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Discussion of Serotonin and Dopamine in the Extrapyramidal System
Discussion of Serotonin and Dopamine in the Extrapyramidal System NILS-ERIK AND~N Department of Pharmacology, University of Goteborg, Goteborg, Sweden
Corpus Striatum The results of Sourkes and Poirier indicate t h a t the course of striatopetal dopamine and 5-HT neurons of monkey and cat are similar t o those of rat. I n this species, some days after extensive removal of the terminals in the neostriatum (the caudate nucleusputamen), we have traced under the fluorescence microscope the nigro-neostriatal dopamine neurons (AndBn et al., 1966a). We have found that most of the nigro-neostriatal dopamine fibers after they leave the area of substantia nigra become aggregated in a bundle which ascends rostrally just medial and dorsomedial to the ventral part of the crus cerebri. At the level of the caudal part of the median eminence the bundle turns into the ventral and rostra1 part of the crus cerebri. Thereafter, the fibers enter into the retrolenticular part of the internal capsule, diverge, and running rostrally and dorsally they approach the neostriatum via the fibers of the capsula interna. I n recent uptake experiments with brain slices (Fuxe and Hamberger, unpublished) we observed that the axons after leaving the cell bodies of the pars compacta of the substantia nigra bend toward the medial lemniscus and the red nucleus before they become aggregated and take a ventral-rostra1 direction. The lesions producing contralateral hypokinesia and ipsilateral striatal dopamine loss reported by Sourkes and Poirier obviously destroyed the nigro-neostriatal pathway just after these fiaers become aggregated. Following lesions in the mesencephalic raphe region, where many 6-HT cell bodies are located, a large part of the forebrain 5-HT was lost ( A n d h , Fuxe, and Ungerstedt, unpublished). In nialamide-treated animals (Dahlstrom and Fuxe, 1964) and after intreventricular injections of 5-HT (Fuxe, Hokfelt, and Ungerstedt, this symposium) we could observe that the fibers from these cell bodies bend toward the basal surface of the brain and enter the medial forebrain bundle at the mamillary body. The lesions in the experiments of Sourkes and Poirier, causing either contralateral tremor or ballism or choreiform movements in combination with ipsilateral striatal 5-HT disappearance, seem to sever the 5-HT fibers when they are bending ventrally-rostrally. The 5-HT in the corpus striatum probably derives from the fibers in the medial forebrain bundle since a lesion of that structure caused a loss of 5-HT terminals in the globus pallidus, the only part of the corpus striatum where the 5-HT normally can be seen histochemitally (And& et al., 1905). I n the neostriatum the 5-HT terminals are hard to detect due to the very dense network of dopamine terminals. We have studied the function of the monoamine nerves in the corpus striatum by observing the asymmetries occurring after a unilateral lesion of the striatopetal mono&mine pathways combined with drugs interfering with the monoamine metabolism ( A n d h et al., 1966b). Using this model, the 5-HT precursor 6-HTP in contrast to the catecholamine precursor dopa did not produce any clear-cut asymmetries although pharmacological effects such as tremor in the forelimbs and hyperextension of the hindlimbs were present. However, reserpine but not haloperidol or chlorpromazine 347
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caused tremor on the side opposite to the lesion. Since reserpine but not haloperidol or chlorpromazine inhibits transmission of nerve impulses mediated by 5-HT (vide infra), this finding may indicate that the tremor is due to functional deficiency of 5-HT in agreement with the data of Sourkes and Poirier. A lesion of the striatopetal dopamine neurons caused a pronounced reduction of both the dopamine and the dopa decarboxylase activity in this brain part, in all probability due to the fact that the amine.and the enzyme normally are present in the same nerve terminals and therefore disappear simultaneously with the anterograde degeneration of the dopamine neurons (AndQnet al., 1966a). The diminution of dopa decarboxylase activity in the denervated corpus striatum explains the reduced formation of .dopamine from dopa in the experiments of Sourkes and Poirier. Injection of dopa to rats unilaterally operated in this way produced turning or rotating of the animals to the side of the lesion (AndBn et aZ., 1966b).
