Interleukin-4 Aggravates LPS-Induced Striatal Neurodegeneration In Vivo via Oxidative Stress and Polarization of Microglia/Macrophages.

The present study investigated the effects of interleukin (IL)-4 on striatal neurons in lipopolysaccharide (LPS)-injected rat striatum in vivo. Either LPS or PBS as a control was unilaterally injected into the striatum, and brain tissues were processed for immunohistochemical and Nissl staining or for hydroethidine histochemistry at the indicated time points after LPS injection. Analysis by NeuN and Nissl immunohistochemical staining showed a significant loss of striatal neurons at 1, 3, and 7 days post LPS. In parallel, IL-4 immunoreactivity was upregulated as early as 1 day, reached a peak at 3 days, and was sustained up to 7 days post LPS. Increased levels of IL-4 immunoreactivity were exclusively detected in microglia/macrophages, but not in neurons nor astrocytes. The neutralizing antibody (NA) for IL-4 significantly protects striatal neurons against LPS-induced neurotoxicity in vivo. Accompanying neuroprotection, IL-4NA inhibited activation of microglia/macrophages, production of reactive oxygen species (ROS), ROS-derived oxidative damage and nitrosative stress, and produced polarization of microglia/macrophages shifted from M1 to M2. These results suggest that endogenous IL-4 expressed in LPS-activated microglia/macrophages contributes to striatal neurodegeneration in which oxidative/nitrosative stress and M1/M2 polarization are implicated.

Connexin 30 deficiency attenuates A2 astrocyte responses and induces severe neurodegeneration in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine hydrochloride Parkinson's disease animal model.

BACKGROUND: The first pathology observed in Parkinson's disease (PD) is 'dying back' of striatal dopaminergic (DA) terminals. Connexin (Cx)30, an astrocytic gap junction protein, is upregulated in the striatum in PD, but its roles in neurodegeneration remain elusive. We investigated Cx30 function in an acute PD model by administering 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) to wild-type (WT) and Cx30 knockout (KO) mice. METHODS: On days 1 and 7 after MPTP administration, we evaluated changes in astrocytic Cx30, Cx43, glial fibrillary acidic protein, and ionised calcium-binding adapter molecule 1 expression by immunostaining and biochemical analysis. Loss of DA neurons was evaluated by tyrosine hydroxylase immunostaining. Gene expression was analysed using A1, A2, pan-reactive astrocyte microarray gene sets, and M1, M2, and M1/M2 mixed microglial microarray gene sets. Real-time PCR and in situ hybridisation were performed to evaluate glial cell-derived neurotrophic factor (Gdnf) and S100a10 expression. Striatal GDNF protein levels were determined by enzyme-linked immunosorbent assay. RESULTS: MPTP treatment induced upregulation of Cx30 and Cx43 levels in the striatum of WT and KO mice. DA neuron loss was accelerated in Cx30 KO compared with WT mice after MPTP administration, despite no change in the striatal concentration of methyl-4-phenylpyridinium+. Astrogliosis in the striatum of Cx30 KO mice was attenuated by MPTP, whereas microglial activation was unaffected. Microarrays of the striatum showed reduced expression of pan-reactive and A2 astrocyte genes after MPTP treatment in Cx30 KO compared with WT mice, while M1, M2, and M1/M2 mixed microglial gene expression did not change. MPTP reduced the number of striatal astrocytes co-expressing Gdnf mRNA and S100ß protein or S100a10 mRNA and S100ß protein and also reduced the level of GDNF in the striatum of Cx30 KO compared with WT mice. CONCLUSIONS: These findings indicate that Cx30 plays critical roles in astrocyte neuroprotection in an MPTP PD model.

Mitochondrial division inhibitor-1 is neuroprotective in the A53T-α-synuclein rat model of Parkinson's disease.

