Estrogen interacts with the IGF-1 system to protect nigrostriatal dopamine and maintain motoric behavior after 6-hydroxdopamine lesions

The most prominent neurochemical hallmark of Parkinson's disease (PD) is the loss of nigrostriatal dopamine (DA). Animal models of PD have concentrated on depleting DA and therapies have focused on maintaining or restoring DA. Within this context estrogen protects against 6-hydroxdopamine (6-OHDA) and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) lesions of the nigrostriatal DA pathway. Present studies tested the hypothesis that neuroprotective estrogen actions involve activation of the insulin-like growth factor-1 (IGF-1) system. Ovariectomized rats were treated with either a single subcutaneous injection of 17beta-estradiol benzoate or centrally or peripherally IGF-1. All rats were infused unilaterally with 6-OHDA into the medial forebrain bundle (MFB) to lesion the nigrostriatal DA pathway. Tyrosine hydroxylase (TH) immunocytochemistry confirmed that rats injected with 6-OHDA had a massive loss of TH immunoreactivity in both the ipsilateral substantia nigra compacta (60% loss) and the striatum (>95% loss) compared to the contralateral side. Loss of TH immunoreactivity was correlated with loss of asymmetric forelimb movements, a behavioral assay for motor deficits. Pretreatment with estrogen or IGF-1 significantly prevented 6-OHDA-induced loss of substantia nigra compacta neurons (20% loss) and TH immunoreactivity in DA fibers in the striatum (<20% loss) and prevented the loss of asymmetric forelimb use. Blockage of IGF-1 receptors by intracerebroventricular JB-1, an IGF-1 receptor antagonist, attenuated both estrogen and IGF-1 neuroprotection of nigrostriatal DA neurons and motor behavior. These findings suggest that IGF-1 and estrogen acting through the IGF-1 system may be critical for neuroprotective effects of estrogen on nigrostriatal DA neurons in this model of PD.     

rAAV-mediated nigral human parkin over-expression partially ameliorates motor deficits via enhanced dopamine neurotransmission in a rat model of Parkinson's disease

We hypothesized that over-expressing the E3 ligase, parkin, whose functional loss leads to Parkinson's disease, in the nigrostriatal tract might be protective in the unilateral 6-hydroxydopamine (6-OHDA) rat lesion model. Recombinant adeno-associated virus (rAAV) encoding human parkin or green fluorescent protein (GFP) was injected into the rat substantia nigra 6 weeks prior to a four-site striatal 6-OHDA lesion. Vector-mediated parkin over-expression significantly ameliorated motor deficits as measured by amphetamine-induced rotational behavior and spontaneous behavior in the cylinder test but forelimb akinesia as assessed by the stepping test was unaffected. rAAV-mediated human parkin was expressed in the nigrostriatal tract, the substantia pars reticulata, and the subthalamic nucleus. However, in lesioned animals, there was no difference between nigral parkin and GFP-transduction on lesion-induced striatal tyrosine hydroxylase (TH) innervation or nigral TH positive surviving neurons. A second lesion experiment was performed to determine if striatal dopamine (DA) neurotransmission was enhanced as measured biochemically. In this second group of parkin and GFP treated rats, behavioral improvement was again observed. In addition, striatal TH and DA levels were slightly increased in the parkin-transduced group. In a third experiment, we evaluated parkin and GFP transduced rats 6 weeks after vector injection without DA depletion. When challenged with amphetamine, parkin treated rats tended to display asymmetries biased away from the treated hemisphere. Nigral parkin over-expression induced increases in both striatal TH and DA levels. Therefore, while parkin over-expression exerted no protective effect on the nigrostriatal DA system, parkin appeared to enhance the efficiency of nigrostriatal DA transmission in intact nigral DA neurons likely due to the observed increases in TH.     

