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Effects of 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine on neostriatal dopamine in mice
0028-3908/84 $3.00 + 0.00 Pergamon Press Ltd
Neuropharmacology Vol. 23, No. 6, pp. 71 I-713, 1984 Printed in Great Britain
EFFECTS
OF l-METHYL-4-PHENYL-1,2,5,6_TETRAHYDROPYRIDINE ON NEOSTRIATAL DOPAMINE IN MICE
R.E. Heikkila,
F.S. Cabbat,
L. Manzino
and R.C. Duvoisin
Department of Neurology University of Medicine and Dentistry of New JerseyRutgers Medical School, Piscataway, N J 08854 U.S.A.
(Accepted
b Aphie
19841
1-Methyl-4-phenyl-1,2,5,6_tetrahydropyridine (MPTP) causes a estruction of the nigrostriatal dopamine pathway in humans as well However, it has been reported that MPTP is inert in as in monkeys. We now report that MPTP, given to mice several small animal species. at 30 mg/kg intraperitoneally, causes severe and long-lasting depletions of dopamine and its major metabolites dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in the neostriatum.
SF’
Since the discovery that the self-administration of 1-methyl-4-phenyl-1,2,5,6tetrahydropyridine (MPTP) along with several other agents induced an irreversible Parkinson syndrome in young drug addicts, including a very extensive dopamine depletion and degeneration of the substantia nigra (Davis et al., 1979; Langston et al., 19831, several investigators have attempted to replicate this phenomenon in laboratory animals. It was subsequently reported that MPTP administration, by itself, was able to cause a severe depletion in the content of neostriatal dopamine and its metabolites in the monkey (Burns et al., 1983) but had a limited effect in guinea pigs and no effect in several other small animal species including rats and cats (Chiueh et al., 1983). Because of the obvious importance of these above observations, several groups of investigators have independently searched for a convenient small animal in which MPTP administration would have effects similar to those observed in the monkey and human. Not surprisingly, MPTP was administered to several small animals, many of them rather uncommon and rarely used as experimental animals. Being interested in animal models of Parkinsonism as well as in understanding how neurotoxic agents exert their actions, we also tried MPTP in several different small animal species. We now report that MPTP causes extensive and long-lasting decrements in the neostriatal content of dopamine and its major metabolites in mice. TABLE 1.
MPTP
Neostriatal levels of dopamine and its metabolites after the injections of MPTP to mice at 30 mg/kg (0.17 mmoles/kgl. Mice were injected daily the number of times indicated and assays done between 5 and 18 days after the last injections. Data represent the mean value? SD for the number of mice indicated.
Number of Injections
Assay Time (Days)
n
DA
21 +
5
10
+ + +
10
5
+ + +
:o"
;:
20
6
;:
:72
13.8?
DOPAC tIlg/g tissue)
1.6
HVA
4.5_+ 1.5
1.9+
0.4
l.O+
0.3
3
5.1 * 1.2
1.2+
; 7
3.0 * 0.7 2.6? 1.0 2.9* 1.4
1.2+ 0.4 1.1 f 0.3 1.4t 0.3
0.9 f 0.3 0.9 + 0.2 1.0 + 0.1
2 3
2.8 + 0.7 2.6 f 0.8 2.6 + 0.5
1.4* 0.3 1.2+ 0.3 1.4-+ 0.2
0.9 i 0.3 0.9 * 0.1 1.2 + 0.2
711
0.5
712
Preliminary
MATERIALS
Notes
AND METHODS
Male, Swiss-Webster mice (Charles River, CF-Wl weighing between 25 and 35 g were injected intraperitoneallywith MPTP, 30 mg/kg (0.17 mmoles/kgl, in distilled water adjusted to pH 8.5 with dilute hydrochloric acid. The mice received multiple MPTP injections, 24 hours apart and levels of dopamine and its metabolites in the neostriatum were assayed at various time periods after the last injection. The neostriata were weighed and homogenized in 0.1 M perchloric acid containing dihydroxybenzylamine(DHBA) as an internal standard, and centrifuged at 27,000 x g for 15 min. The supernatant was used for assays of dopamine, serotonin and their metabolites (Mayer and Shoup, 1983). A Bioanalytical Systems LC-304T liquid chromatograph connected to a Kipp and Zonen dual pen recorder was used for all assays; the