Alpha-synuclein (α-syn) is involved in both familial and sporadic Parkinson's disease (PD). One of the proposed pathogenic mechanisms of α-syn mutations is mitochondrial dysfunction. However, it is not entirely clear the impact of impaired mitochondrial dynamics induced by α-syn on neurodegeneration and whether targeting this pathway has therapeutic potential. In this study we evaluated whether inhibition of mitochondrial fission is neuroprotective against α-syn overexpression in vivo. To accomplish this goal, we overexpressed human A53T-α- synuclein (hA53T-α-syn) in the rat nigrostriatal pathway, with or without treatment using the small molecule Mitochondrial Division Inhibitor-1 (mdivi-1), a putative inhibitor of the mitochondrial fission Dynamin-Related Protein-1 (Drp1). We show here that mdivi-1 reduced neurodegeneration, α-syn aggregates and normalized motor function. Mechanistically, mdivi-1 reduced mitochondrial fragmentation, mitochondrial dysfunction and oxidative stress. These in vivo results support the negative role of mutant α-syn in mitochondrial function and indicate that mdivi-1 has a high therapeutic potential for PD.

Progression of subcortical atrophy and iron deposition in multiple system atrophy: a comparison between clinical subtypes.

Magnetic resonance imaging (MRI) can be useful not only for the diagnosis of multiple system atrophy (MSA) itself, but also to distinguish between different clinical subtypes. This study aimed to investigate whether there are differences in the progression of subcortical atrophy and iron deposition between two variants of MSA. Two serial MRIs at baseline and follow-up were analyzed in eight patients with the parkinsonian variant MSA (MSA-P), nine patients with cerebellar variant MSA (MSA-C), and fifteen patients with Parkinson's disease (PD). The R2* values and volumes were calculated for the selected subcortical structures (caudate nucleus, putamen, globus pallidus, and thalamus) using an automated region-based analysis. In both volume and R2*, a higher rate of progression was identified in MSA-P patients. Volumetric analysis showed significantly more rapid progression of putamen and caudate nucleus in MSA-P than in MSA-C. With regard to R2* changes, a significant increase at follow-up and a higher rate of progression were identified in the putamen of MSA-P group compared to MSA-C and PD groups. This longitudinal study revealed different progression rates of MRI markers between MSA-P and MSA-C. Iron-related degeneration in the putamen may be more specific for MSA-P.

A rat model of striatonigral degeneration generated by simultaneous injection of 6-hydroxydopamine into the medial forebrain bundle and quinolinic acid into the striatum.

A double toxin-double lesion strategy is well-known to generate a rat model of striatonigral degeneration (SND) such as multiple system atrophy-parkinsonian type. However, with this model it is difficult to distinguish SND from Parkinson's disease (PD). In this study, we propose a new rat model of SND, which is generated by simultaneous injection of 6-hydroxydopamine into the medial forebrain bundle and quinolinic acid into the striatum. Stepping tests performed 30 min after intraperitoneal L-dopa administration at 6 weeks post-surgery revealed an L-dopa response in the PD group but not the SND group. Apomorphine-induced rotation tests revealed no rotational bias in the SND group, which persisted for 2 months, but contralateral rotations in the PD group. MicroPET scans revealed glucose hypometabolism and dopamine transporter impairment on the lesioned striatum in the SND group. Tyrosine hydroxylase immunostaining in the SND group revealed that 74.7% of nigral cells on the lesioned side were lost after lesion surgery. These results suggest that the proposed simultaneous double toxin-double lesion method successfully created a rat model of SND that had behavioral outcomes, multitracer microPET evaluation, and histological aspects consistent with SND pathology. This model will be useful for future study of SND.

Sirtuin 2 (SIRT2) enhances 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced nigrostriatal damage via deacetylating forkhead box O3a (Foxo3a) and activating Bim protein.

