- Email: [email protected]
Effects of various opiates including specific delta and mu agonists on dopamine release from nigrostriatal dopaminergic neurons in vitro in the rat and in vivo in the cat
Life Sciences, Vol° 31, pp. 2291-2294 Printed in the U.S.A.
Pergamon Pres~
EFFECTS OF VARIOUS OPIATES INCLUDING SPECIFIC DELTA AND MU AGONISTS ON DOPAMINE RELEASE FR~I NIGROSTRIATAL D O P ~ I N E R G I C NEURONS IN VITRO IN THE RAT ~ND IN VIVO IN THE CAT IM.F. Chesselet, A. Cheramy, T.D. Reisine, C. Lubetzki, J. Glowinski, 2M.C. Fournie-Zaluski and B. Roques iGroupe N.B., College de France, Paris, France 2Faculte de Pharmacie, University Descartes, Paris, France
(Received in final form June 14, 1982)
Summary The effect of various opiates on the release of newly synthesized 3H-dopamine (3H.DA) was studied both in vitro in rat striatal slices and in vivo in caudate nuclei and substantiae nigrae of halothane anaesthetized cats, using a push pull cannula technique. The hexapeptide Tyr-D-Ser-Gly-Phe-Leu-Thre (DSTLE) which acts preferentially on S receptors stimulates 3H. DA release in vitro and produces a biphasic increase of the local 3H-amine outflow when applied in the cat caudate nucleus in vivo. Similar effects were induced by D Ala 2met-enkephalinamide. By contrast, morphine, fentanyl and the potent a~onist Tyr-D-AIa-GIy-NH-C6-HI3 (Trimu 4) did not affect 3H.DA release in vitro. In vivo their effects differ from those of the S agonists and from each other: morphine nroduced s delaved long lasting increase in 3H.DA release while fentanyl and Trimu 4 induced an immediate increase of the 3H-amine outflow followed by a delayed inhibition. The local effects of $ and 11 s~onists also differed as re~ard to their sensitivity to naloxone but in ad@itinn, when co-applied with the opiate antagonist, D-Ala2-met-enkephalinamide and morphine induced changes in 3~.DA release in the contralateral csu~ate nucleus and the contralateral or ipsi]stera] substantis nigra respectively. Therefore, in the striatum, some opiates may, under certain circumstances, act directly or indirectlv upon striatal efferences i~vo]ved in the control of the contralateral nigro-striatsl dopaminergic neurons. In conclusion, thanks to the use of recently developed opiate agonists exhibiting a high preference for either $ or , receptors it has been shown that these receptors are involved in different mechanisms regulating the striatal dopaminergic transmission. In addition, these results suggest that less specific agents, bv interacting with several types of receptors, induce a unique combination of events responsible for local and distal control of DA release from nigro-striatal dopaminergic neurons. Recent studies on the characterization and distribution of opiate binding sites have led to the proposal that the D and $ type of opiate receptors described in the periphery have their counterpart in the mammalian brain (1,2,3, 4). Interestingly, in a given species the respective density of each type of opiate sites varies from one area to another (5,6) and furthermore, autoradiographic studies have shown in such a highly organized structure as the cortex that the ~ and N bindin~ sites are located in different layers (7). These data strongly suggest that each receptor type is involved in different func0024-3205/82/202291-04503.00/0 Copyright (c) 1982 Pergamon Press Ltd.
