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Opiate-receptors in substantia nigra: Role in the regulation of striatal tyrosine hydroxylase activity
PRELIMINARY NOTES OPIATE-RECEPTQRS TN SUBSTANTIA NIGRA: ROLE IN TAE REGULA'FION Ol? STR32iTAL TYRQSINE HYDROXYLASE ACTIVl'TY
K.
Gale*,
F.
Moronir
M.
Kumakura
and
A, Guidotti
Laboratory of PreclinicalPharmacology,National fnstitute of Mental Health, Saint Elizabeths Hospital, Washington,D.C. 20032 *Departmentof Pharmacology,Georgetown University, Washington, D.C, 20(107
Morphine injected into the substantianiqxa fSN) of rats failed to change DA metabolism in striatusn but blocked the haloperidol induced activation of striatal tyrosine hydroxylase (TX). This blockade was mediated by stimulationof opiate recept3rs in SN because it was antagoni2xXi by naltrexone, The binding of H-enkephalinamideto opiate receptors in SN was not altered after lesion of the niqrostriatalDA pathways with 6hydroxydopaminebut was decreased (approximately30%) after brain hemitransection. These results sugqest that intraniqralmorphine prevents the haloperidol induced activation of striatal TfIby modulating the release of endogenous transmittersor modulators on DA neurons. Afferent projections to substantia nigra (SN) include pathways Containing GABA, substance P and serotonin. These transmittershave been shown to affect both efectrophysioloqicaland neurochemical. function of efferent pathways from SN, including the nigrostriataldopamine (DA) projection ffor a review see Dray and Straughan, 19761. Recently, met-enkephalinhas been maasured ih SN (Yang et al., X978), and although the concentrationof this peptide in SB is not as high as that found in other nuclei of the basal ganglia, opiate receptors a~@ abundant in SN (Pollardet al., 19781. This suggests that endogenous opiate peptides may represent an important neurotransmittersystem in the regulationof the activity of SN neurons. At present, the only evideaee in the literaturewhich suggests a functional role for the opiate'receptarsin SN is the observation that Unilateral intraniqxal injection of morphine or beta-endorphininduces contralateralcircling behavior which can be reversed by naloxone (Iwamotoand Way, 1978). In order tm determine whether activation of these opiate receptors in 5% could have functional importance for the regulation of nigrostriatalDA neurons, we chose to study the effect of intranigrallyapplied morphine on the tyrosine hydroxylase ITHI activity in the DA terminals of striatum and to characterize the neuronal population of SN where the opiate receptors are located. After systemic or intrastriatalinjectionsof morphineI an increase in DA metabolism was found in rat striatum. However, 30 min. after injection of morphine directly into SN (in doses of 1,2,5,10 and 20 Ug of the f.90) salt, dissolved in 0,s ~1 saline) we found no change in striatal IF.TA content (Guidottiet al., 1979). These results suggest that unlike the opiate receptors in striatum, the opiate receptors in SN do not mediate the morphine-inducedstimulation of strkatal RA metabolismIn order to determine whether activation of opiate receptors in .W night exert an inhibitory effect on DA neuronsI we examined whether intrraniqraf morphine could inhibit the activation of striatal TN.induced by haloperidol. The action of haluperidolon striatal TH is mediated via the transsynaptisregulation of DA neurons by niqral afferents, including the striate-dgral axon8 containing GABA or substance P (Gale et al., 1978). We therefore expected that if the nigxal opiate receptors exert an inhibitory influence on DA neurons (eitherdirectly ox indirectly),then intraniqralmorphine would be able to antagonize the activation of striatal TN elicited by haloperidol, AS shown in Fig, 1, although fntranigralmorphine by itself did not change the affinity of striatal 9z-X for nMPa , it completely prevented the increase in affinity of striatal Ttrfor DMPE4 in animals trentttdw$th haloperifiol.This effect of intranigralmorphine was reversed by systemic (Fig+ I) or intranigral. injection (Table 1) of the opiate-receptoxantagonist, naltrexone, but was nat reversed by intranigral injection of bicuculline methiodide (Table 1) a EABA receptor antagonist. The specificityof the action of morphine was further sup&orted by experiments which demonstrated that the ability of muscimol to antagonize the haloperidol-induced
427
428
Preliminary Notes
activation of striatal TH was reversed by intranigral bicuculline methiodide but not by naltrexone (Table 1).