Spinal Cord Motor functions are influenced by 5-HT also at other levels in the central nervous system than in the corpus striatum, e.g., in the spinal cord. All the monoamine nerve terminals of the spinal cord belong t o neurons whose cell bodies are present in the lower brainstem. Injection of monoamine precursors produces a formation of amines also in the caudal part of a transected cord and the functional changes occurring there cannot be due to a n action of amines formed in other parts of the central nervous system. First it was shown that injection of 5-HTP, like dopa, caused an increase of the flexor reflex in the acutely spinalized rabbit (Carlsson et d., 1963) and cat (AndBn et al., 1964b). An electrophysiological analysis revealed that 5-HTP like dopa inhibited the transmission of impulses from the flexor reflex afferents to the primary afferents, the motoneurons, and certain ascending fibers (And& et al. 1964a). A delayed increase of the discharge of flexor motoneurons evoked by a train of impulses in the flexor reflex afferents was thereby obtained, probably due to opening of a pathway normally concealed in the spinalized animal (AndBn et al., 1964b). I n addition, 5-HTP but not dopa injection produced increased excitability of the motoneurons and a n enhancement of the monosynaptic reflex (AndBn et d.,1964a, Anderson and Shibuya, 1966). The injection of 5-HTP mimicked the effect of electrical stimulation of reticulo-spinal neurons in the caudal medulla oblongata, where the cell bodies of the bulbo-spinal 5-HT neurons occur. The effect of stimulation of this pathway was reduced by reserpine. Thus, reserpine seems to inhibit the central 5-HT transmission in the same way as the peripheral NE transmission (AndBn et al., 1964a). I n rabbit and cat the spinal effects elicited by 5-HTP and dopa are similar; therefore, in these two species it is not easy to determine the functional roles of 5-HT and NE neurons in the spinal cord. I n the spinal rat, however, the two monoamine precursors produced different actions: dopa produced a n increased flexor reflex whereas 5-HTP produced hyperextension, athetoid movements, and a weak tremor of about the same magnitude as that elicited in intact rats. These effects are central because they could not be reproduced by infusion of NE and 5-HT. Moreover, they are elicited by the amines because the effects of the amino acids were blocked after decarboxylase inhibition (m-hydroxybenzylhydrazine)and potentiated after MA0 inhibition (nialamide, harmaline). Phenoxybenzamine, haloperidol, and chlorpromazine blocked the effects of dopa on the spinal reflexes without changing the formation of catecholamines but they failed to alter the actions of 5-HTP. These findings indicate t h a t these drugs block the cate-
DISCUSSION
349
cholamine but not the 5-HT receptors of the postsynaptic cells, i.e., catecholamines and 5-HT act on different receptors. Many drugs that block peripheral 5-HT receptors may also block 5-HT receptors in the central nervous system. Some of these drugs (bromolysergic acid diethylamideBOL, methysergide, cyproheptadine, atropine, morphine, benzquinarnide) were tested for blockage of the spinal 5-HTP actions, but all of them failed in this respect. On the other hand, lysergic acid diethylamide (LSD) showed a n effect on the spinal reflexes similar t o that seen after 5-HTP injection and the same was true for Np-dimethyltryptamine and tryptamine. Since the actions of LSD and the tryptamines were not diminished after pre-treatment with reserpine and H 22/54 ( a-propyldopacetamide, an inhibitor of tryptophan hydroxylase) i t is possible that these compounds stimulate the 6-HT receptors. An action on the spinal reflexes similar t o that of 5-HTP could be seen some hours after injecting a large dose of an MA0 inhibitor (nialamide 500 mg/kg) to a spinalized rat treated with reserpine the day before. Under these conditions 5-HT but not NE accumulated in the spinal cord including the part caudal t o the transection. This accumulation could be reduced by pretreatment with H 22/54, a n inhibitor of 5-HT synthesis; such a pretreatment reduced the action of nialamide on spinal reflexes. Harmaline, like nialamide, affectad spinal reflexes; however, this drug action appeared somewhat faster and lasted for only a few hours. A possible 5-HT receptor blockade by harmaline was tested by injectionof this drug 30 minutes before the 5-HTP. However, harmaline failed t o block the action of 5-HTP. Therefore, the pharmacological effects of harmaline in the spinal cord seem t o be due to a reversible MA0 inhibition. ACKNOWLEDGMENT This work has been supported by the Swedish Medical Research Council (K67-14X502-03), “Svenska livforsiikringsbolags niimnd for medicinsk forskning”, and “Therese och Johan Anderssons minne . ’ 9 REFERENCES AndBn, N.-E., Jukes, M. G. M., and Lundberg, A. (1964a). Nature 202, 1222. A n d h , N.-E., Jukes, M. G. M., Lundberg, A., and Vyklickj., L. (1964b). Nature 202, 1344. AndBn, N.-E., Dahlstrom, A., Fuxe, K., and Larsson, K. (1965). Li,fe Sci. 4, 1275. AndBn, N.-E., Dahlstrom, A., Fuxe, K., Larsson, K., Olson, L., and Ungerstedt, U. (1966a). Acta Phywiol. Scand. 67, 313. And&, N.-E., Dahlstrom, A., Fuxe, K., and Larsson, K. (1966b). Acta Phamacol. Toxicol. 24, 263. Anderson, E. G., and Shibuya, T. (1966). J. Pharnzacol. Exptl. Therap. 158, 352. Carlsson, A., Magnusson, T., and Rosengren, E. (1963). Experientia 19, 359. Dahlstrom, A., and Fuxe, K. (1964). Acta Physiol. Scand. Suppl. 232.