Aging of the nigrostriatal system in the squirrel monkey

Increasing incidence of Parkinson's disease with advancing age suggests that age-related processes predispose the nigrostriatal dopaminergic system to neurodegeneration. Several hypotheses concerning the effects of aging on nigrostriatal neurons were assessed in this study using a non-human primate model. First, we examined the possibility that the total number of dopaminergic neurons decline in the substantia nigra as a function of age. Stereological counting based on both tyrosine hydroxylase immunoreactivity (TH-ir) and neuromelanin (NM) content revealed no difference in cell number between young, middle-aged and old squirrel monkeys. We then determined whether advancing age changed the relative proportion of neurons characterized by 1) TH-ir in the absence of NM, 2) the presence of both TH-ir and NM, or 3) NM without TH-ir. Indeed, a progressive age-related depletion of TH only cells was paralleled by an increase in NM only neurons. The possibility that these changes could underlie a functional impairment of the nigrostriatal system was supported by striatal dopamine measurements showing a decrease in older monkeys. Finally, we tested the hypotheses that aging may enhance cell vulnerability to injury and that different dopaminergic subpopulations display varying degrees of susceptibility. When monkeys were exposed to the neurotoxicant 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, cell loss was markedly more pronounced in older animals, and the ranking of vulnerability was TH only < TH/NM < NM only cells. The data indicate that, even in the absence of an overall neuronal loss, changes in the characteristics of dopaminergic cells reflect functional deficits and increased vulnerability to injury with age. NM content appears to be an important marker of these age-related effects.     

Transient striatal GLT-1 blockade increases EAAC1 expression, glutamate reuptake, and decreases tyrosine hydroxylase phosphorylation at ser(19)

Three glutamate transporters, GLT-1, GLAST, and EAAC1, are expressed in striatum. GLT-1 and, to a lesser extent, GLAST are thought to play a primary role in glutamate reuptake and mitigate excitoxicity. Progressive tyrosine hydroxylase (TH) loss seen in Parkinson's disease (PD) is associated with increased extracellular glutamate. Glutamate receptor antagonists reduce nigrostriatal loss in PD models. These observations suggest that excess synaptic glutamate contributes to nigrostriatal neuron loss seen in PD. Decreased GLT-1 expression occurs in neurodegenerative disease and PD models, suggesting decreased GLT-1-mediated glutamate reuptake contributes to excitotoxicity. To determine how transient GLT-1 blockade affects glutamate reuptake dynamics and a Ca(2+)-dependent process in nigrostriatal terminals, ser(19) phosphorylation of TH, the GLT-1 inhibitor dihydrokainic acid (DHK) was delivered unilaterally to striatum in vivo and glutamate reuptake was quantified ex vivo in crude synaptosomes 3h later. Ca(2+)-influx is associated with excitotoxic conditions. The phosphorylation of TH at ser(19) is Ca(2+)-dependent, and a change resulting from GLT-1 blockade may signify the potential for excitotoxicity to nigrostriatal neurons. Synaptosomes from DHK infused striatum had a 43% increase in glutamate reuptake in conjunction with decreased ser(19) TH phosphorylation. Using a novel GLAST inhibitor and DHK, we determined that the GLAST-mediated component of increased glutamate reuptake increased 3-fold with no change in GLAST or GLT-1 protein expression. However, GLT-1 blockade increased EAAC1 protein expression ~20%. Taken together, these results suggest that GLT-1 blockade produces a transient increase in GLAST-mediated reuptake and EAAC1 expression coupled with reduced ser(19) TH phosphorylation. These responses could represent an endogenous defense against excitotoxicity to the nigrostriatal pathway.     

Allopregnanolone increases the number of dopaminergic neurons in substantia nigra of a triple transgenic mouse model of Alzheimer's disease