2292
~ and 6 Opiates and DA Release
Vol. 31, No.s 20 & 21, 1982
tions. Although it is well established that at least some p receptors play a role in the control of pain perception (3,8), much less is known about the physiological responses following B receptor stimulation. According to binding studies, the striatum, a structure innervated by a dense network of dopaminergic terminals originating from the substantia nigra, contains one of the highest density of both ~ and S opiate receptors (5,6) in the brain. When injected at the periphery or into the ventricle, p as well as $ opiate agonists modulate striatal dopaminergic transmission (9,10). However, the site and the mechanism of this interaction remains uncertain. In fact, although several studies suggest that opiates increased dopamine (DA) turnover by acting at intrastriatal opiate sites, (11,12), morphine was unable to stimulate the release of dopamine in rat striatal slices (13) One possible explanation for these discrepancies could be that several mechanisms involving different types of opiate receptors intervene in complex opiate actions on nigrostriatal dopaminergic neurons. This led us to investigate the effects of preferential S and ~ opiate agonists on the release of dopamine from nigrostriatal dopaminergic nerve terminals when locally applied in the striatum. For this purpose, the actions of several new compounds showing a high selectivity for either 8 or p opiate sites was analyzed and compared to those of more classical opiate agonists in two types of experiments:in vitro, in superfused rat striatal slices (14) and in vivo in halothane anaesthetized cats in which both caudate nuclei (CN) and substantiae nigrae (SN) were perfused by means of stereotoxically placed push pull cannulae (15). In both cases, the tissue was superfused with a medium containing L-3-5-3H-tyrosine (50 Ci/mmol, 50 pCi/ml) and the release of newly formed 3H-DA was estimated in serial superfusate fractions after biochemical analysis (15). In vitro experiments. A clear difference was observed in the effects of compounds known to act preferentially on S or p opiate receptors in rat striatal slices (16,17). D-Ala2-D-Leu5-enkephalin (DADL) which acts preferentially on S than on p receptors (18,19) increased the spontaneous, calcium dependent, release of newly synthetized 3H-dopam%ne when added to the superfusion fluid at a concentration as low as 5 x 10-9M. The involvement of S receptors in this effect is strongly supported by the fact that it can be similarly induced by Try-DSer-Gly-Phe-LeuThr, DSTLE (5 x I0-8,10-7M), a peptide which highly discriminates between $ versus D sites in peripheral tissue (20,21) as well as in the brain (4). Moreover, DSTLE's effect on 3H-DA release can be blocked by naloxone only if the concentration of the antagonist exceeds that of the agonist. For example, the increase in 3H-DA release induced by DSTLE 5 x IO-8M was blocked by naloxone IO-7M but this concentration of naloxone does not prevent the effect of DSTLE 10-7M. These results reveal that the receptors involved in DSTLE's action have a low sensitivity to naloxone, which is a characteristic feature of ~opiate receptors (1,2)19). D-Ala2-met-enkephalinamide (D-Ala2-met-enk,10-6M), an analog of the endogenous compound met-enkephalin resistant to enkephalin degradation, also increased 3H-DA release in rat striatal slices without changing the amine synthesis. According to the previously discussed data, the increase in 3H-DA release induced by D-Ala2-met-enk is most probably due also to its action on 8 opiate sites. In fact, none of the preferential p agonists tested could produce any effect on 3H-DA release in rat striatal slices, even at relatively hig~ concentration. This was the case for morphine (5 x IO-6M), fentanyl (5 x i0- M) and the tripeptide Tyr-D-AIa-GIy-NH-C 6 HI3 (Trimu 4, IO-6M) which exhibits a marked preferential affinity for p opiate receptors (8,21) and binding sites (4). These results suggest that B opiate receptors may be involved in a local, presynaptic regulation of dopaminergic transmission by some striatal endogenous opiates. Whether this interaction is direct or indirect cannot be interpreted from our experiments, although the possibility of a direct interaction is suggested by several facts. First, we have shown that the D-Ala2-met-enk's effect on 3H-DA release persisted when nerve activity was blocked by tetrodotoxin (5 x i0-711)(17). Secondly, in autoradio~raphic