operating potential was 750 mv. RESULTS Control levels of dopamine and its metabolites are presented in Table 1. After five injections of MPTP at 30 mg/kg, with assays ten days after the last injection, there was a substantial decrement in levels of dopamine, DOPAC and HVA compared to control values (Table 11. After ten or twenty injections of MPTP, with assays done between 5 and 18 days after the last injection, the content of dopamine and its metabolites was markedly reduced from control values. Under these conditions, there was approximately an 80% loss of dopamine, a 70% loss of DOPAC and a 50% loss of HVA. DISCUSSION The data of the present study demonstrate that MPTP administration causes a large decrement in the neostriatal level of dopamine and of its metabolites in male Swiss-Webstermice. In other experiments, MPTP administration brought about decrements in neostriatal dopamine and its metabolites in other strains of mice including CF-1, CD-1 and C-57BL. The data indicate that this effect in mice is long-lasting. There was no apparent recovery up to 18 days after 10 injections or up to 17 days after 20 injections (Table 1). In these same experiments, there was no obvious difference in levels of neostriatal serotonin or 5-hydroxyindoleaceticacid between control mice and MPTP-treated mice. The actions of MPTP are thus specific in mice and similar to those previously seen in monkeys. In experiments parallel to those presented in Table 1, we injected rats, guinea pigs and gerbils with the same 30 mg/kg dose of MPTP. In these three species, there was no apparent decrement in the content of neostriatal dopamine, DOPAC and HVA with up to ten injections of MPTP. Thus under identical conditions, the mouse is clearly more sensitive to the neurotoxic effects of MPTP than the rat, guinea pig or gerbil.
In conclusion, we report that MPTP causes a large and apparently permanent depletion in levels of striatal dopamine and its metabolites in mice. In other experiments, MPTP administration caused a long-lasting decrement in the capacity of synaptosomal preparations prepared from mouse neostriatum to take up 3H-dopamine. Moreover, MPTP administration caused a severe loss of nerve cells in the zona compacta of the substantia nigra. The mouse should prove to be a convenient and relatively inexpensive small animal with which to study the metabolism of MPTP. We expect that the mouse will be valuable for investigators to explore in depth the mechanism by which MPTP selectively destroys the nigrostriatal dopaminergic system. Further, it provides a model of a highly selective neuronal system degeneration which may yield clues to the pathogenesis of human Parkinson's disease. ACKNOWLEDGEMENTS Supported in part by a grant from the USPHS and by a grant from the United Parkinson Foundation. REFERENCES Burns, R.S., Chiueh, C.C., Markey, S.P., Ebert, M.H., Jacobowitz, D.M., Kopin, I.J. (1983). A primate model of parkinsonism: Selective destruction of dopaminergic neurons in the pars compacta of the substantia nigra by N-methyl-4-phenyl-1,2,3,6tetrahydropyridine. Proc. Natl. Acad. Sci. U.S.A. -80:4546-4550. Chiueh, C.C., Markey, S.P., Burns, R.S., Johannessen, J., Jacobowitz, D.M., Kopin, I.J. N-methyl-4-phenyl-1,2,3,6_tetrahydropyridine, a parkinsonian syndrome (1983). causing agent in man and monkey, produces different effects in guinea pig and rat. The Pharmacologist,-25:131.
Preliminary
Notes
713
Davis, G.C., William, A.C., Markey, S.P., Ebert, M.H., Caine, E.D., Reicher, C.M., Kopin, I.J. (1979). Chronic parkinsonism secondary to intravenous injection of meperidine Psych. Res. 1:249-254. analogues. Langston, J.W., Ballard, P., Tetrud, J.W., Irwin. I. (1983). humans due to a product of meperidine-analog synthesis.
Chronic parkinsonism in Science, 219:979-980. --
Mayer, G.S., Shoup, R.E. (1983). Simultaneous multiple electrode liquid chromatographIC-electrochemical assay for catecholamines, indoleamines and metabolites in brain J. Chromatog. -255:533-544. tissue.