Sirtuins are NAD-dependent protein deacetylases that were shown to have beneficial effects against age-related diseases. SIRT2 is a strong deacetylase that is highly expressed in brain. It has been associated with neurodegenerative diseases. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is a dopaminergic neurotoxin that replicates most of the clinical features of Parkinson disease (PD) and produces a reliable and reproducible lesion of the nigrostriatal dopaminergic pathway and neurodegeneration after its systemic administration. Chronic administration of MPTP induces lesion via apoptosis. We show here that SIRT2 deacetylates Foxo3a, increases RNA and protein levels of Bim, and as a result, enhances apoptosis in the MPTP model of PD. We also show that neurodegeneration induced by chronic MPTP regimen is prevented by genetic deletion of SIRT2 in mouse. Deletion of SIRT2 leads to the reduction of apoptosis due to an increase in acetylation of Foxo3a and a decrease in Bim levels. We demonstrate that SIRT2 deacetylates Foxo3a, activates Bim, and induces apoptosis only in 1-methyl-4-phenylpyridinium-treated cells. Therefore, designing SIRT2 inhibitors might be helpful to develop effective treatments for PD.

Peripheral inflammation increases the deleterious effect of CNS inflammation on the nigrostriatal dopaminergic system.

Evidence supports the role of inflammation in the development of neurodegenerative diseases. In this work, we are interested in inflammation as a risk factor by itself and not only as a factor contributing to neurodegeneration. We tested the influence of a mild to moderate peripheral inflammation (injection of carrageenan into the paws of rats) on the degeneration of dopaminergic neurons in an animal model based on the intranigral injection of lipopolysaccharide (LPS), a potent inflammatory agent. Overall, the treatment with carrageenan increased the effect of the intranigral injection of LPS on the loss of dopaminergic neurons in the SN along with all the other parameters studied, including: serum levels of the inflammatory markers TNF-α, IL-1ß, IL-6 and C-reactive protein; activation of microglia, expression of proinflammatory cytokines, the adhesion molecule ICAM and the enzyme iNOS, loss of astrocytes and damage to the blood brain barrier (BBB). The possible implication of BBB rupture in the increased loss of dopaminergic neurons has been studied using another Parkinson's disease animal model based on the intraperitoneal injection of rotenone. In this experiment, loss of dopaminergic neurons was also strengthened by carrageenan, without affecting the BBB. In conclusion, our data show that a mild to moderate peripheral inflammation can exacerbate the degeneration of dopaminergic neurons caused by a harmful stimulus.

Evaluation of a multiple system atrophy model in rats using multitracer microPET.

BACKGROUND: A double toxin-double lesion strategy is appropriate for mimicking of striatonigral degeneration. Because knowledge of human pathology is limited, animal models must be well characterized prior to testing of therapeutic approaches to treat multiple system atrophy. In double-toxin animal models, however, reduced contralateral rotation after apomorphine injection is restored within a few weeks via an unknown mechanism; the animals thus revert to PD status. We assessed this phenomenon using multitracer microPET and tissue staining. METHODS: Five adult male Wistar rats received injections of 6-hydroxydopamine (6-OHDA) into the right medial forebrain bundle (MFB), followed 3 weeks later by injections of quinolinic acid (QA) into the ipsilateral striatum. Apomorphine-induced rotation tests were performed 1 week after each injection, and 6 and 10 weeks after QA injection. Rotarod tests were performed weekly after 6-OHDA injection. MSA-p status was characterized by microPET 5 and 10 weeks after QA injection using the tracers 2-deoxy-2-[(18)F]-fluoro-D-glucose ([(18)F]-FDG) and [(18)F]-N-(3-fluoropropyl)-2-carbomethoxy-3-(4-iodophenyl)nortropane ([(18)F]-FP-CIT). Histological changes were evaluated by tyrosine hydroxylase (TH) and cresyl violet staining. RESULTS: The numbers of apomorphine-induced rotations increased contralaterally after 6-OHDA lesions were created, but decreased significantly after QA administration (p = 0.007). Five weeks after QA injection, however, contralateral rotation again increased and persisted for 1 month. Rotarod rotation differed significantly between the intact and PD states (p < 0.05), but not between the PD and MSA-p states. MicroPET revealed glucose hypometabolism and dopamine transporter (DAT) impairment on the lesioned side of the striatum 1 and 2 months after QA lesion surgery. Loss of nigral cells was confirmed by TH immunostaining, and striatal atrophy was observed upon cresyl violet staining. CONCLUSION: Pathological changes consistent with MSA-p can be generated by the double toxin-double lesion method and persist during follow-up. Behavioral tests, such as drug-induced rotation and rotarod tests, are not appropriate for long-term follow-up in the MSA-p model, suggesting the need for development of more appropriate behavioral tests.

