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Effects of ethylenediamine on morphine analgesia and tolerance-dependence in mice
Gen. Pharmac. Vol. 16, No. 5, pp. 529-531, 1985
0306-3623/85 $3.00+0.00
Printed in Great Britain
Pergamon Press Ltd
EFFECTS OF ETHYLENEDIAMINE ON MORPHINE ANALGESIA A N D TOLERANCE-DEPENDENCE IN MICE E. CONTRERAS and L. TAMAYO Departamento de Ciencias Fisiolbgicas, Universidad de Concepcibn, Casilla 2407 Concepcirn, Chile (Received 14 January 1985) Abstract--1. Ethylenediamine, a GABA receptor agonist induced a small hyperalgesic state in mice, but increased morphine analgesia. The interaction with this morphine effect was not dose-dependent. 2. Ethylenediamine significantly antagonized tolerance development at relatively low doses (5-10 mg/kg). 3. The GABA mimetic agent increased the frequency of abstinence signs in the naloxone-precipitated morphine withdrawal in mice. 4. The effect of ethylenediamine on morphine withdrawal was suppressed by the irreversible GABA transaminase inhibitor, 7-vinyl GABA.
off the plate or kicking the legs. Each animal was tested twice before drug administration and the values recorded were averaged to obtain a baseline. The effects of ethylenediamine were similarly studied. Reactions times were determined after 30 min for 1 hr 30 min. Ethylenediamine was given i.p. immediately before the administration of the test dose of morphine which was injected s.c. In order to express the total analgesic response the area under the time-response curve was calculated from values obtained every 30 min during the I hr 30 min period. (Winter and Flataker, 1950).
INTRODUCTION
Several lines of investigation have implicated 7 aminobutyric acid ( G A B A ) in the mediation of morphine effects (DeFeudis, 1982). The administration of G A B A agonists, muscimol (Biggio et al., 1977; Sawynok and LaBella, 1982) or its analogue, 4,5,6,7-tetrahydroisoxazolo 5,-4-c pyridin-3-ol (THIP) (Hill et al., 1981; Grognet et aL, 1982) produce analgesia. The inhibitors of G A B A transaminase, aminooxyacetic acid (Yoneda et al., 1976) -vinyl G A B A and ~-acetylenic G A B A (Contreras et aL, 1979) potentiate the antinociceptive action of morphine in mice. In addition Buckett (1980) demonstrated an antinociceptive effect of y-vinyl G A B A . Recently, Perkins et al. (1981) and Bokish et al. (1982) have reported that the electrophoretic administration of ethylenediamine depresses neuronal activity of the central nervous system in various animal species. The drug exerts a displacement of [3H]GABA specifically bound to recognition sites for this neurotransmitter (Bowery et al., 1982) and it promotes the release of [~4C]GABA from rat striatal slices (Lloyd et al., 1982). These results suggest that ethylenediamine may induce changes in the acute and chronic morphine effects. Therefore, it was decided to study the influence of the drug in morphine analgesia and in the induction of tolerance to and physical dependence on the narcotic analgesic.
Induction of tolerance and physical dependence Tolerance and dependence were induced by a slow release preparation of morphine of the following composition: morphine (as free base) 300mg, sorbital sesquioleate 0.80ml, liquid paraffin 4.20ml and 0.9~ NaCI 5.00ml. Each mouse received 300 mg/kg s.c. of morphine. The degree of tolerance was expressed as the ratio of the mean analgesic effect of a test dose of morphine in naive and in primed mice. The antagonism or attenuation of tolerance was determined by the statistical difference between the effects of the test dose in primed untreated animals and those observed in primed treated mice. A level of probability of 0.01 was accepted as significant. This criterion was reinforced by the requirement that a decrease of at least 3 units in the degree of tolerance had to be observed between the values of the treated and the primed untreated groups.