Vol. 31, No.s 20 & 21, 1982
~ and 6 Opiates and DA Release
2293
experiments, preferential 8 ligands labelled receptors having a diffuse distribution in the striatum (7). Some of them could be associated with dopaminergic nerve terminals since 6-hydroxydopamine induced lesions of dopaminergic neurons led to a decrease in the number of striatal opiate sites in binding experiments (22,23) and also to a marked reduction of the diffusely located opiate sites in the autoradiographic studies (24). In vivo experiments. Experiments performed in the cat confirmed that preferential 8 and ~ agonists modulated dopaminergic transmission in a different manner. In fact, when locally applied in one cat CN for 30 min, D-Ala2-met-enk ~I0-6M) as well as DSTLE (5 x 10-8M) induced a marked immediate increase in JH-DA release (25). The initial increase (first i0 min fraction of application) was followed by a return of 3H-DA release to control levels and by a second increase when the opiate was removed from the superfusion medium. The mechanism of this biphasic increase is unknown but it should be noted that the amplitude of the two peaks is highly correlated in each experiment. Naloxone (10-6M) did not significantly reduce the amplitude of the 3H-DA increase produced by either D-AIa 2met-enk (10-6M)or DSTLE (5 x 10-8M) (Chesselet et al., submitted). The results were quite different when the preferential ~ agonists which were ineffective in vitro in changing 3H-DA release in the concentration tested, were applied in the cat CN. In fact, in vivo those agents modified 3H-DA release but in a way very different from what was described with the preferential S agonists. Morphine (10-6M) induced a delayed, moderate and prolonged increase in 3H-DA release which was completely reversed by naloxone (i0- M) (25). Trimu4(10-6M) also induced a delayed slight increase in the 3H amine release but this effect was transient and since these experiments were prolonged for up to 70 min after the removal of the peptide from the superfusion medium, a progressive decrease in 3H-DA release was observed. This effect lasted from 30 to 70 min after the end of the peptide's application. Fentanyl, a compound considered to be a potent morphinomimetic, produced a more complex effect that consisted of an initial biphasic increase in 3H-DA release followed by a marked inhibition. However, the biphasic increase was clearly different from the effect produced by 8 agonists since the two peaks are not positively correlated and the second peak as well as the following inhibition were completely blocked by naloxone. Thus the analysis of the local effects of various opiates applied in tbe cat CN on 3H-DA release confirm that agents acting on S and ~ receptors affect dopaminergic transmission through different mechanisms. However, differences also appeared among various preferential ~ agonists, showing that the agents tested cannot be simply classified in two pharmacological categories. This appears even more clearly when considering the effects produced by these opiates in the contralateral CN and both SN. DSTLE and fentanyl which have much preferential affinity for $ and N receptors respectively do not produce any change in 3H-DA release in the distal structures studied either when applied alone or in the presence of naloxone. By contrast, compounds having a wider spectrum of action, i.e. which may act to different degrees on 2, ~ and other opiate sites depending on the concentration utilized (19) had more complex effects on 3H-DA