MPTP对小鼠帕金森病造模条件探讨

目的观察不同剂量1-甲基-4-苯基-1,2,3,6-四氢吡啶(MPTP)对小鼠行为学及脑黑质酪氨酸羟化酶、纹状体多巴胺含量的影响,探讨MPTP致帕金森病(Parkinson′s disease,PD)样小鼠模型的最佳条件。方法C57BL小鼠分别给与MPTP不同剂量处理,测定各组小鼠爬竿时间检测动物运动协调性,应用免疫组化方法和高效液相法观察不同模型组多巴胺能神经元的变化。结果模型组各组均出现不同程度爬竿时间延长,酪氨酸羟化酶阳性细胞数减少和多巴胺含量减少。结论MPTP处理可造成小鼠的帕金森病样症状,在此种动物模型中,应根据科研目的选择MPTP的应用剂量和给药途径。     

Estrogen Alters MPTP-Induced Neurotoxicity in Female Mice: Effects on Striatal Dopamine Concentrations and Release

Abstract: The effects of estrogen on MPTP-induced neurotoxicity of the nigrostriatal dopaminergic system were examined in C57Bl and CD-1 mice. Ovariectomized mice with and without estrogen were treated with MPTP or its vehicle. The effects of these treatments on striatal dopamine concentrations and l -DOPA-stimulated dopamine and l -3,4-dihydroxyphenylacetic acid (DOPAC) release in vitro were determined. Dopamine concentrations of C57Bl mice receiving estrogen before MPTP were significantly greater than those of non-estrogen-treated MPTP mice as well as estrogen-treated mice receiving the MPTP vehicle. Dopamine concentrations of the CD-1 mice did not differ with these treatments. l -DOPA-evoked dopamine release values of estrogen-treated C57Bl mice were significantly increased compared with non-estrogen-treated mice. No such differences were observed in the MPTP-treated C57Bl mice. DOPAC release rates were similar to that of dopamine in these C57Bl mice. In the CD-1 mice estrogen also produced a significant increase in l -DOPA-evoked dopamine release; however, this response was unaltered by MPTP treatment. A significant increase in l -DOPA-evoked DOPAC output was obtained only for estrogen-treated CD-1 mice. Both strains show very similar responses to the estrogen treatment, but differential responses of dopamine release to l -DOPA between the C57Bl and CD-1 mice were obtained with regard to the interactive effects of estrogen and MPTP. Our results suggest that in addition to its role as modulator, estrogen may also function as a neuroprotectant against MPTP neurotoxicity of the nigrostriatal dopaminergic system in the C57Bl mouse.     

Involvement of cytochrome P450 2E1 in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced mouse model of Parkinson's disease

Elucidation of the biochemical steps leading to the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced degeneration of the nigrostriatal dopamine (DA) pathway has provided new clues to the pathophysiology of Parkinson's disease. In line with the enhancement of MPTP toxicity by diethyldithiocarbamate (DDC), here we demonstrate how other cytochrome P450 (CYP) 2E1 inhibitors, such as diallyl sulphide (DAS) and phenylethylisothiocyanate (PIC), also potentiate the selective DA neurone degeneration in C57/bl mice. In addition, we show that CYP 2E1 is present in the brain and in the basal ganglia of this mouse strain, as measured by RT-PCR, western blot analysis and immunohistochemistry. A kinetic analysis of MPTP and its metabolites, by means of the microdialysis technique in the striatum, indicates that no detoxification metabolic pathway is affected by any of these inhibitors. This does not rule out, however, that an undetected detoxification pathway involving CYP 2E1 is operating. In order to provide direct evidence for this isozyme involvement, CYP 2E1 knockout mice were challenged with MPTP or the combined treatment. Here we show that these transgenic mice have a low sensitivity to MPTP alone, similar to their wild-type counterparts, suggesting that it is likely that transgenic mice compensate for the missing enzyme. However, DDC pretreatment completely fails to enhance MPTP toxicity in CYP 2E1 knockout mice, whereas this enhancement is regularly present in wild-type animals. This study indicates that the occurrence of CYP 2E1 in C57/bl mouse brain is relevant to MPTP toxicity, and suggests that this isozyme may have a detoxificant role related to the efflux transporter of the toxin.     