MATERIALS AND METHODS
Adult female mice from a strain raised at our laboratory were maintained under standardized conditions of light (0800--1800 hr), temperature (22 ___2°C) and food. The drugs used were morphine hydrochloride (May and Baker), naloxone hydrochloride (gift from Endo Labs Inc.), ethylenediamine dihydrochloride (L. Light and Co. Ltd) and 7-vinyl GABA (gift from Centre de Recherche Merrel International, Strasbourg, France).
Naloxone precipitated abstinence After 30hr of the slow-release administration of morphine, mice were injected with a dose of naloxone (4 mg/kg) that produced signs of severe abstinence such as diarrhoea, paw tremors, body shakes, and others indicated in the results. The number of mice that showed these signs in a 20min observation period was recorded. The z2-test was used for statistical analysis of the differences between the control (untreated) and treated mice. A level of probability of 0.05 was accepted as significant.
Analgesia Analgesia was assessed by the hot plate method (Eddy et al., 1950) at 55 + 0.5°C. The responses consisted of jumping O.P, 1 6 / ~ 3
529
RESULTS AND DISCUSSION
Ethylenediamine per se exerts a small hyperalgesic state as seen by thermal stimulation in mice; this effect is significant only at the highest dose level tested
530
E. CONTRERAS a n d L. TAMAYO Table 1. Effects of ethylenediamine on morphine analgesia in mice Drug, dose (mg/kg), time after after ethylenediamine administration Saline (control) Morphine, 5 (control)
Analgesic effect ~ 22 __.23 323 _+40 Ethylenediamine alone
Ethylenediamine
20, 30 min 50, 30min 100, 30 min 150, 30 min 200, 30 min
-36 -49 -25 -41 -61
± ± ± ± ±
Ethylenediamine plus morphine (5 mg/kg)
40 51 17 46 142
510 ± 495 ± 521 + 472 ± 547 ±
23 22 36 37 40
P 0.001 0.001 0.005 0.02 0.005
tAssessed by the hot plate procedure in mice and expressed as the area under the time-response curve relating the increase in reaction time and time intervals + SEM. P refers to the statistical significance with reference to the effect of morphine alone. 2Significantly different from saline. (P < 0.005). Number of mice: 14 per group. Table 2. Effect of ethylenediamine on morphine tolerance in mice Analgesic response to a test dose of morphine 5 mg/kg s.c. in mice (hot plate). Area under the time-response curve ± SEM Experimental condition Saline Morphine (untreated) Ethylenediamine 5 mg/kg Ethylenediamine 5 mg/kg twice Ethylenediamine 10 mg/kg Ethylenediamine 20 mg/kg Ethylenediamine 40 mg/kg twice
Degree of tolerance t
Naive
Primed
22 + 23
25 ± 23
323 _ 40
35 ± 41
9.2
340 _ 41
147 ± 473
2.3
326 ± 37
290 ± 613
1.1
330 _ 38
345 ± 523
0.9
568 ± 572
407 ± 433
1.4
486 + 422
406 ± 683
1.2
tRatio of the mean analgesic effect to a test dose in naive and in primed mice. 2Significantly different than the effects of morphine in naive untreated mice. P < 0.01. 3Significantly higher than the effects of morphine in primed untreated mice. P < 0.01. Number of mice: 14 per group. Ethylenediamine was administered 15 rain before the priming dose of morphine, when administering a second dose, this was given 8 hr after the priming dose of the analgesic.