htranigral morpthe blockade
ofHabperii-inducedktivatii
of Striatal TH
l
Mmpkm
\lOvg intronigrd)
0.37 0 HdopriQ((lmg/kg i.P.1
Figure 1.
DOUBLE RECIPROCAL PMTS OF THE INITIAL VELOCITY OFl$RIATAL TH AGAINST VARIOUS CONCENTRATIONS OF DNPH4 IN THE PRESENCE OF 0.1 ml4 C-TYROSINB. TH activity was determined using a dopadecarboxylasecoupled enzymatic reaction to measure 14 CO2 formed from carboxyl labelled tyrosine according to the method of Eivkovic et al. (1974). Haloperidol was injected 30 min prior sacrifice, morphine sulfate 10 min before haloperidol and naltrexone 10 min after haloperidol. Each point represent the mean obtained from six animals. The standard error of the Km values was between 5 and 10%. Stereotaxic coordinates for the intranigral injections were the same as those used by Gale and Guidotti (1976). The volume The Km for TH of untreated rats was 0.83 mM. injected into SN was 0.5 W. Table 1.
Relationship Between Opiate Receptors and GABA Receptors in SN for The Control of Haloperidol-InducedActivation of Striatal TH A Intravenous Injection
B First Intranigral Injection
C Second Intranigral Injection Saline (Km of
Bicuculline Naltrexone TH for DNPH4; mM)
Striatal
SALINE
Saline Muscimol Morphine
.85 .80 .83
.83 .88 .87
.79 .87 .85
HALOPERIDOL
Saline Muscimol Morphine
.37* .85 .73
.32* .91 .30*
.42* .40* .78
A) Saline or haloperidol (0.3 umoles/kg) was administered 30 min prior to sacrifice. B) Saline (0.5 W), morphine sulfate (10 llgin 0.5 ~1) or muscimol (100 ng in 0.5 ~1) was injected 10 min before haloperidol; C) Saline (0.5 ~11, naltrexone (10 vg in 0.5 ~1) or bicuculline methiodide (500 ng in 0.5 ~1) was injected 10 min after haloperidol. TH measurements were done as described in the legend of Fig. 1. Values represent the mean of 6-8 animals. Standard errors were less than 15% of the mean. *Significantlydifferent (P<.Ol) from saline (systemic+ intranigral treated controls. These results suggest that activation of opiate receptors in SN inhibits the activity of DA neurons. In order to determine whether this effect could be mediated by a direct action of morphine on DA cells in SN, we measured opiate receptor binding in SN after unilateral destruction of nigrostriatal DA neurons by 6-hydroxydopamine (6-OHDA). Although 6-OHDA treatment resulted in the destruction of more than 85% of the nigrostriatal DA3neurons (based on TH measurements), no significant change was obtained in the binding of H enkephalinamide in the
Preliminary Notes
crude ~2 fraction of the lesioned SN (Table 2). In contrast, there was a marked (68%) decrease in the binding of 3H enkephalinamide to SN P2 preparation from rats with a mechanical hemitransection of all the connections between the SN and striatum (Table 2). This lesion reduced both GABA and substance P content in SN by 60-85% (Gale et al., 1978). Table 2.