Intrastriatal injection of colchicine induces striatonigral degeneration in mice.

Recent studies from environmental toxicology and molecular genetics demonstrate that midbrain dopamine (DA) neurons are particularly vulnerable to microtubule depolymerizing agents, indicating the involvement of microtubule dysfunction in the pathogenesis of Parkinson's disease. Here we show that intrastriatal injection of colchicine (COL), a well-known microtubule disruptor, induced degeneration of striatonigral pathway. Microtubule disruption caused by unilateral injection of COL blocked the retrograde axonal transport of fluorogold previously injected into striatum and induced substantial death of striatal and DA neurons in substantia nigra pars compacta. Furthermore, COL-induced pathologic changes were associated with robust glial reaction, which may be conducive to the degeneration of striatonigral pathway. We also found that intrastriatal injection of COL resulted in side bias in spontaneous turning activities and apomorphine-induced rotational behavior. Together, our results provide in vivo data lending support to the concept that microtubule dysfunction may play a significant role in the death of DA neurons, though glial reaction may be involved and contribute to the degenerative process. Moreover, intrastriatal COL may serve as another experimental model of striatonigral degeneration (Parkinson's variant of multiple system atrophy), given the concurrent loss of both striatal and DA neurons.

Failure of neuronal protection by inhibition of glial activation in a rat model of striatonigral degeneration.

Previous studies in rodent models of neurodegenerative disorders have demonstrated that minocycline exerts neuroprotective effects unrelated to its antimicrobial action. The purpose of the present study was to analyze whether minocycline exhibits neuroprotective activity in a rat model of striatonigral degeneration (SND), the core pathology underlying levodopa-unresponsive parkinsonism associated with multiple system atrophy (MSA). We observed no significant effect of minocycline on locomotor impairment in double-lesioned SND rats. Minocycline significantly suppressed astroglial and microglial activation (P < 0.01); however, 3'5'-monophosphate-regulated phosphoprotein (DARPP 32) immunohistochemistry revealed no significant differences in striatal lesion volume of minocycline-treated versus untreated control SND rats. Furthermore, there was no protection of nigral dopaminergic neurons in the double-lesion model. We conclude that despite its astrocytic and microglial suppression, minocycline failed to attenuate lesion-induced neuronal damage in the SND rat model.

Striatal dopamine transporter in dementia with Lewy bodies and Parkinson disease: a comparison.