in this work. However, the proposed GABA agonist induces a synergistic effect on the analgesic effect of morphine, sharing the same ability as other GABA mimetics (Biggio et al., 1977; Hill et al., 1981). The synergistic effect of ethylenediamine on morphine analgesia is not dose-related since the different doses assayed exhibited the same potency (Table 1). In tolerant mice, the drug induced a greater increase of analgesia than in the naive mice. In tolerant mice the
effect of the GABA mimetic was initially dosedependent following saturation kinetics (Table 2). With respect to the precipitated morphine withdrawal, ethylenediamine increased the frequency of the abstinence signs when administered either during the course of dependence or 60 min before naloxone (Table 3). The results presented here are in partial agreement with previous observations in drugs affecting GABA
Table 3. Effects of ethylenediamine on the incidence of abstinence signs in morphine treated mice ~
Abstinence sign Jumping Body shakes Paw tremors Abnormal posturing Diarrhoea
Saline
Ethylenediamine during the course of dependence 2
Ethylenediamine during the course of dependence 3
Ethylenediamine 20 mg/kg 1 hr before naloxone
y-vinyl GABA 400 mg/kg 4 hr before and ethylenediamine 20 mg/kg 1 hr before naloxone
6/12 0/12 7/12
8/14 2/14 12/14
8/14 5_5~4 14/14
9/14 ~ 13/14
1~ 3/12 2A]2
8/12 11/ 12
8/14 8/14
10/14 8/14
12/14 7~
0~
qnduced by naloxone, 4 mg/kg i.p., 30 hr after the administration of a sustained release preparation of morphine (300 mg/kg s.c.). 2Ethylenediamine 20 mg/kg administered 15 min before the priming dose of morphine. 3Ethylenediamine 50 mg/kg administered 15 min before the priming dose of morphine. Underlined values are statistically different from those observed in untreated mice (P < 0.05).
531
Ethylenediamine and morphine effects functions. Thus, irreversible GABA transaminase inhibitors increased analgesia, attenuated tolerance and greatly reduced the abstinence behavior in mice (Contreras et al., 1979), whereas in the present experiments ethylenediamine increased the abstinence syndrome. These discrepancies in drugs acting through GABAergic mechanisms may indicate that ethylenediamine exerts a relatively low intrinsic activity at GABA recognition sites in contrast with the GABA transaminase inhibitors which increase the amounts of the inhibitor neurotransmitter in the nervous tissue. The ability of y-vinyl GABA to suppress the enhancing action of ethylenediamine in the abstinence behavior in mice, also supports a possible low intrinsic activity of the GABA receptor agonist. The results of the present work would confirm the involvement of GABA in some morphine effects as it has been suggested by several authors. SUMMARY Recent studies have implicated the inhibitory neurotransmitter y-aminobutyric acid (GABA) in the morphine effects. Ethylenediamine specifically binds to GABA receptors and releases GABA from nervous tissue in the rat, In the present work the influence of ethylenediamine has been studied in morphine analgesia and tolerance to and physical dependence on morphine in mice. Ethylenediamine increased morphine analgesia in the hot plate test in mice but this effect was not dose-related. Low doses of ethylenediamine (5-10 mg/kg) significantly antagonized tolerance development to morphine in mice. The drug increased the incidence of abstinence signs in the naloxone precipitated morphine withdrawal when administered either during the course of dependence or 1 hr before the precipitating dose of naloxone. The effect of ethylenediamine on the naloxone precipitated abstinence was suppressed by the irreversible GABA transaminase inhibitor, y-vinyl GABA. The results of the present paper would support the involvement of GABA in some morphine effects. Acknowledgement--This work was supported by Grant