release. D-Ala2-met-enk (10-6M) induced only a slight inhibition in 3H-DA release in the substantia nigra when applied alone into one CN but, in the presence of naloxone (IO-6M), it produced an increased release in the contralateral CN and SN while the decrease in the ipsilateral nigra was reversed. Thus the presence of naloxone unmasked distal stimulating effects of D-Ala2-met-enk whereas the antagonist itself only produced a slight and delayed inhibition of 3H-DA release in both CN. A puzzling alteration of the opiate effects while in the presence of naloxone was also observed for morphine: morphine itself had no significant effect on 3H-DA efflux in the distal structures studied but, in the presence of naloxone, an inhibition of 3H-DA release was observed locally, in the ipsilateral nigra and in the contralateral CN when compared to the values of 3H-DA release in the control or in the morphine experiments. Thus, at least in three structures, the changes
2294
D and ~ Opiates and DA Release
Vol. 31, No.s 20 & 21, 1982
induced by morphine and D-Ala2-met-enk in the presence of naloxone are in opposite directions. It is known that naloxone has different affinities for different classes of opiate receptors (1,19). Therefore one hypothesis to explain these paradoxical results may be that naloxone by blocking some actions of the opiate studied revealed effects resulting from their actions on unblocked sites, which are normally counteracted or masked by effects produced at naloxone-sensitive sites. The anatomical pathways which may be involved in these distal effects will be discussed elsewhere, but a few remarks can be made concerning the possible signification of these results. First, as already suspected from the diversity of previous experimental results, the interaction between opiates and striatal dopaminergic transmission is certainly complex. Part of this interaction seems to occur in the striatum, through an indirect or more probably direct action of opiates on dopaminergic nerve terminals. The present results suggest that this interaction involves, at least partially, 8 opiate sites, providing one of the first evidences for a functional role of this class of opiate receptors in the mammalian brain. Secondly, because of their lack of specificity for one particular receptor, most opiates induce a unique pattern of events resulting from their particular spectrum of action. This strongly suggest that, in the brain, different classes of opiate receptors are involved in different opiate biological responses and are therefore of physiological relevance. i. J.A.H. LORD, A.A. WATERFIELD, J. HUGHES and H.W. KOSTERLITZ, Nature 267 495-499 (1977) 2. K.J. CHANG and P. CUATRECASAS, J. Biol. Chem. 254 2610-2618 (1979) 3. H.W. KOSTERLITZ, J.A.H. LORD, S.J. PATERSON and A.A. WATERFIELD, Br. J. Pharmacol. 68 333-342 (1980) 4. B.P. ROQUES, M.C. FOURNIE-ZALUSKI, G. GACEL, M. DAVID, J.C. MEUNTER, B. MAIGRET and J.L. MORGAT, Regulatory Peptides, Functional and Pharmacological Aspects, E. Costa and M. Trabucchi, eds., in press,Raven Press, NY 5. K.J. CHANG, B.R. COOPER, E. HAZb~ and P. CUATRECASAS, Molec. Pharmacol. 16 91-104 (1979) 6. M.NINCOVIC,S.P.HUNT,P.C.EMSON and L.L.IVERSEN,Brain Res.214 163-167 (1981) 7. R.R. GOODMAN, S.H. SNYDER, M.J. KUHAR and W.S. YOUNG, Proc. Nat. Acad. Sci. USA 77 6239-6243 (1980) 8. G. GACEL, M.C. FOURNIE-ZALUSKI, E. FELLION and B.P. ROQUES, J. Med. Chem. 24 1119-1124 (1981) 9. B.H.C. WESTERINK and J. KORF, Eur. J. Pharmacol. 