3 H-EnkephalinamideBinding in Rat SN After C-OHDA or Hemitransection 3H-Enkephalinamide(fmol/mgprot) Lesion
Intact Side
6-OHDA
105 zt12
95
Hemitransection
110 t
35 f
8
Lesioned Side
i
% Decrease Due to Lesion
15 6*
68
Bats (250 g) were anesthetized and 6-OHDA (8 ug in 4 ~1) was injected into SN at the rate of .25 ul/min. Stereotaxic coordinates were the same as those used for intranigral injections in Fig. 1. Animals were sacrificed 2 weeks after 6-OHDA treatment. Measurement of TH demonstrated a decrease of 85-98%. Hemitransectionsanterior to SN (accordingto McGeer et al., 1973) were performed one week prior to sacrifice. Each value represe ts the mean of 3 determinations; for 9 each determination, nigras from 3-5 animals were pooled. H-Enkephalinamidebinding was measured according to the method of Pert and Snyder (1974) with minor modifications. Tissues were homogenized in 0.32 M sucrose. The crude P2 fraction was washed once with ice-cold water and resuspended in 50 mM Tris;HCl (pH 7.4). The membranes were preincubated at 37O for 30 min and then incubated with 4 nM H-enkephalinamide (38 Ci/nnnol)at O" for 1 hr. After incubation, samples were centrifuged and the pellets were washed once with 1 ml of ice-cold water and dissolved in tissu~4solubilizerto measure radioactivity. Non-specific binding was measured in the presence of 10 M morphine, and was subtracted from tot31 binding to calculate specific binding. In separate experiments the kinetic constants of H-enkephalinamidebinding in control SN were determined. Two ED values (high affinity = 0.33 nM and low affinity = 1.6 nM) were observed. The Bmax was 20 fmol/mg prot for the high affinity component and 130 fmol/mg prot for the low affinity component. The observation that there is no detectable loss of opiate receptors associated with the 6OHDA-induced degeneration of DA cells in SN , makes it unlikely that enkephalinergicneurons synapse with DA neurons in SN. On the other hand, since the major portion of opiate receptors in SN appears to be associated with other (non-DA) neuronal connections between the SN and forebrain, it is likely that morphine may act indirectly by influencing the release of a transmitter from another pathway which forms synaptic connections with DA neurons in SN. Our previous studies have suggested that striatonigralpathways containing GABA and substance P are probably involved in the activation of striatal TH induced by haloperidol (Gale et al., 1978). It is possible that activation of opiate receptors in SN could modify the release of one of these neuromodulators. For example, morphine could prevent the haloperidol effect on striatal TH by causing the release of GABA in SN, since it has been demonstrated that stimulation of nigral GABA receptors by muscimol blocks the activation of striatal TH (Gale and Guidotti, 1976). However, the inability of the GABA receptor antagonist, bicuculline methiodide, to reverse the action of morphine, makes this possibility unlikely. Among other possibilities we would like to suggest that opiate receptors may be located on the substance-P containing axons which terminate in SN. It is possible that morphine could reduce the excitatory influence of substance P by inhibiting its release onto DA cells. Consistent with this possibility is the recent observatio9 that lesions of the striatonigral pathways with kainic acid cause a marked reduction in H enkephaline binding in SN (Schwartz, 1979). These data are also consistent with observations that substance P axons in CNS (Jesse1 and Iversen, 1977) contain opiate receptors, suggesting a presynaptic regulation of substance P neurons by opiate interneurons. At the same time we must entertain the possibility that opiate receptors located on other neural pathways may exert a regulatory influence on DA neurons through interactions which have yet to be identified. In addition, wherever the opiate receptors are located in SN, it appears that they may be able to influence efferent pathways other than those containing DA, since we observed that the contralateral postural asymmetry produced by intranigral morphine was not blocked by haloperidol or 6-OHDA pretreatment (data not shown). Thus, it appears that opiate peptides may represent an additional neuromodulatory factor in SN which, by activating opiate receptors located on terminals containing substance P and/or other neurotransmitters,can influence the activity of dopaminergic or nondopaminergic nigral projections.