20.33.13 from Direccirn de Investigacirn, Universidad de Concepcirn. REFERENCES
Biggio D., Della Bella D., Frigeni V. and Guidotti A. (1977) Potentiation of morphine analgesia by muscimol. Neuropharmacology 16, 149-150. Bokisch A. J., Bold J. M., Perkins M, N., Roberts C. J., Stone T. W. and Walker R. J. (1982) Actions of ethylenediamine on Limulus and Helix central neurons and on rat cerebellar and sympathetic ganglion neurons. Br. J. Pharmac. 76, 297P. Bowery N. G., Hill D. R., Hudson A. L., Perkins M. N. and Stone T. W. (1982) GABA receptor binding in physiological salt solution. Br. J. Pharmac. 76, 47P. Buckett W. R. (1980) Irreversible inhibitors of GABA transaminase induce antinociceptive effects and potentiate morphine. Neuropharmacology 19, 715-722. Contreras E., Tamayo L. and Quijada L. (1979) Effects of the irreversible inhibition of GABA transaminase upon some morphine effects. Neuropharmacology 18, 309-313. DeFeudis F, V. (1982) GABA-ergic analgesia--A naloxone insensitive system. Pharmac. Res. Commun. 14, 383-390. Eddy M. B., Touchberry C. F. and Lieberman J. E. (1950) Synthetic analgesics: A: Methadone isomers and derivatives. J. Pharmac. exp. Ther. 98, 121-137. Grognet A., Hertz F. and DeFeudis F. V. (1982) Comparison of the analgesic actions of THIP and morphine. Gen. Pharmac. 14, 585-589. Hill R. C., Maurer R., Buescher H. H. and Roemer D. (198l) Analgesic properties of the GABA-mimetic THIP. Eur. J. Pharmac. 69, 221-224. Lloyd H. G. E., Perkins M. N. and Stone T. W. (1982) Ethylenediamine as a specific releasing agent of 7-aminobutyric acid in rat striatal slices. J. Neurochem. 38, 1168-1169. Perkins M. N., Bowery N. G., Hill D. R. and Stone T. W. (1981) Neural responses to etbylenediamine: Preferential blockade by bicuculline. Neurosci. Left. 23, 325-328. Sawynok J. and LaBella F. S. (1982) On the involvement of GABA in the analgesia produced by baclofen, muscimol and morphine. Neuropharmacology 21, 397-403. Winter C. A. and Flataker L. Studies on heptazone (6-morpholino 4,4, diphenyl, 3-heptanone hydrochloride) in comparison with other analgesic drugs. 3'. Pharmac. exp. Ther. 98, 305-317. Yoneda Y., Takashima S. and Kuriyama K. (1976) Possible involvement of GABA in morphine analgesia. Biochem. Pharmac. 25, 2669-2670.
0306-3623/85 $3.00+0.00
Printed in Great Britain
Pergamon Press Ltd
EFFECTS OF ETHYLENEDIAMINE ON MORPHINE ANALGESIA A N D TOLERANCE-DEPENDENCE IN MICE E. CONTRERAS and L. TAMAYO Departamento de Ciencias Fisiolbgicas, Universidad de Concepcibn, Casilla 2407 Concepcirn, Chile (Received 14 January 1985) Abstract--1. Ethylenediamine, a GABA receptor agonist induced a small hyperalgesic state in mice, but increased morphine analgesia. The interaction with this morphine effect was not dose-dependent. 2. Ethylenediamine significantly antagonized tolerance development at relatively low doses (5-10 mg/kg). 3. The GABA mimetic agent increased the frequency of abstinence signs in the naloxone-precipitated morphine withdrawal in mice. 4. The effect of ethylenediamine on morphine withdrawal was suppressed by the irreversible GABA transaminase inhibitor, 7-vinyl GABA.
off the plate or kicking the legs. Each animal was tested twice before drug administration and the values recorded were averaged to obtain a baseline. The effects of ethylenediamine were similarly studied. Reactions times were determined after 30 min for 1 hr 30 min. Ethylenediamine was given i.p. immediately before the administration of the test dose of morphine which was injected s.c. In order to express the total analgesic response the area under the time-response curve was calculated from values obtained every 30 min during the I hr 30 min period. (Winter and Flataker, 1950).