More than a third of Alzheimer's disease (AD) patients show nigrostriatal pathway disturbances, resulting in akinesia (inability to initiate movement) and bradykinesia (slowness of movement). The high prevalence of this dysfunction of dopaminergic neuron in the nigrostriatal pathway in AD suggests that the risk factors for AD appear also significant risk factors for substantia nigra pars compacta (SNpc) lesions. Previously, we have demonstrated that allopregnanolone (APα) promotes neurogenesis and improves the cognitive function in a triple transgenic mouse model of AD (3xTgAD). In this study, we sought to exam 1) the SNpc lesions in 3xTgAD mice and 2) the impact of APα on promoting the regeneration of new dopaminergic neurons in SNpc of the 3xTgAD mice. The number of Nissl-stained total neurons, tyrosine hydroxylase (TH) positive neurons, and BrdU/TH double positive newly formed neurons were analyzed with unbiased stereology. In the SNpc of 3xTgAD mice, TH positive neurons was 47+- 18 % (p = 0.007), total neurons was 62 +-11.6 % (p = 0.016), of those in the SNpc of non-Tg mice, respectively. APα treatment increased the TH positive neurons in the SNpc of 3xTgAD mice to 93.2 +- 18.5 (p = 0.021 vs. 3xTgAD vehicle) and the total neurons to 84.9+- 6.6 (p = 0.046 vs. 3xTgAD vehicle) of non-Tg mice. These findings indicate that there is a loss of neurons, specifically the TH positive neurons in SNpc of 3xTgAD mice, and that APα reverses the lesion in SNpc of 3xTgAD by increasing the formation of new TH neurons.     

Glial cell line-derived neurotrophic factor prevents death,but not reductions in tyrosine hydroxylase,of injured nigrostriatal neurons in adult rats

Glial cell line-derived neurotrophic factor (GDNF) promotes survival of mesencephalic dopaminergic neurons in vitro and when injected locally into the brains of lesioned adult animals. Here, we show that GDNF (3 μg per day and higher) can promote the survival of all (retrogradely labeled) axotomized nigrostriatal dopaminergic neurons of adult rats when continuously infused for 2 weeks close to the substantia nigra, compared to only ∼30% survival with control infusions. Based on our previous observations, GDNF was as potent as ciliary neurotrophic factor and neurotrophin-4 and approximately five to ten times more potent than brain-derived neurotrophic factor and was most effective in promoting survival. GDNF prevented neuronal death induced by 6-hydroxydopamine to a lesser extent than after axotomy. GDNF treatments begun 1 week after axotomy could maintain those neurons that had not yet died. When a 2 week GDNF treatment was interrupted, most of the GDNF-rescued neurons died over the following 2 weeks. This suggests that longer trophic factor treatments or nigrostriatal connections are needed to achieve permanent survival. Measurements of tyrosine hydroxylase (TH) immunoreactivity of the rescued neuronal cell bodies suggest that GDNF cannot prevent the lesion-induced loss of this rate-limiting enzyme for dopamine synthesis. In fact, GDNF induced a decrease in TH in normal animals, suggesting an active down-regulation of TH synthesis. Levels of TH immunoreactivity were recovered between 7 and 14 days after withdrawal of a 2 week GDNF infusion, in the neurons that survived axotomy. These results may have implications for developing new treatment strategies for Parkinson's disease. J. Comp. Neurol. 388:484–494, 1997. © 1997 Wiley-Liss, Inc.     

GPA protects the nigrostriatal dopamine system by enhancing mitochondrial function

Guanidinopropionic acid (GPA) increases AMPK activity, mitochondrial function and biogenesis in muscle and improves physiological function, for example during aging. Mitochondrial dysfunction is a major contributor to the pathogenesis of Parkinson's disease. Here we tested whether GPA prevents neurodegeneration of the nigrostriatal dopamine system in MPTP-treated mice. Mice were fed a diet of 1% GPA or normal chow for 4 weeks and then treated with either MPTP or saline. Indices of nigrostriatal function were examined by HPLC, immunohistochemistry, stereology, electron microscopy and mitochondrial respiration. MPTP intoxication decreased TH neurons in the SNpc of normal chow-fed mice; however GPA-fed mice remarkably exhibited no loss of TH neurons in the SNpc. MPTP caused a decrease in striatal dopamine of both normal chow- and GPA-fed mice, although this effect was significantly attenuated in GPA-fed mice. GPA-fed mice showed increased AMPK activity, mitochondrial respiration and mitochondrial number in nigrostriatal TH neurons, suggesting that the neuroprotective effects of GPA involved AMPK-dependent increases in mitochondrial function and biogenesis. MPTP treatment produced a decrease in mitochondrial number and volume in normal chow-fed mice but not GPA-fed mice. Our results show the neuroprotective properties of GPA in a mouse model of Parkinson's disease are partially mediated by AMPK and mitochondrial function. Mitochondrial dysfunction is a common problem in neurodegeneration and thus GPA may slow disease progression in other models of neurodegeneration.     