Neurochemical and Histochemical Characterization of Neurotoxic Effects of 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine on Brain Catecholamine Neurones in the Mouse

Systemic administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) caused a rapid and long-lasting reduction of both 3,4-dihydroxyphenylalanine (dopamine, DA) and noradrenaline (NA) in mouse brain, as observed histo- and neurochemically. The depleting effects were more pronounced after repeated MPTP administration and the most marked reductions were observed after 2 X 50 mg MPTP/kg s.c., when DA in striatum and NA in frontal cortex were reduced by greater than 90% 1 week after MPTP. Mice with such catecholamine depletions were markedly sedated and almost completely immobilized. The behavioural syndrome after MPTP resembled that seen after reserpine, a monoamine-depleting drug. MPTP also caused a long-lasting reduction of catecholamine uptake in striatal DA and cortical NA nerve terminals and reduced tyrosine hydroxylase activity in these regions. There was no evidence that MPTP caused any marked DA and NA cell body death. MPTP given acutely transiently elevated serotonin levels. The results are compatible with a neurotoxic action of MPTP on both DA and NA nerve terminals. The nigro-striatal DA and the locus coeruleus NA neurone systems appeared to be most susceptible. Synthesis and utilization of residual striatal DA and cortical NA were increased, as often observed in partially denervated monoamine-innervated brain regions. Both DA and NA showed a gradual recovery, which took months to become complete and may have been related to a regrowth of catecholamine nerve terminals.     

Epidermal Growth Factor Enhances Striatal Dopaminergic Parameters in the 1-Methyl-4-Phenyl-l, 2, 3, 6-Tetrahydropyridine-Treated Mouse

Intracerebroventricular infusion of epidermal growth factor (EGF) into mice with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced degeneration of dopaminergic nigrostriatal neurons partially enhanced the content of dopamine (DA) and 3,4-dihydroxyphenylacetic acid as well as the activity of tyrosine hydroxylase in the striatum. EGF also enhanced these parameters in control, unlesioned animals. Neurotrophic activity also was observed in embryonic mesencephalic cultures, where EGF enhanced DA uptake after a lesion with the neurotoxic metabolite of MPTP, 1-methyl-4-phenylpyridinium ion. Our in vivo and in vitro studies suggest that EGF may be a neurotrophic factor for dopaminergic neurons, or may act indirectly by inducing the release of a dopaminergic trophic factor from other cells.     

Involvement of monoamine oxidase inhibition in neuroprotective and neurorestorative effects of Ginkgo biloba extract against MPTP-induced nigrostriatal dopaminergic toxicity in C57 mice.

The present study investigated the neuroprotective and neurorestorative effects of Ginkgo biloba extract (EGb 761) and its two components ginkgolides A (BN52020) and B (BN52021) in mice. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) (30 mg/kg/d i.p. for six days) significantly reduced striatal dopamine (DA) levels in C57 mice measured by high performance liquid chromatography with electrochemical detection (HPLC-EC). When C57 mice were pretreated with EGb 761 (20, 50, 100 mg/kg/d i.p.) for 7 days and then treated with the same extract 30 min before MPTP injection for 6 days, the neurotoxic effect of MPTP was antagonized in a dose-dependent fashion. Similar treatment with ginkgolides A and B (5, 10, 50 mg/kg/d i.p.) showed no protective effect. When C57 mice were treated with EGb 761 (50 mg/kg/d i.p.) after MPTP-lesion, the recovery of striatal dopamine (DA) levels was accelerated. However, similar treatment with ginkgolides A or B (10 mg/kg/d i.p.) did not show any effect. EGb 761, but not ginkgolides A and B, nonselectively inhibited mouse brain MAO activity in vitro (IC50 = 36.45 +/- 1.56 microg/ml) tested by an improved fluorimetric assay. The results demonstrate that EGb 761 administered before or after MPTP treatment effectively protects against MPTP-induced nigrostriatal dopaminergic neurotoxicity and that the inhibitory effect of EGb 761 on brain MAO may be involved in its neuroprotective effect.     