38 281-291 (1976) i0. P.L. WOOD, M. STOTLAND, J.W. RICHARD and A. RACKH~M, J. Pharmacol. Exp. Ther. 215 697-703 (1980) II. G.BIGGIO,M.CASA,M.CORDA,C.DIBELLO and G.L.GESSA,Science 200 552-554 (1978) 12. P.MOLEMAN and J.BRUINVELS, Nature 281 686-687 (1979) 13. S. ARBILLA and S.Z. LANGER, Nature 271 559-568 (1976) 14. M.F. GIORGUIEFF, M.L. LE FLOCH, J. GLOWINSKI and M.J. BESSON, J. Pharmacol. Exp. Ther. 200 535-544 (1977) 15. V. LEVIEL, M.F. CHESSELET, J. GLOWINSKI and A. CHERAMY, Brain Res. 223 257-272 (1981) 16. M.F. CHESSELET, C. LUBETZKI, A. CHERAMY, T.D. REISINE and J. GLOWINSKI, Presynaptic Receptors, J. DeBelleroche ed., in press Ellis Howood pub. 17. C.LUBETZKI,M.F.CHESSELET and J.GLOWINSKI,J.Pharmacol. Exp. Ther. in press 18. M. WUSTER, R. SCHULZ and A. HERZ, Neurosci. Lett. 15 193-198 (1979) 19. H.W.KOSTERLITZ and S.J.PATERSON, Proc. R.Soc.Lond. 210 113-122 (1980) 20. G.GACEL, M.C.FOURNIE-ZALUSKI and B.ROQUES, FEBS Lett. 118 245-247 (1980) 21. M.C. FOURNIE-ZALUSKI, G. GACEL, B. MAIGRET, S. PREMILAT and B.P. ROQUES, Molec. Pharmacol. 20 484-491 (1981) 22. H. POLLARD, C. LLORENS and J.C. SCHWARTZ Nature 268 745-747 (1977) 23. T.D. REISINE, J.I. NAGY, K. BEAUMONT, H.C. FIBIGER and H.I. YAMAMURA, Brain Res. 177 241-252 (1979) 24. L.C. MURRIN, J.T. COYLE and M.J. KUHAR, Life Sci. 27 1175-1183 (1980) 25. M.F.CHESSELET,A.CHERAM~,T.D.REISINE and J.GLOWINSKI,Nature 291 320-322(1981)
Pergamon Pres~
EFFECTS OF VARIOUS OPIATES INCLUDING SPECIFIC DELTA AND MU AGONISTS ON DOPAMINE RELEASE FR~I NIGROSTRIATAL D O P ~ I N E R G I C NEURONS IN VITRO IN THE RAT ~ND IN VIVO IN THE CAT IM.F. Chesselet, A. Cheramy, T.D. Reisine, C. Lubetzki, J. Glowinski, 2M.C. Fournie-Zaluski and B. Roques iGroupe N.B., College de France, Paris, France 2Faculte de Pharmacie, University Descartes, Paris, France
(Received in final form June 14, 1982)
Summary The effect of various opiates on the release of newly synthesized 3H-dopamine (3H.DA) was studied both in vitro in rat striatal slices and in vivo in caudate nuclei and substantiae nigrae of halothane anaesthetized cats, using a push pull cannula technique. The hexapeptide Tyr-D-Ser-Gly-Phe-Leu-Thre (DSTLE) which acts preferentially on S receptors stimulates 3H. DA release in vitro and produces a biphasic increase of the local 3H-amine outflow when applied in the cat caudate nucleus in vivo. Similar effects were induced by D Ala 2met-enkephalinamide. By contrast, morphine, fentanyl and the potent a~onist Tyr-D-AIa-GIy-NH-C6-HI3 (Trimu 4) did not affect 3H.DA release in vitro. In vivo their effects differ from those of the S agonists and from each other: morphine nroduced s delaved long lasting increase in 3H.DA release while fentanyl and Trimu 4 induced an immediate increase of the 3H-amine outflow followed by a delayed inhibition. The local effects of $ and 11 s~onists also differed as re~ard to their sensitivity to naloxone but in ad@itinn, when co-applied with the opiate antagonist, D-Ala2-met-enkephalinamide and morphine induced changes in 3~.DA release in the contralateral csu~ate nucleus and the contralateral or ipsi]stera] substantis nigra respectively. Therefore, in the striatum, some opiates may, under certain circumstances, act directly or indirectlv upon striatal efferences i~vo]ved in the control of the contralateral nigro-striatsl dopaminergic neurons. In conclusion, thanks to the use of recently developed opiate agonists exhibiting a high preference for either $ or , receptors it has been shown that these receptors are involved in different mechanisms regulating the striatal dopaminergic transmission. In addition, these results suggest that less specific agents, bv interacting with several types of receptors, induce a unique combination of events responsible for local and distal control of DA release from nigro-striatal dopaminergic neurons. Recent studies on the characterization and distribution of opiate binding sites have led to the proposal that the D and $ type of opiate receptors described in the periphery have their counterpart in the mammalian brain (1,2,3, 4). Interestingly, in a given species the respective density of each type of opiate sites varies from one area to another (5,6) and furthermore, autoradiographic studies have shown in such a highly organized structure as the cortex that the ~ and N bindin~ sites are located in different layers (7). These data strongly suggest that each receptor type is involved in different func0024-3205/82/202291-04503.00/0 Copyright (c) 1982 Pergamon Press Ltd.