Reversible changes in the activities and amounts of tyrosine hydroxylase in dopamine neurons of the substantia nigra in response to axonal injury as studied by immunochemical and immunocytochemical methods

The effects of unilateral electrolytic lesions of axons of nigrostriatal dopamine neurons of the rat brain on the activity and amount of the neurotransmitter synthesizing enzyme tyrosine hydroxylase (TH) in the axon terminals in the striatum and in the parent cell bodies in the substantia nigra (A9 group) were examined. Lesions in posterolateral hypothalamus damaging axons close to cell bodies resulted, by 72 h, in a permanent anterograde reduction of TH activity to 20% of control in the ipsilateral striatum. By immunochemical titration and immunocytochemical localization with a specific antibody to TH, the reduction was demonstrated as due to loss of enzyme protein. In the reactive A9 cell bodies TH activity was increased to 175% of control by 24 h followed by a gradual and permanent fall to 40% of control by day 14. The retrograde reduction of TH was due to a loss of enzyme protein in turn reflecting retrograde cell death. Striatal lesions of moderate size resulted in a fall in the activity and amount of TH in A9 substantia nigra cell bodies by day 7 to 60–65% of control, with full recovery by day 21. We conclude that the retrograde reaction in central dopamine neurons is dependent upon the proximity of the lesion to the parent cell body: proximal lesions result in retrograde cell death; distal lesions result in a reversible retrograde reduction in the amount and activity of TH. The findings confirm previous evidence that: (a) a reduced accumulation of neurotransmitter synthesizing enzymes is a biochemical concomitant of the retrograde reaction in central neurons; (b) that intrinsic neurons of the CNS may undergo a retrograde response which is reversible. The reduction of TH may indicate a reordering of protein biosynthesis favoring production of protein required for axonal regeneration at the expense of those involved in neurotransmission.     

Microsomal epoxide hydrolase deletion enhances tyrosine hydroxylase phosphorylation in mice after MPTP treatment

Parkinson's disease (PD) is the most prevalent neurodegenerative movement disorder. Epidemiological studies have suggested most cases of PD are linked to environmental risk factors. Microsomal epoxide hydrolase (mEH) is a conserved enzyme that catalyzes hydrolysis of a large number of epoxide intermediates such as drugs and epoxides of environmental toxins. We hypothesize that changes in mEH are involved in the pathogenesis of PD by modulating the vulnerability of dopaminergic neurons to environmental stress. Herein we reported that acute treatment with the neurotoxin MPTP (1-methyl-4-phemyl-1,2,3,6-tetrahydropyridine) markedly increased the mEH immunoreactivity in the nigrostriatal system of C57BL/6 mice. Next, mEH knockout (KO) mice were used, and we found that tyrosine hydroxylase (TH)-positive cell loss was significantly lower in the substantia nigra of mEH KO mice compared with wild-type (WT) mice after MPTP treatment. The mean dopamine turnover ratios were significantly increased in MPTP-treated mEH KO mice compared with WT. In addition, TH is the rate-limiting enzyme for dopamine biosynthesis, and its activity is mainly regulated by TH phosphorylation at Ser-31 (pSer31) and Ser-40 (pSer40). Double immunofluorescence showed that both pSer31 and pSer40 are completely colocalized in total TH-positive cells. However, immunoblotting confirmed that there was a significantly higher level of pSer31 in mEH-KO mice when compared with WT mice after MPTP, and no marked differences among TH and its phosphorylation levels occurred after saline injection. These data suggested that mEH deficiency facilitates TH phosphorylation in the nigrostriatal dopamine system, which may be associated with an increased resistance of dopaminergic neurons to environmental toxins.     