OBJECTIVE: To study the nigrostriatal pathways in 21 patients with dementia with Lewy bodies (DLB), 19 drug naive Parkinson disease (PD) patients, and 16 controls using a dopaminergic presynaptic ligand [123I]-2beta-carbometoxy-3beta-(4-iodophenyl)-N-(3-fluoropropyl) nortropane (FP-CIT) and SPECT in order to assess similarities or differences between DLB and PD. METHODS: A SPECT scan was carried out 3 to 4 hours after administration of 185 MBq (IV) of FP-CIT. Using occipital cortex as a radioactivity uptake reference, ratios for the caudate nuclei and the anterior and posterior putamina of both hemispheres were calculated. From the FP-CIT binding measurements, asymmetry indices and caudate:putamen ratios were derived. RESULTS: The DLB and PD groups had lower FP-CIT binding in all striatal areas than controls (analysis of variance: p < 0.001 in all measures). DLB patients also had significantly lower binding in the caudate nucleus than the PD patients. There was greater asymmetry of uptake in the posterior putamina of PD patients than DLB patients (p < 0.04) and controls (p < 0.01). The mean caudate:putamen ratio for the DLB group was not significantly different from that of the controls, while the mean caudate:putamen ratio of the PD group was higher than that of the control group (p < 0.001) and the DLB group (p < 0.001). CONCLUSION: This study showed differences between PD and DLB in the pattern of striatal dopaminergic dysfunction. DLB patients do not have the characteristic selective degeneration of ventrolateral nigral neurons seen in PD. This could explain some of the clinical differences between DLB and PD.

Potential involvement of cannabinoid receptors in 3-nitropropionic acid toxicity in vivo.

Several neurotransmitter systems are involved in the pathogenesis of Huntington's disease. Here, we examined the involvement of cannabinoid CB(1) receptors in striatal degeneration in the rat model of this disease generated by administration of 3-nitropropionic acid (3NP). Several days before onset of striatal degeneration, G-protein activation by cannabinoid agonists was significantly decreased whereas density and mRNA levels of CB(1) receptors remained essentially normal. This change was transient, CB(1) receptors recovering full functionality after few days. Later, at onset of striatal degeneration, profound alterations of CB(1) receptors were detected, including marked reductions of their density, mRNA levels and coupling to G proteins. In these rats, the administration of the cannabinoid agonist Delta(9)-tetrahydrocannabinol was neuroprotective, which indicates that the early loss of CB(1) receptor signaling could be instrumental in 3NP toxicity. In conclusion, the present study supports the hypothesis that cannabinoid receptors, possibly the CB(1) receptor subtype, may be involved in HD pathogenesis and could be an interesting therapeutic target to slow disease progression.

Neuropathological and behavioral changes induced by various treatment paradigms with MPTP and 3-nitropropionic acid in mice: towards a model of striatonigral degeneration (multiple system atrophy).

We characterized two models of dual nigral and striatal lesions replicating the lesion pattern of striatonigral degeneration, the neuropathological hallmark of parkinsonism associated with multiple system atrophy (SND/MSA-P). For this purpose, we used systemic administration of 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) and 3-nitropropionic acid (3-NP) in C57BL mice. One group of animals was first injected with MPTP followed by 3NP (MPTP+3-NP model). In the second group 3-NP was injected first, followed by MPTP (3-NP+MPTP model). The behavioral and neuropathological characteristics of these two models were compared to those observed after single 3-NP or MPTP intoxication. Results showed that, compared to control mice, spontaneous nocturnal locomotor activity was preserved in the MPTP+3-NP model, whereas it was reduced by 27% ( P<0.05) in the 3-NP+MPTP model and in animals treated with either 3-NP (27%, P<0.05) or MPTP (23%, P<0.05) alone. Quantitative histological evaluation based on Nissl staining and DARPP-32 immunohistochemistry revealed that 3-NP alone and 3-NP+MPTP treatment produced a marked (greater than 50%) loss of striatal neurons, whereas MPTP+3-NP treatment attenuated loss of striatal neurons by 43%. Further, loss of tyrosine hydroxylase-positive neurons in substantia nigra pars compacta (SNc) was attenuated after 3-NP+MPTP treatment compared to that observed after MPTP (40% vs 74%, P<0.001) and MPTP+3NP treatment (55% vs 74%, P<0.01). Our results show that MPTP-induced nigral lesions attenuate 3-NP toxicity and, reciprocally, that 3-NP-induced striatal lesions reduce MPTP toxicity. This suggests that complex integrative mechanisms are likely to regulate the vulnerability of the striatum and SNc to cell death in SND/MSA-P.

