Differential effects of carboxyfullerene on MPP+/MPTP-induced neurotoxicity

The effects of carboxyfullerene on a well-known neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and its active metabolite 1-methyl-4-phenyl-pyridinium (MPP+) were investigated. In chloral hydrate-anesthetized rats, cytosolic cytochrome c was elevated in the infused substantia nigra 4 h after an intranigral infusion of MPP+. Five days after local application of MPP+, lipid peroxidation (LP) was elevated in the infused substantia nigra. Furthermore, dopamine content and tyrosine hydroxylase (TH)-positive axons were reduced in the ipsilateral striatum. Concomitant intranigral infusion of carboxyfullerene abolished the elevation in cytochrome c and oxidative injuries induced by MPP+. In contrast, systemic application of carboxyfullerene did not prevent neurotoxicity induced by intraperitoneal injection of MPTP. In mice, systemic administration of MPTP induced a dose-dependent depletion in striatal dopamine content. Simultaneous injection of carboxyfullerene (10 mg/kg) actually potentiated MPTP-induced reduction in striatal dopamine content. Furthermore, systemic administration of carboxyfullerene (30 mg/kg) caused death in the MPTP-treated mice. An increase in the striatal MPP+ level and reduction in hepatic P450 level were observed in the carboxyfullerene co-treated mice. These data showed that systemic application of carboxyfullerene appears to potentiate MPTP-induced neurotoxicity while local carboxyfullerene has been suggested as a neuroprotective agent. Furthermore, an increase in striatal MPP+ level may contribute to the potentiation by carboxyfullerene of MPTP-induced neurotoxicity.     

Trace dosages of the neurotoxins MPTP and MPP+ may affect brain dopamine in vivo.

The present study has examined the effects of systemically administered MPTP and MPP+ upon striatal DA and Dopac of C57 mice, also treated concurrently with either saline or reserpine. MPTP followed by saline did not affect DA level but decreased that of Dopac only at 5.0 mg/kg and higher dosages. The potency of MPTP affecting DA increased greatly when the neurotoxicant was followed by either 5.0 or 10.0 mg/kg reserpine; MPTP at 0.10 mg/kg and higher dosages significantly reversed the DA depleting effects of reserpine. MPP+ (1.0 or 10.0 mg/kg) with saline did not affect either DA or Dopac. In contrast, MPP+ at 0.10 mg/kg and higher dosages, when followed by 10.0 mg/kg reserpine, dose-dependently enhanced the DA depleting effects of reserpine. In agreement with the earlier results obtained in vitro, the present study indicates that MPTP administration at trace level dosages may lead to an inhibition of MAO in vivo. The effect of systemically given MPP+ on DA, however, appears to be more complex in nature, conceivably comprised of actions at the striatal neurones including the intraneuronal vesicles and, possibly, at the substantia nigra which may affect striatum in turn. That MPP+ may have reached brain areas in these experiments is also indicated by the observation of a significant striatal level of 3H-MPP+ after its systemic administration. In conclusion, irrespective of MPTP and MPP+ action mechanisms, trace levels of these neurotoxicants appear to affect brain dopamine neurons.     

Acute peripheral catecholaminergic changes in rat after MPTP and MPP+ treatment

The effects of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and its main metabolite 1-methyl-4-phenylpyridinium ion (MPP+) on the peripheral catecholaminergic system of the rat were investigated. MPTP and MPP+ injections (20 mg/kg i.p.) caused a marked acute depletion of heart noradrenaline, up to 75% twelve hours after the administration, and a decrease of adrenal gland adrenaline. The time-course of the effect of MPTP and MPP+ is reported, together with a decrease in the tyrosine hydroxylase activity after MPTP treatment, more evident in the adrenal glands. Pargyline (50 mg/kg i.p.) is not able to prevent such a neurotoxic peripheral effect.     