Sequential administration of GDNF into the substantia nigra and striatum promotes dopamine neuron survival and axonal sprouting but not striatal reinnervation or functional recovery in the partial 6-OHDA lesion model

Glial cell line-derived neurotrophic factor (GDNF) has prominent survival-promoting effects on lesioned nigrostriatal dopamine neurons, but understanding of the conditions under which functional recovery can be obtained remains to be acquired. We report here the time course of nigrostriatal axon degeneration in the partial lesion model of Parkinson's disease and the morphological and functional effects of sequential administration of GDNF in the substantia nigra (SN) and striatum during the first 5 weeks postlesion. By 1 day postlesion, the nigrostriatal axons had retracted back to the level of the caudal globus pallidus. Over the next 6 days axonal retraction progressed down to the SN, and during the following 7 weeks 74% of tyrosine hydroxylase-positive (TH(+)) and 84% of retrogradely labeled nigral neurons were lost, with a more pronounced loss in the rostral part of the SN. GDNF administration protected 70 and 72% of the nigral TH(+) and retrogradely labeled cell bodies, respectively, but did not prevent the die-back of the lesioned nigrostriatal axons. Although clear signs of sprouting were observed close to the injection site in the striatum as well as in the globus pallidus, the overall DA innervation of the striatum [as measured by [(3)H]-N-[1-(2-benzo(b)thiopenyl)cyclohexyl]piperidine-binding autoradiography] was not improved by the GDNF treatment. Moreover, the lesion-induced deficits in forelimb akinesia and drug-induced rotation were not attenuated. We conclude that functional recovery in the partial lesion model depends not only on preservation of the nigral cell bodies, but more critically on the ability of GDNF to promote significant reinnervation of the denervated striatum.     

Modification of dopaminergic markers expression in the striatum by neonatal exposure to glutamate during development

Monosodium l-glutamate (MSG) was administered subcutaneously to male neonatal rats, and the effect on developmental profile of tyrosine hydroxylase (TH), D1, D2 receptors, and dopamine (DA) transporter expression in the striatum was examined using Western blot. In addition, TH-immunopositive neurons at substantia nigra (SN) were also examined. MSG treatment (4mg/g of body weight, administered on postnatal days 1, 3, 5, and 7) resulted in a reduction of D1 and D2 receptor expression from 30 days of age and persisted to adulthood (120 days of age), while DA transporter expression was significantly reduced from 14 days of age to adulthood. TH immunopositive neurons at SN showed a significant reduction, as well as TH expression on postnatal days 10, 30, 60, and 120 at striatum was reduced. No changes of TH were observed at 14 days of age. Results indicate that an over-stimulation of the glutamatergic system by neonatal exposure to a high glutamate concentration induces a partial loss in TH-positive neurons in the SN and an important reduction in dopaminergic markers expression in the striatum, suggesting that early excitotoxicity could contribute to developmental alterations in the nigrostriatal pathway, which may be associated with various disorders of the basal ganglia.     

A brief overview of tyrosine hydroxylase and α-synuclein in the Parkinsonian brain

Parkinson's disease (PD) is associated with neurodegeneration of the nigrostriatal tract and is accompanied with loss of tyrosine hydroxylase (TH) and dopamine (DA). Development of neuroprotective strategies targeting PD is often undermined by lack of proper understanding of processes contributing to the pathology. In this mini review we have tried to briefly outline the involvement of TH and α-synuclein in PD. Aberrant expression of α-synuclein is toxic to dopaminergic neurons. It interacts with ubiquitin-proteasomal processing system, implicated in oxidative injury and mitochondrial dysfunction which ultimately induce neurodegenration and cell death. The contributions of DJ-1 in TH regulation have also been discussed. Brain specific TH expression with the combined use of the pegylated immunoliposome (PILs) gene transfer technology and brain specific promoters as a new approach to treat PD has also been included.     

Paradoxical sleep deprivation modulates tyrosine hydroxylase expression in the nigrostriatal pathway and attenuates motor deficits induced by dopaminergic depletion