430
Preliminary
Notes
REFERENCES J. Dray, A. and Straughan, D.W.S. (1976). Synaptic mechanisms in the substantia nigra. Pharm. Pharmacol. 23: 400-405. Gale, K., Costa, E., Toffano, G., Hong, J. S. and Guidotti, A. (1978). Evidence for a role of nigral Y-aminobutyric acid and substance P in the haloperidol-induced activation of striatal tyrosine hydroxylase. J. Pharmacol. Exp. Ther. 206: 29-37. Gale, K. and Guidotti, A. (1976). GABA mediated control of rat neostriatal tyrosine hydroxylase revealed by intranigral muscimol. Nature 263: 691-693. Guidotti, A., Moroni, F., Gale, K. and Kumakura, K. (1979). Opiate receptor stimulation block the activation of striatal tyrosine hydroxylase induced by haloperidol. In: Catecholamine Pergamon Press, New York (in press). Basic and Clinical Frontiers (Usdin, E., Ed.). Iwamoto, E. and Way, L. (1977). Circling behaviour and stereotypy induced by intranigral opiate microinjections. J. Pharmacol. Exp. Ther. 203: 347-359. Jessell, T. M. and Iversen, L. L. (1977). Opiate analgesics inhibit substance P release from Nature 268: 549-551. rat trigeminal nucleus. McGeer, E. G., Fibiger, H. C., McGeer, P. and Brooke, S. (1973). Temporal changes in amine synthetizing enzymes of rat extrapyramidal structures after hemitransections or 6-hydroxydopamine administration. Brain Res. 52: 289-300. Pert, C. B. and Snyder, S. H. (1974). Opiate receptor binding of agonists and antagonists affected differentially by sodium. Mol. Pharmacol. 10: 868-879. Pollard, H., Llorens, C., Schwartz, J. C., Cross, C. and Dray, F. (1978). Localization of opiate receptors and enkephalins in the rat striatum in relationship with the r%gro striatal dopaminergic system. Brain Res. 151: 392-398. Schwartz, J. C., personal communication. Yang, H.-Y.T., Hong, J. S., Fratta, W. and Costa, E. (1978). Rat brain enkephalins: Distribution and biosynthesis. Adv. Biochem. Psychopharmacol. 18: 149-159. Zivkovic, B., Guidotti, A. and Costa, E. (1974). Effects of neuroleptics on striatal tyrosine hydroxylase: Changes in affinity for pteridine cofactor. Mol. Pharmacol. 10: 727-735.
K.
Gale*,
F.
Moronir
M.
Kumakura
and
A, Guidotti
Laboratory of PreclinicalPharmacology,National fnstitute of Mental Health, Saint Elizabeths Hospital, Washington,D.C. 20032 *Departmentof Pharmacology,Georgetown University, Washington, D.C, 20(107
Morphine injected into the substantianiqxa fSN) of rats failed to change DA metabolism in striatusn but blocked the haloperidol induced activation of striatal tyrosine hydroxylase (TX). This blockade was mediated by stimulationof opiate recept3rs in SN because it was antagoni2xXi by naltrexone, The binding of H-enkephalinamideto opiate receptors in SN was not altered after lesion of the niqrostriatalDA pathways with 6hydroxydopaminebut was decreased (approximately30%) after brain hemitransection. These results sugqest that intraniqralmorphine prevents the haloperidol induced activation of striatal TfIby modulating the release of endogenous transmittersor modulators on DA neurons. Afferent projections to substantia nigra (SN) include pathways Containing GABA, substance P and serotonin. These transmittershave been shown to affect both efectrophysioloqicaland neurochemical. function of efferent pathways from SN, including the nigrostriataldopamine (DA) projection ffor a review see Dray and Straughan, 19761. Recently, met-enkephalinhas been maasured ih SN (Yang et al., X978), and although the concentrationof this peptide in SB is not as high as that found in other nuclei of the basal ganglia, opiate receptors a~@ abundant in SN (Pollardet al., 19781. This suggests that endogenous