Prevention of the Nigrostriatal Toxicity of 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine by Inhibitors of 3,4-Dihydroxyphenylethylamine Transport

The 3,4-dihydroxyphenylethylamine (DA, dopamine) uptake inhibitors GBR 13,069, amfonelic acid, WIN-35,065-2, WIN-35,428, nomifensine, mazindol, cocaine, McN-5908, McN-5847, and McN-5292 were effective in preventing [3H]DA and [3H]1-methyl-4-phenylpyridinium (MPP+) uptake in rat and mouse neostriatal tissue slices. These DA uptake inhibitors also were effective in attenuating the MPP+-induced release of [3H]DA in vitro. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) administration to mice (6 X 25 mg/kg i.p.) resulted in a large (70-80%) decrement in neostriatal DA. WIN-35,428 (5 mg/kg), GBR 13,069 (10 mg/kg), McN-5292 (5 mg/kg), McN-5908 (2 mg/kg), and amfonelic acid (2 mg/kg), when administered intraperitoneally 30 min prior to each MPTP injection, fully protected against MPTP-induced neostriatal damage. Other DA uptake inhibitors showed partial protection in vivo at the doses selected. Desmethylimipramine did not prevent [3H]MPP+ uptake or MPP+-induced release of [3H]DA in vitro, and did not protect against MPTP neurotoxicity in vivo. These results support the hypothesis put forth previously by others that the active uptake of MPP+ by dopaminergic neurons is necessary for toxicity.     

Tissue concentrations of MPTP and MPP+ in relation to catecholamine depletion after the oral or subcutaneous administration of MPTP to mice

One hour after MPTP was given to mice at a dose of 30 mg/kg s.c., its concentration in tissues varied in the order kidney greater than liver greater than lung greater than brain greater than heart. When the same dose of MPTP was given orally, concentrations in most tissues were much lower at 1 hr than after s.c. administration, although the MPTP concentration in liver was only slightly lower. The concentrations of MPP+ (a metabolite of MPTP) at 1 hr were as high or higher than those of MPTP in all tissues except kidney, and MPP+ disappeared from the various tissues with half-lives from 3-20 hrs. The highest concentrations of MPP+, both absolute and relative to MPTP, were in heart. After oral administration of MPTP, no MPP+ was found in brain, and MPP+ concentrations in other tissues were lower than those after s.c. dosing. The depletion of heart norepinephrine was similar after MPTP administration by either route of administration even though MPTP and MPP+ concentrations in heart were lower after oral administration, suggesting that other metabolites of MPTP might also contribute to heart norepinephrine depletion.     

Comparative toxicity and antioxidant activity of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine and its monoamine oxidase B-generated metabolites in isolated hepatocytes and liver microsomes

MPTP (1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine) is converted by monoamine oxidase B to its putative toxic metabolite MPP+ (1-methyl-4-phenylpyridinium ion) via MPDP+ (1-methyl-4-phenyl-2,3-dihydropyridinium ion). Both the parent compound and these two major metabolites were toxic to isolated rat hepatocytes with MPDP+ being the most toxic and MPP+ the least effective. MPP+ produced a slight increase in lipid peroxidation above control levels in hepatocytes, while both MPTP and MPDP+ showed antioxidant effects. The latter two compounds also protected against chemically and nonchemically induced lipid peroxidation in rat liver microsomes. MPDP+ was effective at much lower concentrations than MPTP. MPDP+ was also markedly more efficient when NADPH was used to induce microsomal lipid peroxidation. Lipid peroxidation as a consequence of oxygen radical generation is therefore unlikely to be involved in MPTP toxicity in vitro and the rationale of using chain-breaking antioxidants as protective agents in vivo needs a more careful evaluation.     