2292
~ and 6 Opiates and DA Release
Vol. 31, No.s 20 & 21, 1982
tions. Although it is well established that at least some p receptors play a role in the control of pain perception (3,8), much less is known about the physiological responses following B receptor stimulation. According to binding studies, the striatum, a structure innervated by a dense network of dopaminergic terminals originating from the substantia nigra, contains one of the highest density of both ~ and S opiate receptors (5,6) in the brain. When injected at the periphery or into the ventricle, p as well as $ opiate agonists modulate striatal dopaminergic transmission (9,10). However, the site and the mechanism of this interaction remains uncertain. In fact, although several studies suggest that opiates increased dopamine (DA) turnover by acting at intrastriatal opiate sites, (11,12), morphine was unable to stimulate the release of dopamine in rat striatal slices (13) One possible explanation for these discrepancies could be that several mechanisms involving different types of opiate receptors intervene in complex opiate actions on nigrostriatal dopaminergic neurons. This led us to investigate the effects of preferential S and ~ opiate agonists on the release of dopamine from nigrostriatal dopaminergic nerve terminals when locally applied in the striatum. For this purpose, the actions of several new compounds showing a high selectivity for either 8 or p opiate sites was analyzed and compared to those of more classical opiate agonists in two types of experiments:in vitro, in superfused rat striatal slices (14) and in vivo in halothane anaesthetized cats in which both caudate nuclei (CN) and substantiae nigrae (SN) were perfused by means of stereotoxically placed push pull cannulae (15). In both cases, the tissue was superfused with a medium containing L-3-5-3H-tyrosine (50 Ci/mmol, 50 pCi/ml) and the release of newly formed 3H-DA was estimated in serial superfusate fractions after biochemical analysis (15). In vitro experiments. A clear difference was observed in the effects of compounds known to act preferentially on S or p opiate receptors in rat striatal slices (16,17). D-Ala2-D-Leu5-enkephalin (DADL) which acts preferentially on S than on p receptors (18,19) increased the spontaneous, calcium dependent, release of newly synthetized 3H-dopam%ne when added to the superfusion fluid at a concentration as low as 5 x 10-9M. The involvement of S receptors in this effect is strongly supported by the fact that it can be similarly induced by Try-DSer-Gly-Phe-LeuThr, DSTLE (5 x I0-8,10-7M), a peptide which highly discriminates between $ versus D sites in peripheral tissue (20,21) as well as in the brain (4). Moreover, DSTLE's effect on 3H-DA release can be blocked by naloxone only if the concentration of the antagonist exceeds that of the agonist. For example, the increase in 3H-DA release induced by DSTLE 5 x IO-8M was blocked by naloxone IO-7M but this concentration of naloxone does not prevent the effect of DSTLE 10-7M. These results reveal that the receptors involved in DSTLE's action have a low sensitivity to naloxone, which is a characteristic feature of ~opiate receptors (1,2)19). D-Ala2-met-enkephalinamide (D-Ala2-met-enk,10-6M), an analog of the endogenous compound met-enkephalin resistant to enkephalin degradation, also increased 3H-DA release in rat striatal slices without changing the amine synthesis. According to the previously discussed data, the increase in 3H-DA release induced by D-Ala2-met-enk is most probably due also to its action on 8 opiate sites. In fact, none of the preferential p agonists tested could produce any effect on 3H-DA release in rat striatal slices, even at relatively hig~ concentration. This was the case for morphine (5 x IO-6M), fentanyl (5 x i0- M) and the tripeptide Tyr-D-AIa-GIy-NH-C 6 HI3 (Trimu 4, IO-6M) which exhibits a marked preferential affinity for p opiate receptors (8,21) and binding sites (4). These results suggest that B opiate receptors may be involved in a local, presynaptic regulation of dopaminergic transmission by some striatal endogenous opiates. Whether this interaction is direct or indirect cannot be interpreted from our experiments, although the possibility of a direct interaction is suggested by several facts. First, we have shown that the D-Ala2-met-enk's effect on 3H-DA release persisted when nerve activity was blocked by tetrodotoxin (5 x i0-711)(17). Secondly, in autoradio~raphic