opiate peptides may represent an important neurotransmittersystem in the regulationof the activity of SN neurons. At present, the only evideaee in the literaturewhich suggests a functional role for the opiate'receptarsin SN is the observation that Unilateral intraniqxal injection of morphine or beta-endorphininduces contralateralcircling behavior which can be reversed by naloxone (Iwamotoand Way, 1978). In order tm determine whether activation of these opiate receptors in 5% could have functional importance for the regulation of nigrostriatalDA neurons, we chose to study the effect of intranigrallyapplied morphine on the tyrosine hydroxylase ITHI activity in the DA terminals of striatum and to characterize the neuronal population of SN where the opiate receptors are located. After systemic or intrastriatalinjectionsof morphineI an increase in DA metabolism was found in rat striatum. However, 30 min. after injection of morphine directly into SN (in doses of 1,2,5,10 and 20 Ug of the f.90) salt, dissolved in 0,s ~1 saline) we found no change in striatal IF.TA content (Guidottiet al., 1979). These results suggest that unlike the opiate receptors in striatum, the opiate receptors in SN do not mediate the morphine-inducedstimulation of strkatal RA metabolismIn order to determine whether activation of opiate receptors in .W night exert an inhibitory effect on DA neuronsI we examined whether intrraniqraf morphine could inhibit the activation of striatal TN.induced by haloperidol. The action of haluperidolon striatal TH is mediated via the transsynaptisregulation of DA neurons by niqral afferents, including the striate-dgral axon8 containing GABA or substance P (Gale et al., 1978). We therefore expected that if the nigxal opiate receptors exert an inhibitory influence on DA neurons (eitherdirectly ox indirectly),then intraniqralmorphine would be able to antagonize the activation of striatal TN elicited by haloperidol, AS shown in Fig, 1, although fntranigralmorphine by itself did not change the affinity of striatal 9z-X for nMPa , it completely prevented the increase in affinity of striatal Ttrfor DMPE4 in animals trentttdw$th haloperifiol.This effect of intranigralmorphine was reversed by systemic (Fig+ I) or intranigral. injection (Table 1) of the opiate-receptoxantagonist, naltrexone, but was nat reversed by intranigral injection of bicuculline methiodide (Table 1) a EABA receptor antagonist. The specificityof the action of morphine was further sup&orted by experiments which demonstrated that the ability of muscimol to antagonize the haloperidol-induced
427
428
Preliminary Notes
activation of striatal TH was reversed by intranigral bicuculline methiodide but not by naltrexone (Table 1).
htranigral morpthe blockade
ofHabperii-inducedktivatii
of Striatal TH
l
Mmpkm
\lOvg intronigrd)
0.37 0 HdopriQ((lmg/kg i.P.1
Figure 1.
DOUBLE RECIPROCAL PMTS OF THE INITIAL VELOCITY OFl$RIATAL TH AGAINST VARIOUS CONCENTRATIONS OF DNPH4 IN THE PRESENCE OF 0.1 ml4 C-TYROSINB. TH activity was determined using a dopadecarboxylasecoupled enzymatic reaction to measure 14 CO2 formed from carboxyl labelled tyrosine according to the method of Eivkovic et al. (1974). Haloperidol was injected 30 min prior sacrifice, morphine sulfate 10 min before haloperidol and naltrexone 10 min after haloperidol. Each point represent the mean obtained from six animals. The standard error of the Km values was between 5 and 10%. Stereotaxic coordinates for the intranigral injections were the same as those used by Gale and Guidotti (1976). The volume The Km for TH of untreated rats was 0.83 mM. injected into SN was 0.5 W. Table 1.