Different immunoreactivities of the microtubule-binding region of tau and its molecular basis in brains from patients with Alzheimer's disease, Pick's disease, progressive supranuclear palsy and corticobasal degeneration.

The microtubule-associated protein tau accumulates as cytoplasmic inclusions in Alzheimer's disease (AD), Pick's disease (PiD), progressive supranuclear palsy (PSP) and corticobasal degeneration (CBD). We investigated the immunoreactivity of tau-positive structures using a panel of antibodies to epitopes spanning the entire length of the tau molecule. In ethanol-fixed brain tissues, most antibodies to the microtubule-binding domain (MBD) required formic acid (FA) treatment to stain tau inclusions in PSP and CBD. This is in contrast with the intense labeling of neurofibrillary tangles in AD without FA treatment. Pick bodies (PiB) in PiD showed an intermediate pattern with respect to the immunoreactivity of the MBD because accumulated tau in PiB mostly lacks the insertion of exon 10, and the proportion of tau phosphorylated at Ser262 is smaller than in other abnormal tau structures. Such immunohistochemical profiles appeared to correlate with the occurrence of the smeared tau on immunoblot analysis of brain homogenate. The smeared tau was more abundant in AD and PiD than in PSP and CBD. Since the smeared tau was N-terminally truncated and was characteristic of advanced forms of modified tau, these findings suggest that tau accumulated in AD and PiD was processed more markedly than that in PSP and CBD. The MBD of tau may be masked in the presence of the intact N terminus and require FA treatment for antibody recognition in tissue sections. Advanced modification may expose the MBD in brain tissues of AD and PiD. It is suggested that the processing of abnormally accumulated tau characterizes the pathophysiology of each tauopathy.

Dystonia is predictive of subsequent altered dopaminergic responsiveness in a chronic 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine+3-nitropropionic acid model of striatonigral degeneration in monkeys.

We conducted a new chronic sequential 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and 3-nitropropionic acid (3NP) intoxication paradigm in two female monkeys in order to reproduce the striatonigral degeneration type of levodopa-unresponsive parkinsonism. A comparison was made with MPTP-, 3NP-intoxicated and control monkeys. A levodopa-responsive parkinsonism emerged in all MPTP-treated monkeys. During subsequent 3NP intoxication, one of the two MPTP +3NP monkeys exhibited hindlimb dystonia concomitantly with a reduced levodopa response. All MPTP-monkeys had severe cell loss in the substantia nigra pars compacta (>70%), but 3NP-induced discrete lesioned areas and cell loss predominantly in the putamen appeared only in the dystonic and levodopa-unresponsive animal. We propose that the appearance of dystonia after 3NP intoxication following dopaminergic striatal denervation is the key symptom predictive of the loss of dopaminergic response.

Comparison of unilateral and bilateral intrastriatal 6-hydroxydopamine-induced axon terminal lesions: evidence for interhemispheric functional coupling of the two nigrostriatal pathways.