Vol. 31, No.s 20 & 21, 1982
~ and 6 Opiates and DA Release
2293
experiments, preferential 8 ligands labelled receptors having a diffuse distribution in the striatum (7). Some of them could be associated with dopaminergic nerve terminals since 6-hydroxydopamine induced lesions of dopaminergic neurons led to a decrease in the number of striatal opiate sites in binding experiments (22,23) and also to a marked reduction of the diffusely located opiate sites in the autoradiographic studies (24). In vivo experiments. Experiments performed in the cat confirmed that preferential 8 and ~ agonists modulated dopaminergic transmission in a different manner. In fact, when locally applied in one cat CN for 30 min, D-Ala2-met-enk ~I0-6M) as well as DSTLE (5 x 10-8M) induced a marked immediate increase in JH-DA release (25). The initial increase (first i0 min fraction of application) was followed by a return of 3H-DA release to control levels and by a second increase when the opiate was removed from the superfusion medium. The mechanism of this biphasic increase is unknown but it should be noted that the amplitude of the two peaks is highly correlated in each experiment. Naloxone (10-6M) did not significantly reduce the amplitude of the 3H-DA increase produced by either D-AIa 2met-enk (10-6M)or DSTLE (5 x 10-8M) (Chesselet et al., submitted). The results were quite different when the preferential ~ agonists which were ineffective in vitro in changing 3H-DA release in the concentration tested, were applied in the cat CN. In fact, in vivo those agents modified 3H-DA release but in a way very different from what was described with the preferential S agonists. Morphine (10-6M) induced a delayed, moderate and prolonged increase in 3H-DA release which was completely reversed by naloxone (i0- M) (25). Trimu4(10-6M) also induced a delayed slight increase in the 3H amine release but this effect was transient and since these experiments were prolonged for up to 70 min after the removal of the peptide from the superfusion medium, a progressive decrease in 3H-DA release was observed. This effect lasted from 30 to 70 min after the end of the peptide's application. Fentanyl, a compound considered to be a potent morphinomimetic, produced a more complex effect that consisted of an initial biphasic increase in 3H-DA release followed by a marked inhibition. However, the biphasic increase was clearly different from the effect produced by 8 agonists since the two peaks are not positively correlated and the second peak as well as the following inhibition were completely blocked by naloxone. Thus the analysis of the local effects of various opiates applied in tbe cat CN on 3H-DA release confirm that agents acting on S and ~ receptors affect dopaminergic transmission through different mechanisms. However, differences also appeared among various preferential ~ agonists, showing that the agents tested cannot be simply classified in two pharmacological categories. This appears even more clearly when considering the effects produced by these opiates in the contralateral CN and both SN. DSTLE and fentanyl which have much preferential affinity for $ and N receptors respectively do not produce any change in 3H-DA release in the distal structures studied either when applied alone or in the presence of naloxone. By contrast, compounds having a wider spectrum of action, i.e. which may act to different degrees on 2, ~ and other opiate sites depending on the concentration utilized (19) had more complex effects on 3H-DA release. D-Ala2-met-enk (10-6M) induced only a slight inhibition in 3H-DA release in the substantia nigra when applied alone into one CN but, in the presence of naloxone (IO-6M), it produced an increased release in the contralateral CN and SN while the decrease in the ipsilateral nigra was reversed. Thus the presence of naloxone unmasked distal stimulating effects of D-Ala2-met-enk whereas the antagonist itself only produced a slight and delayed inhibition of 3H-DA release in both CN. A puzzling alteration of the opiate effects while in the presence of naloxone was also observed for morphine: morphine itself had no significant effect on 3H-DA efflux in the distal structures studied but, in the presence of naloxone, an inhibition of 3H-DA release was observed locally, in the ipsilateral nigra and in the contralateral CN when compared to the values of 3H-DA release in the control or in the morphine experiments. Thus, at least in three structures, the changes