Interaction between nicotine and MPTP/MPP+ in rat brain endothelial cells

To examine the interaction between nicotine and MPTP/MPP+ in the blood-brain barrier, cellular uptake of MPTP and MPP+ was studied in the presence of nicotine and several compounds, including MPTP/MPP+ analogs and a specific inhibitor of organic cation transporter (OCT) in an adult rat brain microvascular endothelial cell line (ARBEC). The kinetic properties of the uptake of MPTP, MPP+, and nicotine were also examined. In addition, a microdialysis study was performed to evaluate the in vivo effect of nicotine (i.p.) on extracellular levels of MPTP and MPP+ in the brain after intravenous administration of MPTP. The results showed that uptake of MPTP, MPP+, and nicotine was partly mediated by a carrier system that was sensitive to decynium22, a specific OCT inhibitor. RT-PCR showed the presence of OCT1 mRNA in ARBEC. Capacity for uptake of MPTP and nicotine was much higher than that for MPP+ (Km and Vm values of 10.94+/-1.44 microM and 0.049+/-0.007 pmol/mg s, respectively, for MPP+, compared to values of 35.75+/-0.85 microM and 40.95+/-3.56 pmol/mg s for MPTP and 25.29+/-6.44 microM and 51.15+/-14.18 pmol/mg s for nicotine). In addition, nicotine competitively inhibited the uptake of both MPTP and MPP+, with inhibition constants (Ki) of 328 microM and 210 microM, respectively. In vivo microdialysis results showed that nicotine significantly reduced brain extracellular levels of MPTP in the first 30 min (507.4+/-8.5 ng/ml vs. 637.9+/-30.8 ng/ml with and without nicotine pre-treatment, respectively), but did not have significant effect on those of MPP+. In conclusion, nicotine can inhibit in vitro cellular uptake and in vivo transfer of MPTP across the blood-brain barrier, which can be mediated by multiple pathways including OCT1.     

Urinary excretion of MPTP and its primary metabolites in mice

Mice were injected with single doses of MPTP (15 mg/kg, s.c.) containing one microCi of [3H]methyl-MPTP. Approximately 42% of the total injected [3H] was detected in the urine within 3 hours after drug administration. The early urine samples were analyzed using high pressure liquid chromatography. MPTP N-oxide was identified as a major metabolite, with trace amounts of MPP+ and MPTP also detected. The urinary volume and excretion of MPTP metabolites were inhibited by pretreating the animals with probenecid (250 mg/kg, i.p.). These results indicate that large amounts of injected MPTP are rapidly metabolized in the periphery by liver enzymes to form MPTP N-oxide.     

Protective effect of harmalol and harmaline on MPTP neurotoxicity in the mouse and dopamine-induced damage of brain mitochondria and PC12 cells

The present study elucidated the protective effect of beta-carbolines (harmaline, harmalol, and harmine) on oxidative neuronal damage. MPTP treatment increased activities of total superoxide dismutase, catalase, and glutathione peroxidase and levels of malondialdehyde and carbonyls in the basal ganglia, diencephalon plus midbrain of brain compared with control mouse brain. Coadministration of harmalol (48 mg/kg) attenuated the MPTP effect on the enzyme activities and formation of tissue peroxidation products. Harmaline, harmalol, and harmine attenuated both the 500 microM MPP(+)-induced inhibition of electron flow and membrane potential formation and the 100 microM dopamine-induced thiol oxidation and carbonyl formation in mitochondria. The scavenging action of beta-carbolines on hydroxyl radicals was represented by inhibition of 2-deoxy-D-ribose degradation. Harmaline and harmalol (100 microM) attenuated 200 microM dopamine-induced viability loss in PC12 cells. The beta-carbolines (50 microM) attenuated 50 microM dopamine-induced apoptosis in PC12 cells. The compounds alone did not exhibit significant cytotoxic effects. The results indicate that beta-carbolines attenuate brain damage in mice treated with MPTP and MPP(+)-induced mitochondrial damage. The compounds may prevent dopamine-induced mitochondrial damage and PC12 cell death through a scavenging action on reactive oxygen species and inhibition of monoamine oxidase and thiol oxidation.     