The nigrostriatal pathway is very likely involved in sleep regulation, considering the occurrence and high prevalence of sleep-related disorders in patients with Parkinson's disease. Indeed, dopaminergic neurons in the ventral tegmental area were recently shown to fire in bursts during paradoxical sleep (PS), but little is known about the activity of the nigrostriatal dopamine (DA) cells in relation to PS. In view of that we hypothesized that paradoxical sleep deprivation (PSD) may play a relevant role in nigrostriatal tyrosine hydroxylase (TH) expression and, subsequently, in sleep rebound. The present study was designed to determine the effects of PSD in the nigrostriatal pathway in mice by means of neurochemical and behavioral approaches. Intraperitoneal reserpine (1 mg/kg) associated to α-methyl-p-tyrosine (αMT) (250 mg/kg) to produce catecholamine depletion, or rotenone (10 mg/kg) to increase striatal DA turnover were injected 30 min before the 24 h of PSD. Catalepsy and open-field tests indicated that motor deficits induced by reserpine-αMT were counteracted by PSD, which, in contrast, potentiated the motor impairment induced by rotenone. Besides, PSD produced down-regulation on TH expression within the substantia nigra pars compacta and striatum, without affecting the number or the optical density of dopaminergic neurons present in the respective areas. Interestingly, PSD potentiated the downregulation of TH expression in the substantia nigra pars compacta and striatum induced by the co-administration of reserpine-αMT. These results reinforce the notion of a strong participation of DA in PS, as a consequence of the modulation of TH protein expression in the nigrostriatal pathway.     

Delayed increase of tyrosine hydroxylase expression in rat nigrostriatal system after traumatic brain injury

Tyrosine hydroxylase (TH) is the key enzyme for synthesizing dopamine (DA) in dopaminergic neurons and its terminals. Emerging experimental and clinical evidence support the hypothesis of a disturbance in dopamine neurotransmission following traumatic brain injury (TBI). However, the effect of controlled cortical impact (CCI) injury on TH in the nigrostriatal system is currently unknown. To determine if there is an alteration in TH after CCI injury, we examined TH levels at 1 day, 7 days, and 28 days post-injury by utilizing a commercially available antibody specific to TH. Rats were anesthetized and surgically prepared for CCI injury (4 m/s, 3.2 mm) or sham surgery. Injured (N=6) and sham animals (N=6) were sacrificed and coronally sectioned (35 microm thick) through the striatum and substantia nigra (SN) for immunohistochemistry. Additionally, semiquantitative measurements of TH protein in striatal and SN homogenates from injured (N=6) and sham (N=6) rats sacrificed at the appropriate time post-surgery were assessed using Western blot analysis. TH protein is bilaterally increased at 28 days post-injury in nigrostriatal system revealed by immunohistochemistry in injured rats compared to sham controls. Western blot analysis confirms the findings of immunohistochemistry in both striatum and SN. We speculate that the increase in TH in the nigrostriatal system may reflect a compensatory response of dopaminergic neurons to upregulate their synthesizing capacity and a delayed increase in the efficiency of DA neurotransmission after TBI.     

Relationships between various behavioural abnormalities and nigrostriatal dopamine depletion in the unilateral 6-OHDA-lesioned rat

While rotational asymmetry is used as a characteristic behavioural sign of striatal dopamine (DA) loss in unilateral animal models of Parkinson's disease (PD), there is relatively little analysis of how other common behavioural deficits relate to nigrostriatal DA depletion. We analysed the relationships between several deficits induced by unilateral 6-OHDA lesions and striatal neurochemistry, as well as neuronal loss in the dopaminergic substantia nigra (SN). Behaviour was evaluated from before until 6 weeks after surgery and abnormalities appeared in body axis, head position and sensorimotor performance as well as apomorphine-induced rotation. As expected, rotational behaviour correlated with striatal DA loss and not with other striatal neurotransmitters measured. Similar observations were found for sensorimotor deficits ('disengage task'). Both deficits were observed in rats with >70% loss of TH+ nigral neurons and >80% loss of striatal DA. Additional postural abnormalities appeared with mean losses of 87% of nigral DA neurons and 97% striatal DA, consistent with observations in patients with advanced PD. The data show that the repertoire of behavioural abnormalities manifested by hemiparkinsonian rats relate directly to the degree of nigrostriatal DA loss and, therefore, mimic features of PD. Analysis of such behaviours are relevant for chronic therapeutic studies targeting PD.     

