 4.3 ~
34.3 -+ 5.6 34.5+6.0 40.5-+7.7 1.5-+0.1 b 12.4-+5.3 ~ 12.1 +2.8 c
" Rats were injected with vehicle (water) or the peptide (2 m g / k g s.c.) and then injected with buprenorphine (0,5 m g / k g s.c. twice a day) or its vehicle (water). This procedure was repeated for 4 days. On day 5, analgesic response to buprenorphine (0.5 m g / k g s.c.) was determined for each treatment group. b P < 0 . 0 5 vs. the corresponding water-treated group. c P < 0 . 0 5 vs. the w a t e r + b u p r e n o r p h i n e group.
mg/kg) produced 88.8, 77.8 and 34.3% analgesia at 30, 60 and 90 min respectively after its injection in rats treated chronically with water (vehicle). This analgesic response was not modified by daily injections of either MIF or cyclo (Leu-Gly) (P = 0.15). However, the analgesic activity of buprenorphine was modified by the peptides in buprenorphine tolerant rats (P < 0.001). There was also a significant difference in the analgesic response for water and buprenorphine groups ( P < 0.0001). Further analysis indicated that the peptide vehicletreated water and buprenorphine groups differed significantly at 30, 60 and 90 rain (P < 0.0001) in response to a challenge dose of buprenorphine. Similarly, for 30, 60 and 90 min MIF-treated and CLG-treated water and buprenorphine groups differed significantly (P < 0.0001). Analyses of the data by Schaffe's S-test indicated that the treatment with peptides significantly inhibited the development of tolerance to the analgesic effect of buprenorphine. However, the response to buprenorphine in MIF and CLG groups did not differ (table 3). In contrast to 88.8% analgesia at 30 rain after buprenorphine injection in water treated rats, only 14.1% analgesia was observed in rats treated chronically with buprenorphine. The analgesia was practically absent (1.5%) at 60 and 90 min after buprenorphine injection in buprenorphine tolerant rats.
3.3. Effect of peptides on the development of tolerance to the hyperthermic response of buprenorphine Chronic administration of buprenorphine resulted in the development of tolerance to its hyperthermic effect. As shown in table4, thirty min after the buprenorphine (0.5 mg/kg) injection, an increase in body temperature by 1.76°C in nontolerant rats was noted, whereas, only 0.97°C increase was seen in rats which were given buprenorphine chronically. However, the hyperthermic effect of buprenorphine was significantly altered by chronic injections of MIF or CLG (P<0.001). The chronic water and chronic buprenorphine groups also differed significantly (P < 0.001). As seen with analgesia, the hyperthermic response to buprenorphine differed significantly in water-, MIF- or CLG-treated groups at all time intervals. Analyses of data by Schaffe's S-test indicated that daily administration of MIF or cyclo (Leu-Gly) did not alter the hyperthermic response of buprenorphine in rats which were given vehicle (water) chronically. However, daily injections of MIF or cyclo (Leu-Gly) completely blocked the development of tolerance to the hyperthermic effect of buprenorphine. The increase in body temperature after buprenorphine (0.5 mg/kg) injection in chronically vehicle injected and chronically buprenorphine injected rats treated with peptides did not differ.
121
TABLE 4 Effect of Pro-Leu-Gly-NH 2 (MIF) and cyclo (Leu-Gly) (CLG) on tolerance to the hyperthermic response of buprenorphine in the rat. Treatment a
W a t e r + water MIF+water CLG+water Water + buprenorphine M I F + buprenorphine C L G + buprenorphine
n
6 6 6 8 8 8
Increase in rectal temperature, (°C), mean ± S.E.M. Time after buprenorphine injection (min) 30
60
90
1.76 ÷ 0.24 1.64+0.25 1.55-+0.13 0.97_+0.04 b 1.65 -+ 0.10 c 1.61 ± 0.10 c
1.60 + 0.20 1.48=0.24 1.37-+0.13 0.87±0.06 b 1.63 ± 0.10 c 1.48 ± 0.17 c
1.49 ± 0.18 1.30±0.21 1.18-+0.16 0.71 -+0.06 b 1.25 ± 0.10 ~ 1.31 + 0.16 c
a Rats were injected with vehicle (water) or the peptide (2 m g / k g s.c.) and then injected with buprenorphine (0.5 m g / k g s.c. twice a day) or its vehicle (water). This procedure was repeated for 4 days. On day 5, the thermic response to buprenorphine (0.5 m g / k g s.c.) was determined for each treatment group. b P < 0 . 0 5 vs. the w a t e r + w a t e r group. c P < 0 . 0 5 vs. the w a t e r + b u p r e n o r p h i n e group.