2294
D and ~ Opiates and DA Release
Vol. 31, No.s 20 & 21, 1982
induced by morphine and D-Ala2-met-enk in the presence of naloxone are in opposite directions. It is known that naloxone has different affinities for different classes of opiate receptors (1,19). Therefore one hypothesis to explain these paradoxical results may be that naloxone by blocking some actions of the opiate studied revealed effects resulting from their actions on unblocked sites, which are normally counteracted or masked by effects produced at naloxone-sensitive sites. The anatomical pathways which may be involved in these distal effects will be discussed elsewhere, but a few remarks can be made concerning the possible signification of these results. First, as already suspected from the diversity of previous experimental results, the interaction between opiates and striatal dopaminergic transmission is certainly complex. Part of this interaction seems to occur in the striatum, through an indirect or more probably direct action of opiates on dopaminergic nerve terminals. The present results suggest that this interaction involves, at least partially, 8 opiate sites, providing one of the first evidences for a functional role of this class of opiate receptors in the mammalian brain. Secondly, because of their lack of specificity for one particular receptor, most opiates induce a unique pattern of events resulting from their particular spectrum of action. This strongly suggest that, in the brain, different classes of opiate receptors are involved in different opiate biological responses and are therefore of physiological relevance. i. J.A.H. LORD, A.A. WATERFIELD, J. HUGHES and H.W. KOSTERLITZ, Nature 267 495-499 (1977) 2. K.J. CHANG and P. CUATRECASAS, J. Biol. Chem. 254 2610-2618 (1979) 3. H.W. KOSTERLITZ, J.A.H. LORD, S.J. PATERSON and A.A. WATERFIELD, Br. J. Pharmacol. 68 333-342 (1980) 4. B.P. ROQUES, M.C. FOURNIE-ZALUSKI, G. GACEL, M. DAVID, J.C. MEUNTER, B. MAIGRET and J.L. MORGAT, Regulatory Peptides, Functional and Pharmacological Aspects, E. Costa and M. Trabucchi, eds., in press,Raven Press, NY 5. K.J. CHANG, B.R. COOPER, E. HAZb~ and P. CUATRECASAS, Molec. Pharmacol. 16 91-104 (1979) 6. M.NINCOVIC,S.P.HUNT,P.C.EMSON and L.L.IVERSEN,Brain Res.214 163-167 (1981) 7. R.R. GOODMAN, S.H. SNYDER, M.J. KUHAR and W.S. YOUNG, Proc. Nat. Acad. Sci. USA 77 6239-6243 (1980) 8. G. GACEL, M.C. FOURNIE-ZALUSKI, E. FELLION and B.P. ROQUES, J. Med. Chem. 24 1119-1124 (1981) 9. B.H.C. WESTERINK and J. KORF, Eur. J. Pharmacol. 38 281-291 (1976) i0. P.L. WOOD, M. STOTLAND, J.W. RICHARD and A. RACKH~M, J. Pharmacol. Exp. Ther. 215 697-703 (1980) II. G.BIGGIO,M.CASA,M.CORDA,C.DIBELLO and G.L.GESSA,Science 200 552-554 (1978) 12. P.MOLEMAN and J.BRUINVELS, Nature 281 686-687 (1979) 13. S. ARBILLA and S.Z. LANGER, Nature 271 559-568 (1976) 14. M.F. GIORGUIEFF, M.L. LE FLOCH, J. GLOWINSKI and M.J. BESSON, J. Pharmacol. Exp. Ther. 200 535-544 (1977) 15. V. LEVIEL, M.F. CHESSELET, J. GLOWINSKI and A. CHERAMY, Brain Res. 223 257-272 (1981) 16. M.F. CHESSELET, C. LUBETZKI, A. CHERAMY, T.D. REISINE and J. GLOWINSKI, Presynaptic Receptors, J. DeBelleroche ed., in press Ellis Howood pub. 17. C.LUBETZKI,M.F.CHESSELET and J.GLOWINSKI,J.Pharmacol. Exp. Ther. in press 18. M. WUSTER, R. SCHULZ and A. HERZ, Neurosci. Lett. 15 193-198 (1979) 19. H.W.KOSTERLITZ and S.J.PATERSON, Proc. R.Soc.Lond. 210 113-122 (1980) 20. G.GACEL, M.C.FOURNIE-ZALUSKI and B.ROQUES, FEBS Lett. 118 245-247 (1980) 21. M.C. FOURNIE-ZALUSKI, G. GACEL, B. MAIGRET, S. PREMILAT and B.P. ROQUES, Molec. Pharmacol. 20 484-491 (1981) 22. H. POLLARD, C. LLORENS and J.C. SCHWARTZ Nature 268 745-747 (1977) 23. T.D. REISINE, J.I. NAGY, K. BEAUMONT, H.C. FIBIGER and H.I. YAMAMURA, Brain Res. 177 241-252 (1979) 24. L.C. MURRIN, J.T. COYLE and M.J. KUHAR, Life Sci. 27 1175-1183 (1980) 25. M.F.CHESSELET,A.CHERAM~,T.D.REISINE and J.GLOWINSKI,Nature 291 320-322(1981)