INTRODUCTION
Several lines of investigation have implicated 7 aminobutyric acid ( G A B A ) in the mediation of morphine effects (DeFeudis, 1982). The administration of G A B A agonists, muscimol (Biggio et al., 1977; Sawynok and LaBella, 1982) or its analogue, 4,5,6,7-tetrahydroisoxazolo 5,-4-c pyridin-3-ol (THIP) (Hill et al., 1981; Grognet et aL, 1982) produce analgesia. The inhibitors of G A B A transaminase, aminooxyacetic acid (Yoneda et al., 1976) -vinyl G A B A and ~-acetylenic G A B A (Contreras et aL, 1979) potentiate the antinociceptive action of morphine in mice. In addition Buckett (1980) demonstrated an antinociceptive effect of y-vinyl G A B A . Recently, Perkins et al. (1981) and Bokish et al. (1982) have reported that the electrophoretic administration of ethylenediamine depresses neuronal activity of the central nervous system in various animal species. The drug exerts a displacement of [3H]GABA specifically bound to recognition sites for this neurotransmitter (Bowery et al., 1982) and it promotes the release of [~4C]GABA from rat striatal slices (Lloyd et al., 1982). These results suggest that ethylenediamine may induce changes in the acute and chronic morphine effects. Therefore, it was decided to study the influence of the drug in morphine analgesia and in the induction of tolerance to and physical dependence on the narcotic analgesic.
Induction of tolerance and physical dependence Tolerance and dependence were induced by a slow release preparation of morphine of the following composition: morphine (as free base) 300mg, sorbital sesquioleate 0.80ml, liquid paraffin 4.20ml and 0.9~ NaCI 5.00ml. Each mouse received 300 mg/kg s.c. of morphine. The degree of tolerance was expressed as the ratio of the mean analgesic effect of a test dose of morphine in naive and in primed mice. The antagonism or attenuation of tolerance was determined by the statistical difference between the effects of the test dose in primed untreated animals and those observed in primed treated mice. A level of probability of 0.01 was accepted as significant. This criterion was reinforced by the requirement that a decrease of at least 3 units in the degree of tolerance had to be observed between the values of the treated and the primed untreated groups.
MATERIALS AND METHODS
Adult female mice from a strain raised at our laboratory were maintained under standardized conditions of light (0800--1800 hr), temperature (22 ___2°C) and food. The drugs used were morphine hydrochloride (May and Baker), naloxone hydrochloride (gift from Endo Labs Inc.), ethylenediamine dihydrochloride (L. Light and Co. Ltd) and 7-vinyl GABA (gift from Centre de Recherche Merrel International, Strasbourg, France).
Naloxone precipitated abstinence After 30hr of the slow-release administration of morphine, mice were injected with a dose of naloxone (4 mg/kg) that produced signs of severe abstinence such as diarrhoea, paw tremors, body shakes, and others indicated in the results. The number of mice that showed these signs in a 20min observation period was recorded. The z2-test was used for statistical analysis of the differences between the control (untreated) and treated mice. A level of probability of 0.05 was accepted as significant.
Analgesia Analgesia was assessed by the hot plate method (Eddy et al., 1950) at 55 + 0.5°C. The responses consisted of jumping O.P, 1 6 / ~ 3
529
RESULTS AND DISCUSSION
Ethylenediamine per se exerts a small hyperalgesic state as seen by thermal stimulation in mice; this effect is significant only at the highest dose level tested
530