Neuropharmacology Vol. 23, No. 6, pp. 71 I-713, 1984 Printed in Great Britain
EFFECTS
OF l-METHYL-4-PHENYL-1,2,5,6_TETRAHYDROPYRIDINE ON NEOSTRIATAL DOPAMINE IN MICE
R.E. Heikkila,
F.S. Cabbat,
L. Manzino
and R.C. Duvoisin
Department of Neurology University of Medicine and Dentistry of New JerseyRutgers Medical School, Piscataway, N J 08854 U.S.A.
(Accepted
b Aphie
19841
1-Methyl-4-phenyl-1,2,5,6_tetrahydropyridine (MPTP) causes a estruction of the nigrostriatal dopamine pathway in humans as well However, it has been reported that MPTP is inert in as in monkeys. We now report that MPTP, given to mice several small animal species. at 30 mg/kg intraperitoneally, causes severe and long-lasting depletions of dopamine and its major metabolites dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in the neostriatum.
SF’
Since the discovery that the self-administration of 1-methyl-4-phenyl-1,2,5,6tetrahydropyridine (MPTP) along with several other agents induced an irreversible Parkinson syndrome in young drug addicts, including a very extensive dopamine depletion and degeneration of the substantia nigra (Davis et al., 1979; Langston et al., 19831, several investigators have attempted to replicate this phenomenon in laboratory animals. It was subsequently reported that MPTP administration, by itself, was able to cause a severe depletion in the content of neostriatal dopamine and its metabolites in the monkey (Burns et al., 1983) but had a limited effect in guinea pigs and no effect in several other small animal species including rats and cats (Chiueh et al., 1983). Because of the obvious importance of these above observations, several groups of investigators have independently searched for a convenient small animal in which MPTP administration would have effects similar to those observed in the monkey and human. Not surprisingly, MPTP was administered to several small animals, many of them rather uncommon and rarely used as experimental animals. Being interested in animal models of Parkinsonism as well as in understanding how neurotoxic agents exert their actions, we also tried MPTP in several different small animal species. We now report that MPTP causes extensive and long-lasting decrements in the neostriatal content of dopamine and its major metabolites in mice. TABLE 1.
MPTP
Neostriatal levels of dopamine and its metabolites after the injections of MPTP to mice at 30 mg/kg (0.17 mmoles/kgl. Mice were injected daily the number of times indicated and assays done between 5 and 18 days after the last injections. Data represent the mean value? SD for the number of mice indicated.
Number of Injections
Assay Time (Days)
n
DA
21 +
5
10
+ + +
10
5
+ + +
:o"
;:
20
6
;:
:72
13.8?
DOPAC tIlg/g tissue)
1.6
HVA
4.5_+ 1.5
1.9+
0.4
l.O+
0.3
3
5.1 * 1.2
1.2+
; 7
3.0 * 0.7 2.6? 1.0 2.9* 1.4
1.2+ 0.4 1.1 f 0.3 1.4t 0.3
0.9 f 0.3 0.9 + 0.2 1.0 + 0.1
2 3
2.8 + 0.7 2.6 f 0.8 2.6 + 0.5
1.4* 0.3 1.2+ 0.3 1.4-+ 0.2
0.9 i 0.3 0.9 * 0.1 1.2 + 0.2
711
0.5
712
Preliminary
MATERIALS
Notes
AND METHODS
Male, Swiss-Webster mice (Charles River, CF-Wl weighing between 25 and 35 g were injected intraperitoneallywith MPTP, 30 mg/kg (0.17 mmoles/kgl, in distilled water adjusted to pH 8.5 with dilute hydrochloric acid. The mice received multiple MPTP injections, 24 hours apart and levels of dopamine and its metabolites in the neostriatum were assayed at various time periods after the last injection. The neostriata were weighed and homogenized in 0.1 M perchloric acid containing dihydroxybenzylamine(DHBA) as an internal standard, and centrifuged at 27,000 x g for 15 min. The supernatant was used for assays of dopamine, serotonin and their metabolites (Mayer and Shoup, 1983). A Bioanalytical Systems LC-304T liquid chromatograph connected to a Kipp and Zonen dual pen recorder was used for all assays; the operating potential was 750 mv. RESULTS Control levels of dopamine and its metabolites are