Relationship Between Opiate Receptors and GABA Receptors in SN for The Control of Haloperidol-InducedActivation of Striatal TH A Intravenous Injection
B First Intranigral Injection
C Second Intranigral Injection Saline (Km of
Bicuculline Naltrexone TH for DNPH4; mM)
Striatal
SALINE
Saline Muscimol Morphine
.85 .80 .83
.83 .88 .87
.79 .87 .85
HALOPERIDOL
Saline Muscimol Morphine
.37* .85 .73
.32* .91 .30*
.42* .40* .78
A) Saline or haloperidol (0.3 umoles/kg) was administered 30 min prior to sacrifice. B) Saline (0.5 W), morphine sulfate (10 llgin 0.5 ~1) or muscimol (100 ng in 0.5 ~1) was injected 10 min before haloperidol; C) Saline (0.5 ~11, naltrexone (10 vg in 0.5 ~1) or bicuculline methiodide (500 ng in 0.5 ~1) was injected 10 min after haloperidol. TH measurements were done as described in the legend of Fig. 1. Values represent the mean of 6-8 animals. Standard errors were less than 15% of the mean. *Significantlydifferent (P<.Ol) from saline (systemic+ intranigral treated controls. These results suggest that activation of opiate receptors in SN inhibits the activity of DA neurons. In order to determine whether this effect could be mediated by a direct action of morphine on DA cells in SN, we measured opiate receptor binding in SN after unilateral destruction of nigrostriatal DA neurons by 6-hydroxydopamine (6-OHDA). Although 6-OHDA treatment resulted in the destruction of more than 85% of the nigrostriatal DA3neurons (based on TH measurements), no significant change was obtained in the binding of H enkephalinamide in the
Preliminary Notes
crude ~2 fraction of the lesioned SN (Table 2). In contrast, there was a marked (68%) decrease in the binding of 3H enkephalinamide to SN P2 preparation from rats with a mechanical hemitransection of all the connections between the SN and striatum (Table 2). This lesion reduced both GABA and substance P content in SN by 60-85% (Gale et al., 1978). Table 2.
3 H-EnkephalinamideBinding in Rat SN After C-OHDA or Hemitransection 3H-Enkephalinamide(fmol/mgprot) Lesion
Intact Side
6-OHDA
105 zt12
95
Hemitransection
110 t
35 f
8
Lesioned Side
i
% Decrease Due to Lesion
15 6*
68
Bats (250 g) were anesthetized and 6-OHDA (8 ug in 4 ~1) was injected into SN at the rate of .25 ul/min. Stereotaxic coordinates were the same as those used for intranigral injections in Fig. 1. Animals were sacrificed 2 weeks after 6-OHDA treatment. Measurement of TH demonstrated a decrease of 85-98%. Hemitransectionsanterior to SN (accordingto McGeer et al., 1973) were performed one week prior to sacrifice. Each value represe ts the mean of 3 determinations; for 9 each determination, nigras from 3-5 animals were pooled. H-Enkephalinamidebinding was measured according to the method of Pert and Snyder (1974) with minor modifications. Tissues were homogenized in 0.32 M sucrose. The crude P2 fraction was washed once with ice-cold water and resuspended in 50 mM Tris;HCl (pH 7.4). The membranes were preincubated at 37O for 30 min and then incubated with 4 nM H-enkephalinamide (38 Ci/nnnol)at O" for 1 hr. After incubation, samples were centrifuged and the pellets were washed once with 1 ml of ice-cold water and dissolved in tissu~4solubilizerto measure radioactivity. Non-specific binding was measured in the presence of 10 M morphine, and was subtracted from tot31 binding to calculate specific binding. In separate experiments the kinetic constants of H-enkephalinamidebinding in control SN were determined. Two ED values (high affinity = 0.33 nM and low affinity = 1.6 nM) were observed. The Bmax was 20 fmol/mg prot for the high affinity component and 130 fmol/mg prot for the low affinity component. The observation that there is no detectable loss of opiate receptors associated with the 6OHDA-induced degeneration of DA cells in SN , makes it unlikely that enkephalinergicneurons synapse with DA neurons in SN. On the other hand, since the major portion of opiate receptors in SN appears to be associated with other (non-DA) neuronal connections between the SN and forebrain, it is likely that morphine may act indirectly by influencing the release of a transmitter from another pathway which forms synaptic connections with DA neurons in SN. Our previous studies have suggested that