Prolonged Alterations in Canine Striatal Dopamine Metabolism Following Subtoxic Doses of l-Methyl-4-Phenyl- 1,2,3,6-Tetrahydropyridine (MPTP) and 4''-Amino-MPTP Are Linked to the Persistence of Pyridinium Metabolites

Single toxic doses of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP).HCl (2.5 mg/kg i.v.) and 4'-amino-MPTP.2HCl (22.5 mg/kg) induce loss of striatal dopamine (DA) and tyrosine hydroxylase (TH) activity and of nigral DA neurons in the dog. To examine the subacute neurochemical changes induced by low doses of MPTP and 4'-amino-MPTP, dose-response studies of these compounds were carried out in the dog, using 6- and 3-week survival times for these two compounds, respectively. Low single doses of MPTP (1.0, 0.5, and 0.1 mg/kg i.v.) and 4'-amino-MPTP (15, 7.5, and 3.75 mg/kg i.v.) did not cause depletion of canine striatal DA or TH or a loss of nigral neurons. However, levels of the DA metabolites 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) were decreased in a dose-related fashion, with significant loss of DOPAC being evident 6 weeks after the lowest administered dose of MPTP and 3 weeks after 4'-amino-MPTP. This selective loss of DA metabolites following nontoxic doses of MPTP and 4'-amino-MPTP led to a shift in the ratio of DA to DOPAC or HVA, which was characteristic for each compound. The measurement of striatal 1-methyl-4-phenylpyridinium (MPP+) and 4'-amino-MPP+ levels revealed that high concentrations (up to 150 microM) persist in the striatum for weeks following administration of a single nontoxic dose of MPTP or 4'-amino-MPTP. A causal relationship between the striatal concentration of MPP+ or 4'-amino-MPP+ and the change in DA metabolism as reflected in the DA/DOPAC ratio is suggested by a significant correlation between these measures. It is suggested that presynaptic sequestration and retention of MPP+ and 4'-amino-MPP+ by striatal DA terminals result in the inhibition of the monoamine oxidase contained within these terminals.     

Behavioral and biochemical changes following acute administration of MPTP and MPP+

The acute effects of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and 1-methyl-4-phenylpyridinium ion (MPP+) on mouse locomotor activity and striatal dopamine (DA) and 5-hydroxytryptamine (5-HT) levels were investigated. A single dose of either MPTP (10-30 mg/kg, i.p.) or MPP+ (5-20 ug/mouse, i.c.v.) decreased locomotor activity 10-40 min after injection: this locomotor effect was significantly suppressed by either pretreatment with nomifensine or 1-deprenyl alone, or by the combination of desmethylimipramine and 6-hydroxydopamine. Pretreatment with clorgyline did not suppress this behavior and a single dose of haloperidol enhanced the effect. The striatal levels of DA, 3-methoxytyramine and 5-HT increased in parallel with the decrease in locomotor activity caused by MPTP or MPP+. In contrast, levels of 3,4-dihydroxyphenylacetic acid, homovanillic acid and 5-hydroxyindoleacetic acid were decreased by injection of either MPTP or MPP+. Possible mechanism(s) of the behavioral and biochemical changes caused by the acute actions of MPTP and MPP+ with respect to their neurotoxic effects on the nigrostriatal DA system are discussed.     