Partial protection from the dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) by Pro-Leu-Gly-NH2(PLG; MIF-1)

The administration of MPTP to man and monkey has been shown to cause a neurotoxic effect on the nigrostriatal dopamine system. MPTP was injected in C57-BL black mice, 36 mg per kg for 7 days, which resulted in permanent reduction of dopamine and serotonin levels in the striatum. In the mice pretreated with PLG, although the striatal dopamine level was also reduced, mean dopamine and serotonin levels were significantly higher than in mice given MPTP alone. It is concluded that PLG could protect at least partially the neurotoxic effect of MPTP.     

Presynaptic CB(1) cannabinoid receptors control frontocortical serotonin and glutamate release--species differences

Both the serotonergic and endocannabinoid systems modulate frontocortical glutamate release; thus they are well positioned to participate in the pathogenesis of psychiatric disorders. With the help of fluorescent and confocal microscopy, we localized the CB(1) cannabinoid receptor (CB(1)R) in VGLUT1- and 2- (i.e. glutamatergic) and serotonin transporter- (i.e. serotonergic) -positive fibers and nerve terminals in the mouse and rat frontal cortex. CB(1)R activation by the synthetic agonists, WIN55212-2 (1 μM) and R-methanandamide (1 μM) inhibited the simultaneously measured evoked Ca(2+)-dependent release of [(14)C]glutamate and [(3)H]serotonin from frontocortical nerve terminals of Wistar rats, in a fashion sensitive to the CB(1)R antagonists, O-2050 (1 μM) and LY320135 (5 μM). CB(1)R agonists also inhibited the evoked release of [(14)C]glutamate in C57BL/6J mice in a reversible fashion upon washout. Interestingly, the evoked release of [(14)C]glutamate and [(3)H]serotonin was significantly greater in the CB(1)R knockout CD-1 mice. Furthermore, CB(1)R binding experiments revealed similar frontocortical CB(1)R density in the rat and the CD-1 mouse. Still, the evoked release of [(3)H]serotonin was modulated by neither CB(1)R agonists nor antagonists in wild-type CD-1 or C57BL/6J mice. Altogether, this is the first study to demonstrate functional presynaptic CB(1)Rs in frontocortical glutamatergic and serotonergic terminals, revealing species differences.     

Short-term manganese pretreatment partially protects against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine neurotoxicity

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is a neurotoxin that induces parkinsonism in human and non-human primates. Its mechanism of action is not fully elucidated.Recently, the participation of trace metals, such as manganese, on its neurotoxic action has been postulatted. In this work, we studied the effect of manganese administration on the neurochemical consequences of MPTP neurotoxic action. Male Swiss albino mice were treated with manganese chloride (MnCl₂ ·4H₂O; 0.5 mg/ml or 1.0 mg/ml of drinking water) for 7 days, followed by three MPTP administrations (30 mg/Kg, intraperitoneally). Seven days after the last MPTP administration, mice were sacrificed and dopamine and homovanillic acid contents in corpus striatum were analyzed. Striatal concentration of dopamine was found increased by 60% in mice pretreated with 0.5 mg/ml and 52% in the group treated of 1.0 mg/ml as compared versus animals treated with MPTP only. Hornovanillic acid content in both groups treated with manganese was the same as those in control animals. The results indicate that manganese may interact with MPTP, producing an enhancement of striatal dopamine turnover, as the protective effect of manganese was more pronounced in the metabolite than in the neurotransmitter.     

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) neurotoxicity in mice is enhanced by ethanol or acetaldehyde

Persistent neurochemical changes consistent with parkinsonism have been reported in brains of mice treated with repeated high doses of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). We now report that ethanol or acetaldehyde potentiate MPTP-induced damage to mouse striatum. One hour after the combined treatments (ethanol and MPTP or acetaldehyde and MPTP), the animals exhibited a marked and long-lasting catatonic posture and then returned gradually to apparently normal locomotion. Seven days after MPTP administration, depletion of dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in mouse striatum were further potentiated in the group of animals treated with ethanol. This effect was more evident when the treatment was repeated twice and was dose-dependent. Acetaldehyde was more potent than ethanol in enhancing MPTP neurotoxicity. A single exposure to acetaldehyde before and during MPTP treatment produced a very consistent fall of DA, DOPAC and HVA but not serotonin (5HT) or 5-hydroxyindoleacetic acid (5HIAA) in the striatum. This suggests that ethanol effects on MPTP neurotoxicity might be related to acetaldehyde formation.     