Levodopa-induced dyskinesia in MPTP-treated macaques is not dependent on the extent and pattern of nigrostrial lesioning

The extent of nigrostriatal denervation is presumed to play a role in the genesis of levodopa-induced dyskinesia. Yet some parkinsonian patients who have been treated over a long period do not develop dyskinesia, raising the possibility that the pattern of denervation is as important as the extent of lesioning as a risk factor. Here we study the extent and pattern of nigrostriatal denervation in a homogeneous population of parkinsonian macaque monkeys chronically treated with levodopa. Based on the characteristics of the lesioning, non-dyskinetic animals could not be differentiated from those with dyskinesia. Indeed, the number of tyrosine-hydroxylase (TH)-immunopositive neurons in the substantia nigra pars compacta, striatal dopamine transporter (DAT) binding and TH immunostaining, as well as the overall TH striatal content measured by Western blotting were identical. Moreover, the patterns of lesioning assessed by a detailed analysis of the TH- and DAT-immunopositive striatal fibers were comparable in all functional quadrants and at all rostro-caudal levels considered. These data indicate that neither the extent nor the pattern of nigrostriatal lesioning are sufficient to explain the occurrence of levodopa-induced dyskinesia.     

EphB1 null mice exhibit neuronal loss in substantia nigra pars reticulata and spontaneous locomotor hyperactivity

The molecular mechanisms that regulate basal ganglia development are largely unknown. Eph receptor tyrosine kinases are potential participants in this process as they regulate development of other CNS regions and are expressed in basal ganglia nuclei, such as the substantia nigra (SN) and striatum. To address the role of Eph receptors in the development of these nuclei, we analysed anatomical changes in the SN and striatum of mice with null mutations for EphB1. These mice express beta-galactosidase as a marker for cells normally expressing EphB1. In situ hybridization data and a direct comparison of SN neurons expressing tyrosine hydroxylase (TH) and/or the beta-gal marker for EphB1 revealed that EphB1 is not expressed in TH+ neurons of pars compacta (SNc), but is restricted to neurons in pars reticulata (SNr). Consistent with this, we find that EphB1 null mice exhibit a significant decrease in the volume and number of neurons (40% decrease) in SNr, whereas the volume and number of TH+ neurons in SNc is not significantly affected nor are there changes in the distribution of nigrostriatal dopamine neurons. Although EphB1 is expressed in the striatum, EphB1-/- mice exhibit no significant changes in striatal volume and TH fiber density, and have no obvious alterations in striatal patch/matrix organization. Behavioral evaluation of EphB1 null mice in an open-field environment revealed that these mice exhibited spontaneous locomotor hyperactivity. These results suggest that EphB1 is necessary for the proper formation of SNr, and that neuronal loss in SNr is associated with altered locomotor functions.     

Environmental risk factors and Parkinson's disease: selective degeneration of nigral dopaminergic neurons caused by the herbicide paraquat

Environmental toxicants and, in particular, pesticides have been implicated as risk factors in Parkinson's disease (PD). The purpose of this study was to determine if selective nigrostriatal degeneration could be reproduced by systemic exposure of mice to the widely used herbicide paraquat. Repeated intraperitoneal paraquat injections killed dopaminergic neurons in the substantia nigra (SN) pars compacta, as assessed by stereological counting of tyrosine hydroxylase (TH)-immunoreactive and Nissl-stained neurons. This cell loss was dose- and age-dependent. Several lines of evidence indicated selective vulnerability of dopaminergic neurons to paraquat. The number of GABAergic cells was not decreased in the SN pars reticulata, and counting of Nissl-stained neurons in the hippocampus did not reveal any change in paraquat-treated mice. Degenerating cell bodies were observed by silver staining, but only in the SN pars compacta, and glial response was present in the ventral mesencephalon but not in the frontal cortex and cerebellum. No significant depletion of striatal dopamine followed paraquat administration. On the other hand, enhanced dopamine synthesis was suggested by an increase in TH activity. These findings unequivocally show that selective dopaminergic degeneration, one of the pathological hallmarks of PD, is also a characteristic of paraquat neurotoxicity. The apparent discrepancy between pathological (i.e., neurodegeneration) and neurochemical (i.e., lack of significant dopamine loss) effects represents another important feature of this paraquat model and is probably a reflection of compensatory mechanisms by which neurons that survive damage are capable of restoring neurotransmitter tissue levels.