striatonigralpathways containing GABA and substance P are probably involved in the activation of striatal TH induced by haloperidol (Gale et al., 1978). It is possible that activation of opiate receptors in SN could modify the release of one of these neuromodulators. For example, morphine could prevent the haloperidol effect on striatal TH by causing the release of GABA in SN, since it has been demonstrated that stimulation of nigral GABA receptors by muscimol blocks the activation of striatal TH (Gale and Guidotti, 1976). However, the inability of the GABA receptor antagonist, bicuculline methiodide, to reverse the action of morphine, makes this possibility unlikely. Among other possibilities we would like to suggest that opiate receptors may be located on the substance-P containing axons which terminate in SN. It is possible that morphine could reduce the excitatory influence of substance P by inhibiting its release onto DA cells. Consistent with this possibility is the recent observatio9 that lesions of the striatonigral pathways with kainic acid cause a marked reduction in H enkephaline binding in SN (Schwartz, 1979). These data are also consistent with observations that substance P axons in CNS (Jesse1 and Iversen, 1977) contain opiate receptors, suggesting a presynaptic regulation of substance P neurons by opiate interneurons. At the same time we must entertain the possibility that opiate receptors located on other neural pathways may exert a regulatory influence on DA neurons through interactions which have yet to be identified. In addition, wherever the opiate receptors are located in SN, it appears that they may be able to influence efferent pathways other than those containing DA, since we observed that the contralateral postural asymmetry produced by intranigral morphine was not blocked by haloperidol or 6-OHDA pretreatment (data not shown). Thus, it appears that opiate peptides may represent an additional neuromodulatory factor in SN which, by activating opiate receptors located on terminals containing substance P and/or other neurotransmitters,can influence the activity of dopaminergic or nondopaminergic nigral projections.
430
Preliminary
Notes
REFERENCES J. Dray, A. and Straughan, D.W.S. (1976). Synaptic mechanisms in the substantia nigra. Pharm. Pharmacol. 23: 400-405. Gale, K., Costa, E., Toffano, G., Hong, J. S. and Guidotti, A. (1978). Evidence for a role of nigral Y-aminobutyric acid and substance P in the haloperidol-induced activation of striatal tyrosine hydroxylase. J. Pharmacol. Exp. Ther. 206: 29-37. Gale, K. and Guidotti, A. (1976). GABA mediated control of rat neostriatal tyrosine hydroxylase revealed by intranigral muscimol. Nature 263: 691-693. Guidotti, A., Moroni, F., Gale, K. and Kumakura, K. (1979). Opiate receptor stimulation block the activation of striatal tyrosine hydroxylase induced by haloperidol. In: Catecholamine Pergamon Press, New York (in press). Basic and Clinical Frontiers (Usdin, E., Ed.). Iwamoto, E. and Way, L. (1977). Circling behaviour and stereotypy induced by intranigral opiate microinjections. J. Pharmacol. Exp. Ther. 203: 347-359. Jessell, T. M. and Iversen, L. L. (1977). Opiate analgesics inhibit substance P release from Nature 268: 549-551. rat trigeminal nucleus. McGeer, E. G., Fibiger, H. C., McGeer, P. and Brooke, S. (1973). Temporal changes in amine synthetizing enzymes of rat extrapyramidal structures after hemitransections or 6-hydroxydopamine administration. Brain Res. 52: 289-300. Pert, C. B. and Snyder, S. H. (1974). Opiate receptor binding of agonists and antagonists affected differentially by sodium. Mol. Pharmacol. 10: 868-879. Pollard, H., Llorens, C., Schwartz, J. C., Cross, C. and Dray, F. (1978). Localization of opiate receptors and enkephalins in the rat striatum in relationship with the r%gro striatal dopaminergic system. Brain Res. 151: 392-398. Schwartz, J. C., personal communication. Yang, H.-Y.T., Hong, J. S., Fratta, W. and Costa, E. (1978). Rat brain enkephalins: Distribution and biosynthesis. Adv. Biochem. Psychopharmacol. 18: 149-159. Zivkovic, B., Guidotti, A. and Costa, E. (1974). Effects of neuroleptics on striatal tyrosine hydroxylase: Changes in affinity for pteridine cofactor. Mol. Pharmacol. 10: 727-735.