Dopaminergic cells in the substantia nigra are highly vulnerable to the neurodegenerative process of Parkinson's disease. Therefore, mechanisms that enhance their susceptibility to injury bear important implications for disease pathogenesis. Repeated injections with the herbicide paraquat cause oxidative stress and a selective loss of dopaminergic neurons in mice. In this model, the first paraquat exposure, though not sufficient to induce any neurodegeneration, predisposes neurons to damage by subsequent insults. The purpose of this study was to elucidate the mechanisms underlying this "priming" event. We found that a single paraquat exposure was followed by an increase in the number of cells with immunohistochemical, morphological and biochemical characteristics of activated microglia, including induction of NADPH-oxidase. If this microglial response was inhibited by the anti-inflammatory drug minocycline, subsequent exposures to the herbicide failed to cause oxidative stress and neurodegeneration. On the other hand, if microglial activation was induced by pretreatment with lipopolysaccharide, a single paraquat exposure became capable of triggering a loss of dopaminergic neurons. Finally, mutant mice lacking functional NADPH-oxidase were spared from neurodegeneration caused by repeated paraquat exposures. Data indicate that microglial activation and consequent induction of NADPH-oxidase may act as risk factors for Parkinson's disease by increasing the vulnerability of dopaminergic cells to toxic injury.

Alpha-synuclein is likely to play a role in neurodegenerative processes, including the degeneration of nigrostriatal dopaminergic neurons that underlies Parkinson's disease. However, the toxicological properties of alpha-synuclein remain relatively unknown. Here, the relationship between alpha-synuclein expression and neuronal injury was studied in mice exposed to the herbicide paraquat. Paraquat neurotoxicity was compared in control animals versus mice with transgenic expression of human alpha-synuclein driven by the tyrosine hydroxylase (TH) promoter. In control mice, paraquat caused both the formation of alpha-synuclein-containing intraneuronal deposits and the degeneration of nigrostriatal neurons, as demonstrated by silver staining and a reduction of the counts of TH-positive and Nissl-stained cells. Mice overexpressing alpha-synuclein, either the human wild-type or the Ala53Thr mutant form of the protein, displayed paraquat-induced protein aggregates but were completely protected against neurodegeneration. These resistant animals were also characterized by increased levels of HSP70, a chaperone protein that has been shown to counteract paraquat toxicity in other experimental models and could therefore contribute to neuroprotection in alpha-synuclein transgenic mice. The results indicate a dissociation between toxicant-induced alpha-synuclein deposition and neurodegeneration. They support a role of alpha-synuclein against toxic insults and suggest that its involvement in human neurodegenerative processes may arise not only from a gain of toxic function, as previously proposed, but also from a loss of defensive properties.

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This study examines the developmental expression of GABAB receptor subunits (GABAB(1a), GABAB(1b), GABAB(2)) in the pituitary intermediate lobe using in situ hybridization, reverse transcriptase-polymerase chain reaction, immunohistochemistry, and Western blots. Receptor functionality was studied by baclofen-stimulated GTPgammaS binding. In the adult rat pituitary all three transcripts were detected in melanotropes, but not in glia, of the intermediate lobe. No transcripts of any subunit were detected in the neural lobe. Transcripts of GABAB(1a) and GABAB(1b), but not of GABAB(2), were detected in specific subpopulations of cells in the anterior lobe. All three transcripts were detected in melanotropes on gestational day 18 using in situ hybridization. Reverse transcriptase-polymerase chain reactions comparing postnatal day 2 and adult transcript levels in the neurointermediate lobe support in situ hybridization data that GABAB(1a) mRNA levels do not change, GABAB(1b) levels increase, and GABAB(2) levels decrease as the rat matures. Thus, GABAB receptor subunit transcripts are differentially regulated in melanotropes during development. In the adult rat both GABAB(1) and GABAB(2) proteins were detected in the neurointermediate lobe using Western blotting and in melanotropes by immunohistochemistry. Developmentally, GABAB(1) protein was not detected until postnatal day 7, but was clearly expressed by postnatal day 15 while GABAB(2) protein could not be detected until postnatal day 15. Functional receptors were found in the intermediate lobe at postnatal day 15 and in the adult. The demonstration of transcripts for GABAB(1a), GABAB(1b) and GABAB(2) subunits at gestational day 18 contrasted with the failure to detect any protein before postnatal day 7, suggesting that the regulation of GABAB subunit isoforms occurs differentially at both the transcriptional and translational level as development progresses. The disparity in the regulation of the receptor subunits may suggest that GABAB(1) could have other functions besides being part of the GABAB receptor heterodimer.