Effects of Systemic Administration of 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine to Mice on Tyrosine Hydroxylase, l-3,4-Dihydroxyphenylalanine Decarboxylase, Dopamine β-Hydroxylase, and Monoamine Oxidase Activities in the Striatum and Hypothalamus

The effects of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) (30 mg/kg subcutaneously per day for 8 days) to C57BL/6N mice were studied on tyrosine hydroxylase (TH), L-3,4-dihydroxyphenylalanine decarboxylase (DDC), and monoamine oxidase (MAO) activities in the striatum, and TH, DDC, dopamine-beta-hydroxylase (DBH), and MAO activities in the hypothalamus. Treatment with MPTP led to a large decrease in TH activity and a parallel decrease in DDC activity in the striatum, as compared with the saline controls. In contrast, MPTP administration did not cause a decrease of the activities of TH, DDC, and DBH in the hypothalamus. There was also no reduction in MAO activities of striatum and hypothalamus. These data indicate that MPTP administration to mice results in specific degeneration of the dopaminergic nigrostriatal pathway and that DDC in the mouse striatum may mainly be localized in the dopaminergic neurons with TH.     

帕金森病小鼠模型行为学检测方法的比较研究

目的比较目前常用的5种帕金森病(PD)小鼠模型行为学检测方法在PD研究中的作用。方法用MPTP建立C57BL小鼠PD模型,通过行为学检测(自主活动计数、滚轴实验、游泳实验、爬杆实验、悬挂实验)、免疫组织化学和荧光分光光度法,对比5种行为学检测方法的平均数与变异系数,观察MPTP对PD小鼠模型的行为学、黑质多巴胺(DA)神经元和纹状体酪氨酸羟化酶免疫反应阳性(TH-ir)神经纤维以及纹状体DA水平的影响。结果给与MPTP后,小鼠行为学计数降低,爬杆实验未能得到检测结果,悬挂实验变异系数很高,结果有明显的偶然性,滚轴实验结果变异系数中等,平均数呈现一定的上升趋势,自主活动计数中移动与站立和游泳实验的平均数则呈现明显的下降趋势,变异系数很低,而黑质DA神经元数目减少约58%,纹状体TH-ir神经纤维密度减低,纹状体DA水平明显降低约88%,两组相比差异有显著性(P<0.01)。结论MPTP所致的C57BL小鼠的神经病理、生化改变与PD患者近似,自主活动计数和游泳实验优于其他行为学检测方法。     

Treatment with GM1 Ganglioside Restores Striatal Dopamine in the 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine-Treated Mouse

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), 30 mg/kg i.p. daily for 7 days, was administered to mice. This dosage regimen resulted in an approximately 50% reduction of striatal dopamine (DA) level. Chronic administration of GM1 ganglioside (II3NeuAc-GgOse Cer), beginning between 1 to 4 days after terminating MPTP dosing, resulted in partial restoration of the striatal DA level. From dose- and time-response studies, it appeared that 30 mg/kg i.p. of GM1 administered daily for approximately 23 days resulted in an approximately 80% restoration of the DA level and complete restoration of the 3,4-dihydroxyphenylacetic acid (DOPAC) content. This dosage of GM1 also restored the turnover rate of DA in the striatum to near normal. Discontinuing GM1 treatment resulted in a fall of DA and DOPAC levels to values found in mice treated with MPTP alone. There was no evidence for regeneration of nerve terminal amine reuptake in the GM1-treated mice as evaluated by DA uptake into synaptosomes. Our biochemical findings in animals suggest that early GM1 ganglioside treatment of individuals with degenerative diseases of dopaminergic nigrostriatal neurons might be fruitful.     