Effect of dopaminergic neurotoxin MPTP/MPP+ on coenzyme Q content

Coenzyme Q10, an endogenous lipophilic antioxidant, plays an indispensable role in ATP synthesis. The therapeutic value of coenzyme Q10 in Parkinson's disease and other neurodegenerative disorders is still being tested and the preliminary results are promising. The 1-methyl-4-phenyl-1, 2, 3, 6 tetrahydropyridine (MPTP)-treated mouse is a valid and accepted animal model for Parkinson's disease. 1-methyl-4-phenylpyridinium (MPP(+)) is an active toxic metabolite of MPTP. MPP(+) and MPTP are known to induce oxidative stress and mitochondrial dysfunction. However, the effect of MPP(+) and MPTP on coenzyme Q is not clearly understood. The present study investigated the in vitro and in vivo effect of MPP(+) and MPTP on coenzyme Q content. Coenzyme Q content was measured using HPLC-UV detection methods. In the in vitro studies, MPP(+) (0-50 microM) was incubated with SH-SY5Y human neuroblastoma cells and NG-108-15 (mouse/rat, neuroblastomaxglioma hybrid) cells. MPP(+) concentration dependently increased coenzyme Q10 content in SH-SY5Y cells. In NG-108-15 cells, MPP(+) concentration dependently increased both coenzyme Q9 and Q10 content. In the in vivo study, mice were administered with MPTP (30 mg/kg, twice 16 h apart) and sacrificed one week after the last administration. Administration of MPTP to mice significantly increased coenzyme Q9 and coenzyme Q10 levels in the nigrostriatal tract. However, MPTP did not affect the coenzyme Q content in the cerebellum, cortex and pons. This study demonstrated that MPP(+)/MPTP significantly affected the coenzyme Q content in the SH-SY5Y and NG-108 cells and in the mouse nigrostriatal tract.     

Recommended safe practices for using the neurotoxin MPTP in animal experiments

The metabolism and distribution of the parkinsonian syndrome inducing neurotoxin MPTP has been studied in non-human primates and mice housed in controlled environmental chambers. 14C6-MPTP was prepared and injected at concentrations normally employed for lesioning experiments (30 mg/kg in mice, 0.3 mg/kg in monkeys). All interior surfaces of the chambers which could be reached by animals or their excreta were contaminated with radiolabeled metabolites. Vapor born unmetabolized MPTP was negligible, although significant amounts of MPTP were found in the excreta of mice (less than or equal to 15% injected dose) and small amounts from rhesus monkeys (less than 2%). Procedures to minimize contact with animal fur, bedding and excreta should protect investigators working with MPTP over extended periods. Permanganate oxidation effectively detoxifies solutions of MPTP. MPTP, MPP+, common synthetic intermediates, and the products of MPTP's oxidation are not mutagenic as measured by a Salmonella-microsome assay.     

Diethyldithiocarbamate Potentiates the Neurotoxicity of In Vivo l-Methyl-4-Phenyl-1, 2, 3, 6-Tetrahydropyridine and of In Vitro 1-Methyl-4-Phenylpyridinium

Diethyldithiocarbamic acid (DDC) potentiates in vivo neurotoxicity of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and in vitro neurotoxicity of 1-methyl-4-phenylpyridinium (MPP+). Male C57B1/6 mice were given two or five injections of MPTP (30 mg/kg i.p.) preceded 0.5 h by DDC (400 mg/kg i.p.). The mice were tested for catalepsy, akinesia, or motor activity during and after the period of dosing. Striatal and hippocampal tissues were obtained at 2 and 7 days following the last injection and evaluated for dopamine and norepinephrine levels, respectively. These same tissues were also analyzed for the levels of glial fibrillary acidic protein (GFAP), an astrocyte-localized protein known to increase in response to neural injury. Pretreatment with DDC potentiated the effect of MPTP in striatum and resulted in substantially greater dopamine depletion, as well as a more pronounced elevation in GFAP. In hippocampus, the levels of norepinephrine and GFAP were not different from controls in mice receiving only MPTP, but pretreatment with DDC resulted in a sustained depletion of norepinephrine and an elevation of GFAP, suggesting that damage was extended to this brain area by the combined treatment. Mice receiving MPTP preceded by DDC also demonstrated a more profound, but reversible, catalepsy and akinesia compared to those receiving MPTP alone. Systemically administered MPP+ decreased heart norepinephrine, but did not alter the striatal levels of dopamine or GFAP, and pretreatment with DDC did not alter these effects, but did increase lethality. DDC is known to increase brain levels of MPP+ after MPTP, but our data indicate that this is not due to a movement of peripherally generated MPP+ into CNS.(ABSTRACT TRUNCATED AT 250 WORDS)     