Corpus Striatum The results of Sourkes and Poirier indicate t h a t the course of striatopetal dopamine and 5-HT neurons of monkey and cat are similar t o those of rat. I n this species, some days after extensive removal of the terminals in the neostriatum (the caudate nucleusputamen), we have traced under the fluorescence microscope the nigro-neostriatal dopamine neurons (AndBn et al., 1966a). We have found that most of the nigro-neostriatal dopamine fibers after they leave the area of substantia nigra become aggregated in a bundle which ascends rostrally just medial and dorsomedial to the ventral part of the crus cerebri. At the level of the caudal part of the median eminence the bundle turns into the ventral and rostra1 part of the crus cerebri. Thereafter, the fibers enter into the retrolenticular part of the internal capsule, diverge, and running rostrally and dorsally they approach the neostriatum via the fibers of the capsula interna. I n recent uptake experiments with brain slices (Fuxe and Hamberger, unpublished) we observed that the axons after leaving the cell bodies of the pars compacta of the substantia nigra bend toward the medial lemniscus and the red nucleus before they become aggregated and take a ventral-rostra1 direction. The lesions producing contralateral hypokinesia and ipsilateral striatal dopamine loss reported by Sourkes and Poirier obviously destroyed the nigro-neostriatal pathway just after these fiaers become aggregated. Following lesions in the mesencephalic raphe region, where many 6-HT cell bodies are located, a large part of the forebrain 5-HT was lost ( A n d h , Fuxe, and Ungerstedt, unpublished). In nialamide-treated animals (Dahlstrom and Fuxe, 1964) and after intreventricular injections of 5-HT (Fuxe, Hokfelt, and Ungerstedt, this symposium) we could observe that the fibers from these cell bodies bend toward the basal surface of the brain and enter the medial forebrain bundle at the mamillary body. The lesions in the experiments of Sourkes and Poirier, causing either contralateral tremor or ballism or choreiform movements in combination with ipsilateral striatal 5-HT disappearance, seem to sever the 5-HT fibers when they are bending ventrally-rostrally. The 5-HT in the corpus striatum probably derives from the fibers in the medial forebrain bundle since a lesion of that structure caused a loss of 5-HT terminals in the globus pallidus, the only part of the corpus striatum where the 5-HT normally can be seen histochemitally (And& et al., 1905). I n the neostriatum the 5-HT terminals are hard to detect due to the very dense network of dopamine terminals. We have studied the function of the monoamine nerves in the corpus striatum by observing the asymmetries occurring after a unilateral lesion of the striatopetal mono&mine pathways combined with drugs interfering with the monoamine metabolism ( A n d h et al., 1966b). Using this model, the 5-HT precursor 6-HTP in contrast to the catecholamine precursor dopa did not produce any clear-cut asymmetries although pharmacological effects such as tremor in the forelimbs and hyperextension of the hindlimbs were present. However, reserpine but not haloperidol or chlorpromazine 347
348
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caused tremor on the side opposite to the lesion. Since reserpine but not haloperidol or chlorpromazine inhibits transmission of nerve impulses mediated by 5-HT (vide infra), this finding may indicate that the tremor is due to functional deficiency of 5-HT in agreement with the data of Sourkes and Poirier. A lesion of the striatopetal dopamine neurons caused a pronounced reduction of both the dopamine and the dopa decarboxylase activity in this brain part, in all probability due to the fact that the amine.and the enzyme normally are present in the same nerve terminals and therefore disappear simultaneously with the anterograde degeneration of the dopamine neurons (AndQnet al., 1966a). The diminution of dopa decarboxylase activity in the denervated corpus striatum explains the reduced formation of .dopamine from dopa in the experiments of Sourkes and Poirier. Injection of dopa to rats unilaterally operated in this way produced turning or rotating of the animals to the side of the lesion (AndBn et aZ., 1966b).