过表达PINK1抵抗鱼藤酮引起多巴胺神经元损伤的研究

目的:明确在C57BL/6小鼠纹状体过表达野生型的人源帕金森相关蛋白PINK1能否减轻由侧脑室注射鱼藤酮引起多巴胺神经元损伤。方法:通过向C57BL/6小鼠(雄性,7周龄,18~20g)左侧纹状体中注射带有GFP人源野生型PINK1及突变体PINK1^G309D的慢病毒包装颗粒,两周后向小鼠左侧侧脑室中定位注射鱼藤酮,通过蛋白质印迹,免疫组化和行为学的方法检测PINK1对鱼藤酮引起多巴胺神经元损伤的影响。结果:蛋白质印迹和免疫组化的实验都证明了在C57BL/6小鼠纹状体过表达野生型的PINK1对于鱼藤酮引起多巴胺能神经元的减少有明显的抑制作用(P<0.01),但对鱼藤酮引起的行为学损伤没有明显的改善作用。     

Effects of 1-Methyl-4-Phenyl-1,2,5,6-Tetrahydropyridine and Related Compounds on the Uptake of [3H]3,4-Dihydroxyphenylethylamine and [3H]5-Hydroxytryptamine in Neostriatal Synaptosomal Preparations

1-Methyl-4-phenyl-1,2,5,6-tetrahydropyridine (MPTP) is known to cause a destruction of the dopaminergic nigrostriatal pathway in certain animal species including mice. MPTP and some structurally related analogs were tested in vitro for their capacity to inhibit the uptake of [3H]3,4-dihydroxyphenylethylamine-([3H]DA), [3H]5-hydroxytryptamine ([3H]5-HT), and [3H]gamma-aminobutyric acid [( 3H]GABA) in mouse neostriatal synaptosomal preparations. MPTP was a very potent inhibitor of [3H]5-HT uptake (IC50 value 0.14 microM), a moderate inhibitor of [3H]DA uptake (IC50 value 2.6 microM), and a very weak inhibitor of [3H]GABA uptake (no significant inhibition observed at 10 microM MPTP). In other experiments, MPTP caused some release of previously accumulated [3H]DA and [3H]5-HT, but in each case MPTP was considerably better as an uptake inhibitor than as a releasing agent. The 4-electron oxidation product of MPTP, i.e., 1-methyl-4-phenyl-pyridinium iodide (MPP+), was a very potent inhibitor of [3H]DA uptake (IC50 value 0.45 microM) and of [3H]5-HT uptake (IC50 value 0.78 microM) but MPP+ was a very weak inhibitor of [3H]GABA uptake. These data may have relevance to the neurotoxic actions of MPTP.     

Genotype modulates testosterone-dependent activity and reactivity in male mice

Adult castration significantly reduced the homecage locomotor activity of both inbred C57BL/6J and DBA/2J and outbred Rockland-Swiss (R-S) male mice. Castrated C57BL animals exhibited greater reductions in this behavior than did the other genotypes. Locomotor activity in a novel environment (reactivity) was also reduced by castration but only for inbred males. In both test situations, postcastration reductions in ambulation were prevented by implants of testosterone (T)-containing Silastic capsules. Thus, testicular hormones promote activity and reactivity in the male mouse in a genotype-dependent fashion.     

EPC-K1, a Hydroxyl Radical Scavenger,Prevents 6-Hydroxydopamine-Induced Dopamine Depletion in the Mouse Striatum by Up-Regulation of Catalase Activity