E. CONTRERAS a n d L. TAMAYO Table 1. Effects of ethylenediamine on morphine analgesia in mice Drug, dose (mg/kg), time after after ethylenediamine administration Saline (control) Morphine, 5 (control)
Analgesic effect ~ 22 __.23 323 _+40 Ethylenediamine alone
Ethylenediamine
20, 30 min 50, 30min 100, 30 min 150, 30 min 200, 30 min
-36 -49 -25 -41 -61
± ± ± ± ±
Ethylenediamine plus morphine (5 mg/kg)
40 51 17 46 142
510 ± 495 ± 521 + 472 ± 547 ±
23 22 36 37 40
P 0.001 0.001 0.005 0.02 0.005
tAssessed by the hot plate procedure in mice and expressed as the area under the time-response curve relating the increase in reaction time and time intervals + SEM. P refers to the statistical significance with reference to the effect of morphine alone. 2Significantly different from saline. (P < 0.005). Number of mice: 14 per group. Table 2. Effect of ethylenediamine on morphine tolerance in mice Analgesic response to a test dose of morphine 5 mg/kg s.c. in mice (hot plate). Area under the time-response curve ± SEM Experimental condition Saline Morphine (untreated) Ethylenediamine 5 mg/kg Ethylenediamine 5 mg/kg twice Ethylenediamine 10 mg/kg Ethylenediamine 20 mg/kg Ethylenediamine 40 mg/kg twice
Degree of tolerance t
Naive
Primed
22 + 23
25 ± 23
323 _ 40
35 ± 41
9.2
340 _ 41
147 ± 473
2.3
326 ± 37
290 ± 613
1.1
330 _ 38
345 ± 523
0.9
568 ± 572
407 ± 433
1.4
486 + 422
406 ± 683
1.2
tRatio of the mean analgesic effect to a test dose in naive and in primed mice. 2Significantly different than the effects of morphine in naive untreated mice. P < 0.01. 3Significantly higher than the effects of morphine in primed untreated mice. P < 0.01. Number of mice: 14 per group. Ethylenediamine was administered 15 rain before the priming dose of morphine, when administering a second dose, this was given 8 hr after the priming dose of the analgesic.
in this work. However, the proposed GABA agonist induces a synergistic effect on the analgesic effect of morphine, sharing the same ability as other GABA mimetics (Biggio et al., 1977; Hill et al., 1981). The synergistic effect of ethylenediamine on morphine analgesia is not dose-related since the different doses assayed exhibited the same potency (Table 1). In tolerant mice, the drug induced a greater increase of analgesia than in the naive mice. In tolerant mice the
effect of the GABA mimetic was initially dosedependent following saturation kinetics (Table 2). With respect to the precipitated morphine withdrawal, ethylenediamine increased the frequency of the abstinence signs when administered either during the course of dependence or 60 min before naloxone (Table 3). The results presented here are in partial agreement with previous observations in drugs affecting GABA
Table 3. Effects of ethylenediamine on the incidence of abstinence signs in morphine treated mice ~
Abstinence sign Jumping Body shakes Paw tremors Abnormal posturing Diarrhoea
Saline
Ethylenediamine during the course of dependence 2
Ethylenediamine during the course of dependence 3
Ethylenediamine 20 mg/kg 1 hr before naloxone
y-vinyl GABA 400 mg/kg 4 hr before and ethylenediamine 20 mg/kg 1 hr before naloxone
6/12 0/12 7/12
8/14 2/14 12/14
8/14 5_5~4 14/14
9/14 ~ 13/14
1~ 3/12 2A]2
8/12 11/ 12
8/14 8/14
10/14 8/14
12/14 7~
0~
qnduced by naloxone, 4 mg/kg i.p., 30 hr after the administration of a sustained release preparation of morphine (300 mg/kg s.c.). 2Ethylenediamine 20 mg/kg administered 15 min before the priming dose of morphine. 3Ethylenediamine 50 mg/kg administered 15 min before the priming dose of morphine. Underlined values are statistically different from those observed in untreated mice (P < 0.05).