Spinal Cord Motor functions are influenced by 5-HT also at other levels in the central nervous system than in the corpus striatum, e.g., in the spinal cord. All the monoamine nerve terminals of the spinal cord belong t o neurons whose cell bodies are present in the lower brainstem. Injection of monoamine precursors produces a formation of amines also in the caudal part of a transected cord and the functional changes occurring there cannot be due to a n action of amines formed in other parts of the central nervous system. First it was shown that injection of 5-HTP, like dopa, caused an increase of the flexor reflex in the acutely spinalized rabbit (Carlsson et d., 1963) and cat (AndBn et al., 1964b). An electrophysiological analysis revealed that 5-HTP like dopa inhibited the transmission of impulses from the flexor reflex afferents to the primary afferents, the motoneurons, and certain ascending fibers (And& et al. 1964a). A delayed increase of the discharge of flexor motoneurons evoked by a train of impulses in the flexor reflex afferents was thereby obtained, probably due to opening of a pathway normally concealed in the spinalized animal (AndBn et al., 1964b). I n addition, 5-HTP but not dopa injection produced increased excitability of the motoneurons and a n enhancement of the monosynaptic reflex (AndBn et d.,1964a, Anderson and Shibuya, 1966). The injection of 5-HTP mimicked the effect of electrical stimulation of reticulo-spinal neurons in the caudal medulla oblongata, where the cell bodies of the bulbo-spinal 5-HT neurons occur. The effect of stimulation of this pathway was reduced by reserpine. Thus, reserpine seems to inhibit the central 5-HT transmission in the same way as the peripheral NE transmission (AndBn et al., 1964a). I n rabbit and cat the spinal effects elicited by 5-HTP and dopa are similar; therefore, in these two species it is not easy to determine the functional roles of 5-HT and NE neurons in the spinal cord. I n the spinal rat, however, the two monoamine precursors produced different actions: dopa produced a n increased flexor reflex whereas 5-HTP produced hyperextension, athetoid movements, and a weak tremor of about the same magnitude as that elicited in intact rats. These effects are central because they could not be reproduced by infusion of NE and 5-HT. Moreover, they are elicited by the amines because the effects of the amino acids were blocked after decarboxylase inhibition (m-hydroxybenzylhydrazine)and potentiated after MA0 inhibition (nialamide, harmaline). Phenoxybenzamine, haloperidol, and chlorpromazine blocked the effects of dopa on the spinal reflexes without changing the formation of catecholamines but they failed to alter the actions of 5-HTP. These findings indicate t h a t these drugs block the cate-
DISCUSSION
349
cholamine but not the 5-HT receptors of the postsynaptic cells, i.e., catecholamines and 5-HT act on different receptors. Many drugs that block peripheral 5-HT receptors may also block 5-HT receptors in the central nervous system. Some of these drugs (bromolysergic acid diethylamideBOL, methysergide, cyproheptadine, atropine, morphine, benzquinarnide) were tested for blockage of the spinal 5-HTP actions, but all of them failed in this respect. On the other hand, lysergic acid diethylamide (LSD) showed a n effect on the spinal reflexes similar t o that seen after 5-HTP injection and the same was true for Np-dimethyltryptamine and tryptamine. Since the actions of LSD and the tryptamines were not diminished after pre-treatment with reserpine and H 22/54 ( a-propyldopacetamide, an inhibitor of tryptophan hydroxylase) i t is possible that these compounds stimulate the 6-HT receptors. An action on the spinal reflexes similar t o that of 5-HTP could be seen some hours after injecting a large dose of an MA0 inhibitor (nialamide 500 mg/kg) to a spinalized rat treated with reserpine the day before. Under these conditions 5-HT but not NE accumulated in the spinal cord including the part caudal t o the transection. This accumulation could be reduced by pretreatment with H 22/54, a n inhibitor of 5-HT synthesis; such a pretreatment reduced the action of nialamide on spinal reflexes. Harmaline, like nialamide, affectad spinal reflexes; however, this drug action appeared somewhat faster and lasted for only a few hours. A possible 5-HT receptor blockade by harmaline was tested by injectionof this drug 30 minutes before the 5-HTP. However, harmaline failed t o block the action of 5-HTP. Therefore, the pharmacological effects of harmaline in the spinal cord seem t o be due to a reversible MA0 inhibition. ACKNOWLEDGMENT This work has been supported by the Swedish Medical Research Council (K67-14X502-03), “Svenska livforsiikringsbolags niimnd for medicinsk forskning”, and “Therese och Johan Anderssons minne . ’ 9 REFERENCES AndBn, N.-E., Jukes, M. G. M., and Lundberg, A. (1964a). Nature 202, 1222. A n d h , N.-E., Jukes, M. G. M., Lundberg, A., and Vyklickj., L. (1964b). Nature 202, 1344. AndBn, N.-E., Dahlstrom, A., Fuxe, K., and Larsson, K. (1965). Li,fe Sci. 4, 1275. AndBn, N.-E., Dahlstrom, A., Fuxe, K., Larsson, K., Olson, L., and Ungerstedt, U. (1966a). Acta Phywiol. Scand. 67, 313. And&, N.-E., Dahlstrom, A., Fuxe, K., and Larsson, K. (1966b). Acta Phamacol. Toxicol. 24, 263. Anderson, E. G., and Shibuya, T. (1966). J. Pharnzacol. Exptl. Therap. 158, 352. Carlsson, A., Magnusson, T., and Rosengren, E. (1963). Experientia 19, 359. Dahlstrom, A., and Fuxe, K. (1964). Acta Physiol. Scand. Suppl. 232.