presented in Table 1. After five injections of MPTP at 30 mg/kg, with assays ten days after the last injection, there was a substantial decrement in levels of dopamine, DOPAC and HVA compared to control values (Table 11. After ten or twenty injections of MPTP, with assays done between 5 and 18 days after the last injection, the content of dopamine and its metabolites was markedly reduced from control values. Under these conditions, there was approximately an 80% loss of dopamine, a 70% loss of DOPAC and a 50% loss of HVA. DISCUSSION The data of the present study demonstrate that MPTP administration causes a large decrement in the neostriatal level of dopamine and of its metabolites in male Swiss-Webstermice. In other experiments, MPTP administration brought about decrements in neostriatal dopamine and its metabolites in other strains of mice including CF-1, CD-1 and C-57BL. The data indicate that this effect in mice is long-lasting. There was no apparent recovery up to 18 days after 10 injections or up to 17 days after 20 injections (Table 1). In these same experiments, there was no obvious difference in levels of neostriatal serotonin or 5-hydroxyindoleaceticacid between control mice and MPTP-treated mice. The actions of MPTP are thus specific in mice and similar to those previously seen in monkeys. In experiments parallel to those presented in Table 1, we injected rats, guinea pigs and gerbils with the same 30 mg/kg dose of MPTP. In these three species, there was no apparent decrement in the content of neostriatal dopamine, DOPAC and HVA with up to ten injections of MPTP. Thus under identical conditions, the mouse is clearly more sensitive to the neurotoxic effects of MPTP than the rat, guinea pig or gerbil.
In conclusion, we report that MPTP causes a large and apparently permanent depletion in levels of striatal dopamine and its metabolites in mice. In other experiments, MPTP administration caused a long-lasting decrement in the capacity of synaptosomal preparations prepared from mouse neostriatum to take up 3H-dopamine. Moreover, MPTP administration caused a severe loss of nerve cells in the zona compacta of the substantia nigra. The mouse should prove to be a convenient and relatively inexpensive small animal with which to study the metabolism of MPTP. We expect that the mouse will be valuable for investigators to explore in depth the mechanism by which MPTP selectively destroys the nigrostriatal dopaminergic system. Further, it provides a model of a highly selective neuronal system degeneration which may yield clues to the pathogenesis of human Parkinson's disease. ACKNOWLEDGEMENTS Supported in part by a grant from the USPHS and by a grant from the United Parkinson Foundation. REFERENCES Burns, R.S., Chiueh, C.C., Markey, S.P., Ebert, M.H., Jacobowitz, D.M., Kopin, I.J. (1983). A primate model of parkinsonism: Selective destruction of dopaminergic neurons in the pars compacta of the substantia nigra by N-methyl-4-phenyl-1,2,3,6tetrahydropyridine. Proc. Natl. Acad. Sci. U.S.A. -80:4546-4550. Chiueh, C.C., Markey, S.P., Burns, R.S., Johannessen, J., Jacobowitz, D.M., Kopin, I.J. N-methyl-4-phenyl-1,2,3,6_tetrahydropyridine, a parkinsonian syndrome (1983). causing agent in man and monkey, produces different effects in guinea pig and rat. The Pharmacologist,-25:131.
Preliminary
Notes
713
Davis, G.C., William, A.C., Markey, S.P., Ebert, M.H., Caine, E.D., Reicher, C.M., Kopin, I.J. (1979). Chronic parkinsonism secondary to intravenous injection of meperidine Psych. Res. 1:249-254. analogues. Langston, J.W., Ballard, P., Tetrud, J.W., Irwin. I. (1983). humans due to a product of meperidine-analog synthesis.
Chronic parkinsonism in Science, 219:979-980. --
Mayer, G.S., Shoup, R.E. (1983). Simultaneous multiple electrode liquid chromatographIC-electrochemical assay for catecholamines, indoleamines and metabolites in brain J. Chromatog. -255:533-544. tissue.