531
Ethylenediamine and morphine effects functions. Thus, irreversible GABA transaminase inhibitors increased analgesia, attenuated tolerance and greatly reduced the abstinence behavior in mice (Contreras et al., 1979), whereas in the present experiments ethylenediamine increased the abstinence syndrome. These discrepancies in drugs acting through GABAergic mechanisms may indicate that ethylenediamine exerts a relatively low intrinsic activity at GABA recognition sites in contrast with the GABA transaminase inhibitors which increase the amounts of the inhibitor neurotransmitter in the nervous tissue. The ability of y-vinyl GABA to suppress the enhancing action of ethylenediamine in the abstinence behavior in mice, also supports a possible low intrinsic activity of the GABA receptor agonist. The results of the present work would confirm the involvement of GABA in some morphine effects as it has been suggested by several authors. SUMMARY Recent studies have implicated the inhibitory neurotransmitter y-aminobutyric acid (GABA) in the morphine effects. Ethylenediamine specifically binds to GABA receptors and releases GABA from nervous tissue in the rat, In the present work the influence of ethylenediamine has been studied in morphine analgesia and tolerance to and physical dependence on morphine in mice. Ethylenediamine increased morphine analgesia in the hot plate test in mice but this effect was not dose-related. Low doses of ethylenediamine (5-10 mg/kg) significantly antagonized tolerance development to morphine in mice. The drug increased the incidence of abstinence signs in the naloxone precipitated morphine withdrawal when administered either during the course of dependence or 1 hr before the precipitating dose of naloxone. The effect of ethylenediamine on the naloxone precipitated abstinence was suppressed by the irreversible GABA transaminase inhibitor, y-vinyl GABA. The results of the present paper would support the involvement of GABA in some morphine effects. Acknowledgement--This work was supported by Grant
20.33.13 from Direccirn de Investigacirn, Universidad de Concepcirn. REFERENCES
Biggio D., Della Bella D., Frigeni V. and Guidotti A. (1977) Potentiation of morphine analgesia by muscimol. Neuropharmacology 16, 149-150. Bokisch A. J., Bold J. M., Perkins M, N., Roberts C. J., Stone T. W. and Walker R. J. (1982) Actions of ethylenediamine on Limulus and Helix central neurons and on rat cerebellar and sympathetic ganglion neurons. Br. J. Pharmac. 76, 297P. Bowery N. G., Hill D. R., Hudson A. L., Perkins M. N. and Stone T. W. (1982) GABA receptor binding in physiological salt solution. Br. J. Pharmac. 76, 47P. Buckett W. R. (1980) Irreversible inhibitors of GABA transaminase induce antinociceptive effects and potentiate morphine. Neuropharmacology 19, 715-722. Contreras E., Tamayo L. and Quijada L. (1979) Effects of the irreversible inhibition of GABA transaminase upon some morphine effects. Neuropharmacology 18, 309-313. DeFeudis F, V. (1982) GABA-ergic analgesia--A naloxone insensitive system. Pharmac. Res. Commun. 14, 383-390. Eddy M. B., Touchberry C. F. and Lieberman J. E. (1950) Synthetic analgesics: A: Methadone isomers and derivatives. J. Pharmac. exp. Ther. 98, 121-137. Grognet A., Hertz F. and DeFeudis F. V. (1982) Comparison of the analgesic actions of THIP and morphine. Gen. Pharmac. 14, 585-589. Hill R. C., Maurer R., Buescher H. H. and Roemer D. (198l) Analgesic properties of the GABA-mimetic THIP. Eur. J. Pharmac. 69, 221-224. Lloyd H. G. E., Perkins M. N. and Stone T. W. (1982) Ethylenediamine as a specific releasing agent of 7-aminobutyric acid in rat striatal slices. J. Neurochem. 38, 1168-1169. Perkins M. N., Bowery N. G., Hill D. R. and Stone T. W. (1981) Neural responses to etbylenediamine: Preferential blockade by bicuculline. Neurosci. Left. 23, 325-328. Sawynok J. and LaBella F. S. (1982) On the involvement of GABA in the analgesia produced by baclofen, muscimol and morphine. Neuropharmacology 21, 397-403. Winter C. A. and Flataker L. Studies on heptazone (6-morpholino 4,4, diphenyl, 3-heptanone hydrochloride) in comparison with other analgesic drugs. 3'. Pharmac. exp. Ther. 98, 305-317. Yoneda Y., Takashima S. and Kuriyama K. (1976) Possible involvement of GABA in morphine analgesia. Biochem. Pharmac. 25, 2669-2670.