Chemical oxidation of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and its in vivo metabolism in rat brain and liver

We investigated in vivo the metabolism of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in the brain and liver of rats 45 min after the systemic administration of 50 mg/kg of the neurotoxin. The metabolites present in brain and liver extracts were identified through multiple analytical methods by comparison to authentic compounds obtained from a number of chemical oxidations of MPTP. Our results indicate the presence of approximately 15% unreacted MPTP and relatively large amounts of both 1-methyl-4-phenylpyridinium (MPP+) and a mixture of three nonpolar lactams: 1-methyl-4-phenyl-5,6-dihydro-2(1H)-pyridinone, 1-methyl-4-phenyl-2(1H)-pyridinone, and a previously unreported metabolite 1-methyl-4-phenyl-2-piperidinone. Whereas MPP+ was more prevalent in the brain than in the liver, the lactam metabolites were more predominant in the liver. The amounts of the N-oxide and N-demethylated metabolites of MPTP were minimal.     

Metabolism of N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine in a rat pheochromocytoma cell line, PC12h

The uptake and metabolism of a neurotoxin, N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) were examined in a rat pheochromocytoma cell line, PC12h. These cells which contain only type A monoamine oxidase (MAO-A) oxidize MPTP into N-methyl-4-phenylpyridinium ion (MPP+). By kinetic analysis, the apparent Km value and the maximal velocity of the MPP+ production are 70.4 +/- 6.5 microM and 38.3 +/- 10.0 pmol/min/mg protein, respectively. After 7 days of culture in the presence of MPTP, the cells could oxidize from 25 to 50% of the MPTP added to the culture medium and could accumulate MPP+. The intracellular concentrations of MPTP were almost the same after 7 days of culture in the presence of MPTP from 10 nM to 100 microM. The cells could survive 7 days after exposure to up to 100 microM MPTP. Tyrosine hydroxylase (TH) and MAO activity were not affected by the presence of MPTP. Dopamine (DA) concentrations and a nonspecific enzyme, beta-galactosidase activity in the cells were not affected by the addition of MPTP. These data show that the uptake and oxidative conversion of MPTP take place in the cells having MAO-A alone, and that the neurotoxicity of MPP+ may not be due directly to its storage in subcellular compartments.     

Effects of MPTP, MPP+ and paraquat on mitochondrial potential and oxidative stress

The effect of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), 1-methyl-4-phenylpyridinium (MPP+) and 1,1-dimethyl-4,4-bipyridinium (paraquat) upon the electrical potential across the plasma and mitochondrial membranes within synaptosomes has been investigated. MPTP selectively depressed plasma membrane potential while MPP+ specifically reduced mitochondrial potential. The structurally similar compound paraquat had no effect on either membrane potential. Enhancement of the lipid peroxidative activity with an Fe-ADP complex depressed both potentials. Paraquat effected increased peroxidative activity in brain homogenates that was less pronounced than that due to Fe-ADP. MPTP reduced basal but stimulated Fe-ADP enhanced peroxidation. The mechanisms underlying the toxicity of MPP+ are likely to differ from those of paraquat, primarily involving impaired mitochondrial function rather than increased oxidative stress.     