4. Discussion
Chronic administration of buprenorphine resuited in the development of tolerance to its analgesic and hyperthermic effects in the rat. The development of tolerance to the analgesic effect of buprenorphine has been reported in the mouse also (Cowan et al., 1977b). Although Cox et al. (1976) failed to show tolerance to the hyperthermic effect of opiate agonists, like morphine, others have shown that tolerance indeed develops to the hyperthermic effects of both the endogenous and the exogenous opiates (Huidobro-Toro and Way, 1980; Bhargava, 1981e). This study demonstrates that agonist-antagonist buprenorphine produces hyperthermia and tolerance develops to it. It must be noted that the inhibition of development of tolerance to the analgesic effect of buprenorphine was only partial, whereas the effect on tolerance to the hyperthermic effect was more pronounced. It is possible that different mechanisms may be involved in the two processes. The evidence for this stems from the fact that another hypothalamic peptide, thyrotropin releasing hormone (TRH) which does not antagonize morphine induced analgesia (Martin et al., 1977) was effective in blocking the development of tolerance to the analgesic but not to the hypothermic effect of morphine in mice (Bhargava, 1981f).
The mechanism by which the peptides inhibit the development of opiate-induced tolerance is not evident at present. However, there may be several different ways that the peptides could influence the development of tolerance to opiates. It is possible that these peptides may be exerting a narcotic antagonistic effect, since narcotic antagonists, like naloxone and naltrexone can prevent narcotic tolerance and dependence development (Mushlin and Cochin, 1976; Bhargava, 1978; Huidobro-Toro and Way, 1980). Indeed, MIF has been shown to possess naloxone like activity in some tests. For instance, morphine-induced analgesia was antagonized by MIF when given 10 min prior to morphine injection; however, unlike naloxone, MIF was inactive in reversing the Straub-tail reflex, or the inhibition of electrically induced contraction of the vas deferens (Kastin et al., 1979). Kastin et al. (1980) found that a 15 mg/kg dose of morphine in mice maintained high level of analgesia for more than three hours. Administration of MIF (0.01-10 mg/kg) significantly antagonized the analgesic effect of morphine (15 mg/kg). In contrast, Chiu and Mishra (1979) were unable to observe an antagonism of morphine-induced analgesia by MIF in doses of 1.0-40 mg/kg. A study by Dickinson and Slater (1980) indicated that in rats, chronic treatment with MIF (1 mg/kg per day for 10 days) decreased slightly the analgesic
122
effect of morphine. In contrast, acute treatment with MIF (2 mg/kg) had no effect on morphine (2 or 4 mg/kg) induced analgesia; chronic treatment for 5 days with MIF ( 2 m g / k g per day) did not antagonize analgesia induced by 2 mg/kg of morphine but not that induced by 4 mg/kg of morphine. Our preliminary studies indicate that MIF (0.5-2 mg/kg) treatment antagonizes analgesia induced by 5 mg/kg of morphine but not that produced by 2.5 and 10 mg/kg (unpublished observations). These studies show that complex interactions may be taking place between MIF and opiates. Detailed studies are under way to characterize MIF as an opiate antagonist. It is known that opiates, on chronic administration, induce supersensitivity of brain dopamine receptors (Lal, 1975; Ritzmann et al., 1979; Bhargava, 1980, 1981 a) which results from chronic depression of dopaminergic transmission. The opiate induced supersensitivity of dopamine receptors is blocked by MIF and cyclo (Leu-Gly) (Ritzmann et al., 1979; Bhargava, 1980, 1981a). Buprenorphine also interacts with central dopaminergic systems as evidenced by an elevation in rat forebrain homovanillic acid concentration following buprenorphine treatment (Cowan et al., 1976). Since MIF is known to potentiate the behavioral effects of 1-dopa (Plotnikoff et al., 1971) it is possible that the inhibition of development of tolerance to buprenorphine by peptides may involve central dopaminergic systems.
Acknowledgements These studies were supported in part by a USPHS grant DA-02598 from the National Institute on Drug Abuse. The author thanks George Matwyshyn for providing excellent technical assistance, and Ms. Dorothy Laverne Guilty for her help in preparation of this manuscript.
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