We examined the effect of pretreatment with EPC-K1, a potent hydroxyl radical scavenger, on 6-hydroxydopamine (6-OHDA)-induced reduction of dopamine (DA) and its metabolites in the mouse striatum. EPC-K1 was mixed with diet (0.2%, wt/wt) for 1 or 2 weeks, and then 6-OHDA (60 g in 2l of saline solution) was injected intracereberoventricularly. Mice continued to be fed EPC-K1-containing diet for another one week before they were sacrificed. The concentrations of DA and its metabolites in the striatum were measured by high performance liquid chromatography. 6-OHDA reduced the level of DA and its metabolites in the striatum. Pretreatment with EPC-K1 for 2 weeks, but not for 1 week, abrogated the neurotoxic effect of 6-OHDA on striatal concentrations of DA and its metabolites. Measurement of striatal concentrations of thiobarbituric acid reactive substances, glutathione, and malonaldehyde plus 4-hydroxynonenal, and the activities of superoxide dismutase and catalase in EPC-K1 treated mice showed an increase in catalase activity after 2 weeks of such treatment. No other changes in anti-oxidants levels were noted. Our results suggest that EPC-K1 counteracts the neurotoxicity of 6-OHDA by increasing catalase activities.     

Selective stimulation of serotonin2c receptors blocks the enhancement of striatal and accumbal dopamine release induced by nicotine administration

The effects of acute and repeated nicotine administration on the extracellular levels of dopamine (DA) in the corpus striatum and the nucleus accumbens were studied in conscious, freely moving rats by in vivo microdialysis. Acute intraperitoneal (i.p.) injection of nicotine (1 mg/kg) increased DA outflow both in the corpus striatum and the nucleus accumbens. Repeated daily injection of nicotine (1 mg/kg, i.p.) for 10 consecutive days caused a significant increase in basal DA outflow both in the corpus striatum and the nucleus accumbens. Acute challenge with nicotine (1 mg/kg, i.p.) in animals treated repeatedly with this drug enhanced DA extracellular levels in both brain areas. However, the effect of nicotine was potentiated in the nucleus accumbens, but not in the corpus striatum. To test the hypothesis that stimulation of 5-HT (5-hydroxytryptamine, serotonin)(2C) receptors could affect nicotine-induced DA release, the selective 5-HT(2C) receptor agonist RO 60-0175 was used. Pretreatment with RO 60-0175 (1 and 3 mg/kg, i.p.) dose-dependently prevented the enhancement in DA release elicited by acute nicotine in the corpus striatum, but was devoid of any significant effect in the nucleus accumbens. RO 60-0175 (1 and 3 mg/kg, i.p.) dose-dependently reduced the stimulatory effect on striatal and accumbal DA release induced by an acute challenge with nicotine (1 mg/kg, i.p.) in rats treated repeatedly with this alkaloid. However, only the effect of 3 mg/kg RO 60-0175 reached statistical significance. The inhibitory effect of RO 60-0175 on DA release induced by nicotine in the corpus striatum and the nucleus accumbens was completely prevented by SB 242084 (0.5 mg/kg, i.p.) and SB 243213 (0.5 mg/kg, i.p.), two selective antagonists of 5-HT(2C) receptors. It is concluded that selective activation of 5-HT(2C) receptors can block the stimulatory action of nicotine on central DA function, an effect that might be relevant for the reported antiaddictive properties of RO 60-0175.     

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)     

Rat-1 Fibroblasts Engineered with GAD65 and GAD67 cDNAs in Retroviral Vectors Produce and Release GABA

Abstract: The effects of the adenosine A₁ agonist N ₆-cyclohexyladenosine (CHA) on MPTP-induced dopamine (DA) depletion in the striatum of C57BL/6 mice were studied. Twenty hours after a single injection of MPTP (30 mg/kg, s.c.), the toxin caused 62% depletion of striatal DA. CHA (0.2–3 mg/kg, s.c.), when given together with MPTP, prevented the toxin-induced DA depletion in a dose-dependent manner. This protective action was apparently mediated by the A¹ receptors, because this effect was selectively antagonized by pretreating the animals with the A₁ antagonist 8-cyclopentyl-1,3-dipropylxanthine (25 mg/kg, i.p.) but not with the A₂ antagonist 1,3-dipropyl-7-methylxanthine (25 mg/kg, i.p.). When CHA (3 mg/kg) was injected 5 h after MPTP administration, at which point striatal DA levels were already reduced significantly, a rapid and complete recovery of the striatal DA levels occurred. These neurochemical data suggest that the A₁ agonist CHA is potentially useful as a neuroprotective agent against MPTP-induced toxicity.