Genetic ablation of tumor necrosis factor-alpha (TNF-alpha) and pharmacological inhibition of TNF-synthesis attenuates MPTP toxicity in mouse striatum

The impact of pro-inflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) in the pathology of Parkinson's disease (PD) and in MPTP neurotoxicity remains unclear. Here, male TNF-alpha (-/-) deficient mice and C57bL/6 mice were treated with MPTP (4 x 15 mg/kg, 24 h intervals) and in one series, thalidomide was administered to inhibit TNF-alpha synthesis. Real-time RT-PCR revealed that the striatal mRNA levels of TNF-alpha, of the astrocytic marker glial fibrillary acidic protein (GFAP) and of the marker for activated microglia, macrophage antigen complex-1 (MAC-1), were significantly enhanced after MPTP administration. Thalidomide (50 mg/kg, p.o.) partly protected against the MPTP-induced dopamine (DA) depletion, and TNF-alpha (-/-) mice showed a significant attenuation of striatal DA and DA metabolite loss as well as striatal tyrosine hydroxylase (TH) fiber density, but no difference in nigral TH and DA transporter immunoreactivity. TNF-alpha deficient mice suffered a lower mortality (10%) compared to the high mortality (75%) seen in wild-type mice after acute MPTP treatment (4 x 20 mg/kg, 2 h interval). HPLC measurement of MPP(+) levels revealed no differences in TNF-alpha (-/-), wild-type and thalidomide treated mice. This study demonstrates that TNF-alpha is involved in MPTP toxicity and that inhibition of TNF-alpha response may be a promising target for extending beyond symptomatic treatment and developing anti-parkinsonian drugs for the treatment of the inflammatory processes in PD.     

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) does not destroy nigrostriatal neurons in the scorbutic guinea pig

Guinea pigs were injected subcutaneously with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in maximal tolerated doses (8 mg/kg, once daily) for 10 or 15 days. No neurological effects were noted, other than sedation and hypotonia lasting a few hours after each injection, either in animals maintained on normal diet or in animals fed an ascorbate-deficient diet and rendered severely scorbutic. Subsequent chemical analyses of the striatum showed no evidence of lasting damage to nigrostriatal dopaminergic neurons in MPTP treated guinea pigs on normal diet, and minimal evidence of permanent damage to these neurons in scorbutic animals. MPTP was undetectable in the urine of MPTP-treated animals, although a metabolite, presumably 1-methyl-4-phenylpyridinium ion (MPP+) was regularly present in urine. The relative lack of neurotoxicity of MPTP in the guinea pig remains unexplained. This species clearly is not a suitable small animal for MPTP-induced parkinsonism.     

1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine Metabolism and l-Methyl-4-Phenylpyridinium Uptake in Dissociated Cell Cultures from the Embryonic Mesencephalon

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), a contaminant found in a synthetic illicit drug, can elicit in humans and monkeys a severe extrapyramidal syndrome similar to Parkinson's disease. It also induces alterations of the dopamine (DA) pathways in rodents. MPTP neurotoxicity requires its enzymatic transformation into 1-methyl-4-phenylpyridinium (MPP+) by monoamine oxidase followed by its concentration into target cells, the DA neurons. Here, we show that mesencephalic glial cells from the mouse embryo can take up MPTP in vitro, transform it into MPP+, and release it into the culture medium. MPTP is not taken up by neurons from either the mesencephalon or the striatum in vitro (8 days in serum-free conditions). However, mesencephalic neurons in culture revealed a high-affinity uptake mechanism for the metabolite MPP+, similar to that for DA. The affinity (Km) for DA uptake is fivefold higher than that for MPP+ (0.2 and 1.1 microM, respectively), whereas the number of uptake sites for MPP+ is double (Vmax = 25 and 55 pmol/mg of protein/min for DA and MPP+, respectively). Mazindol, a DA uptake inhibitor, blocks the uptake of DA and MPP+ equally well under these conditions. Moreover, by competition experiments, the two molecules appear to use the same carrier(s) to enter DA neurons. Small concentrations of MPP+ are also taken up by striatal neurons in vitro. The amount taken up represented less than 10% of the MPP+ uptake in mesencephalic neurons. Depolarization induced by veratridine released comparable proportions of labeled DA and MPP+ from mesencephalic cultures.(ABSTRACT TRUNCATED AT 250 WORDS)