Partial lesions of the nigrostriatal dopamine system can be induced reliably by the intrastriatal injection of 6-hydroxydopamine (6-OHDA) and are considered to be analogous to the early stages of human Parkinson's disease. Previous studies have established a clear correlation between different doses and placements of the 6-OHDA toxin and the degree of neurodegenerative changes and behavioral impairments. In the present study, the influence of the interdependence between the two nigrostriatal systems in both hemispheres on the effects on sensorimotor behavioral performances after terminal 6-OHDA lesions was investigated. The behavioral effects were correlated to the extent of nigral dopamine neuron cell and striatal tyrosine-hydroxylase (TH)-positive fiber loss. Sprague-Dawley rats receiving unilateral intrastriatal 6-OHDA injections (4 x 5 microg) exhibited a 30-70% reduction in striatal TH-positive fiber density along an anterior-posterior gradient, an 80% loss of nigral dopamine neurons and a mild degree of behavioral impairments as revealed by amphetamine-induced rotational asymmetry, and a reduced performance in the stepping and postural balance tests. When the same amount of toxin was injected twice into both hemispheres (2 x 4 x 5 microg), additional behavioral deficits were observed, consisting of a significant, but temporary, weight loss, a stable reduction in general locomotor activity and explorational behavior, and a long-term deficit in skilled forelimb use. This is interesting in light of the morphological findings, in which uni- and bilaterally lesioned animals did not differ significantly in the extent of TH-immunoreactive fiber and dopamine neuron loss within the nigrostriatal system in each lesioned hemisphere. These results indicate that the interdependent regulation of the two nigrostriatal systems may provide some compensatory support for the function and behavioral performance of the lesioned side via the normal unlesioned side, which is lost in animals with bilateral lesions of the nigrostriatal system. Therefore, this model of uni- and bilateral partial lesions of the nigrostriatal system, as characterized in the present study, may foster further exploration of compensatory functional mechanisms active in the early stages of Parkinson's disease and promote development of novel neuroprotective and restorative strategies.

Fibroblast growth factor-2-producing fibroblasts protect the nigrostriatal dopaminergic system from 6-hydroxydopamine.

We tested the hypothesis that fibroblasts, which had been genetically engineered to produce fibroblast growth factor-2 (FGF-2), can protect nigrostriatal dopaminergic neurons. Three groups of rats received either a burr hole only (n=5) or implantation of fibroblasts, which had been genetically engineered to produce beta-galactosidase (beta-gal) (n=8) or FGF-2 (n=8), at two sites in the right striatum. Two weeks later, the animals received an injection of 25 microg of 6-hydroxydopamine hydrobromide (6-OHDA) midway between the two implant sites. The group that received FGF-2-fibroblasts had significantly fewer apomorphine-induced rotations than the groups that received a burr hole only or beta-gal-fibroblasts at weeks 2 and 3 following lesioning with 6-OHDA. Testing for amphetamine-induced rotation revealed a mild reduction in rotation in the beta-gal-fibroblast group compared to the burr hole only group, but a striking attenuation of amphetamine-induced rotation in the FGF-2-fibroblast group. There was also preservation of TH-IR neurons on the lesioned side relative to both control groups. The size of the grafts and the gliosis surrounding the injection sites did not differ between the FGF-2-fibroblast and beta-gal-fibroblast groups. To further characterize the production of FGF-2 by the FGF-2-fibroblasts, we implanted FGF-2-fibroblasts and beta-gal-fibroblast into the striatum of rats but did not lesion the animals with 6-OHDA. The animals were then sacrificed at 1, 2 and 5 weeks following implantation. Prior to implantation the FGF-2 fibroblasts contained 148 ng/mg of FGF-2-immunoreactive (FGF-2-IR) material per mg of protein of cell lysate. After implantation FGF-2-IR material was noted in the grafts of FGF-2-fibroblasts, most conspicuously at 1 and 2 weeks following implantation. We also noted FGF-2-IR material in the nuclei of reactive astrocytes adjacent to the implants, and OX-42-immunoreactive (OX-42-IR) cells adjacent and occasionally within the implants. Our work indicates that fibroblasts genetically engineered to produce FGF-2 and implanted in the striatum can protect the nigrostriatal dopaminergic system and may be useful in the treatment of Parkinson's disease.

Effects of nigrostriatal dopamine denervation on ionotropic glutamate receptors in rat caudate-putamen.

Changes in ionotropic glutamate NMDA, AMPA and KA receptor binding in rat caudate-putamen were examined by quantitative in vitro receptor autoradiography 5 weeks after lesioning nigrostriatal dopaminergic projections. In this animal model of Parkinson's disease, density of binding in caudate-putamen increased at KA, but not NMDA or AMPA receptors. The findings indicate that nigrostriatal dopamine denervation can selectively enhance KA receptor levels in rat basal ganglia, suggest that KA receptors contribute to the pathophysiology of Parkinson's disease, and may suggest innovative treatments.

Tremor is associated with PET measures of nigrostriatal dopamine function in MPTP-lesioned monkeys.

Unilateral intracarotid artery (ICA) MPTP infusion, along with sequential systemic doses of MPTP, produces near complete degeneration of the nigrostriatal pathway on the side of infusion (ipsilateral) and variable levels of damage in the contralateral hemisphere accompanied by varying levels of parkinsonism (overlesioned hemiparkinsonian model). Positron emission tomography and the dopamine (DA) metabolism tracer [(18)F]6-fluoro-l-m-tyrosine (FMT) were used to evaluate the relationship between DA metabolism and clinical features of parkinsonism in 14 overlesioned hemiparkinsonian monkeys. Monkeys were rated on a parkinsonian scale that included ratings of bradykinesia, fine motor skills (FMS), and rest tremor. Because the monkeys tended to show more severe clinical signs on the side of the body contralateral to ICA MPTP infusion, we calculated asymmetry scores for each of the clinical features as well as for FMT uptake (K(i)) in the caudate and putamen. Tremor asymmetry was associated with FMT uptake asymmetry in the putamen. No such relationship was observed for FMS or bradykinesia. The overall severity of tremor (mild, moderate/severe) was associated with FMT uptake in the caudate and putamen. Postmortem biochemical analysis for a subset of monkeys showed that the monkeys with moderate/severe tremor had significantly lower DA levels in both caudate and putamen than those with mild tremor. In addition, K(i) values were significantly correlated with DA levels in both caudate and putamen. These findings support the idea that nigrostriatal degeneration contributes to rest tremor.

Complex motor disturbances in a sequential double lesion rat model of striatonigral degeneration (multiple system atrophy).

This study characterizes paw reaching, stepping and balance abnormalities in a double lesion rat model of striatonigral degeneration, the core pathology underlying levodopa unresponsive parkinsonism associated with multiple system atrophy. Extensive unilateral nigral or striatal lesions induced by 6-hydroxydopamine or quinolinic acid, respectively, produced a similarly marked contralateral paw reaching deficit without further deterioration following a secondary (complementary) lesion of ipsilateral striatum or substantia nigra. Contralateral stepping rates were reduced by unilateral 6-hydroxydopamine lesions without further deterioration following the secondary striatal lesion. In contrast, initial unilateral striatal quinolinic acid injections induced bilateral stepping deficits that significantly worsened contralaterally following the secondary nigral lesion. Contralateral sidefalling rates were significantly increased following primary nigral and striatal lesions. Secondary nigral but not secondary striatal lesions worsened contralateral sidefalling rates. Histological studies revealed subtotal (>90%) depletion of dopaminergic neurons in substantia nigra pars compacta and variable degrees of striatal degeneration depending on the lesion sequence. Animals pre-lesioned with 6-hydroxydopamine showed significantly larger residual striatal surface areas following the secondary striatal quinolinic acid lesion compared to animals with primary striatal quinolinic acid lesions (P<0.001). These findings are in line with previous experimental studies demonstrating that striatal dopamine depletion confers neuroprotection against subsequent excitotoxic injury. Whether loss of dopaminergic neurons protects against the striatal disease process occurring in multiple system atrophy (Parkinson-type) remains to be elucidated. In summary, this is the first experimental study to investigate spontaneous motor behaviour in a unilateral double lesion rat model. Our observations are consistent with a complex interaction of nigral and striatal lesions producing distinct behavioural and histological changes depending on the lesion sequence. Tests of forelimb akinesia and complex motor behaviour appear to provide a reliable tool that will be helpful for monitoring the effects of interventional strategies such as embryonic neuronal transplantation in the rat model of striatonigral degeneration.