The vesicular monoamine transporter 2 (VMAT2; SLC18A2) is responsible for packaging dopamine into vesicles for subsequent release and has been suggested to serve a neuroprotective role in the dopamine system. Here, we show that mice that express ϳ5% of normal VMAT2 (VMAT2 LO) display age-associated nigrostriatal dopamine dysfunction that ultimately results in neurodegeneration. Elevated cysteinyl adducts to L-DOPA and DOPAC are seen early and are followed by increased striatal protein carbonyl and 3-nitrotyrosine formation. These changes were associated with decreased striatal dopamine and decreased expression of the dopamine transporter and tyrosine hydroxylase. Furthermore, we observed an increase in ␣-synuclein immunoreactivity and accumulation and neurodegeneration in the substantia nigra pars compacta in aged VMAT2 LO mice. Thus, VMAT2 LO animals display nigrostriatal degeneration that begins in the terminal fields and progresses to eventual loss of the cell bodies, ␣-synuclein accumulation, and an L-DOPA responsive behavioral deficit, replicating many of the key aspects of Parkinson's disease. These data suggest that mishandling of dopamine via reduced VMAT2 expression is, in and of itself, sufficient to cause dopamine-mediated toxicity and neurodegeneration in the nigrostriatal dopamine system. In addition, the altered dopamine homeostasis resulting from reduced VMAT2 function may be conducive to pathogenic mechanisms induced by genetic or environmental factors thought to be involved in Parkinson's disease.

Mutations in the gene encoding leucine-rich repeat kinase 2 (LRRK2) have been identified as the cause of familial Parkinson's disease (PD) at the PARK8 locus. To begin to understand the physiological role of LRRK2 and its involvement in PD, we have investigated the distribution of LRRK2 mRNA and protein in the adult mouse brain. In situ hybridization studies indicate sites of mRNA expression throughout the mouse brain, with highest levels of expression detected in forebrain regions, including the cerebral cortex and striatum, intermediate levels observed in the hippocampus and cerebellum, and low levels in the thalamus, hypothalamus and substantia nigra. Immunohistochemical studies demonstrate localization of LRRK2 protein to neurones in the cerebral cortex and striatum, and to a variety of interneuronal subtypes in these regions. Furthermore, expression of LRRK2 mRNA in the striatum of VMAT2-deficient mice is unaltered relative to wild-type littermate controls despite extensive dopamine depletion in this mouse model of parkinsonism. Collectively, our results demonstrate that LRRK2 is present in anatomical brain regions of direct relevance to the pathogenesis of PD, including the nigrostriatal dopaminergic pathway, in addition to other regions unrelated to PD pathology, and is likely to play an important role in the normal function of telencephalic forebrain neurones and other neuronal populations.

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Dopamine cytotoxicity is thought to contribute towards the selective loss of substantia nigra pars compacta dopamine neurons and disease progression in Parkinson's disease. However, the long-term toxicity of dopamine in vivo has not previously been established. The vesicular monoamine transporter 2 (VMAT2) sequesters monoamines into synaptic vesicles, a process that, in addition to being important in normal transmission, may also act to keep intracellular levels of monoamine neurotransmitters below potentially toxic thresholds. The homozygous VMAT2-hypomorphic mouse has an insertion in the VMAT2 gene (Slc18a2). Consequently, VMAT2-deficient mice (VD(-/-)) have an approximately 95% reduction in VMAT2 expression and an equivalent level of dopamine depletion in the striatum which results in moderate motor impairment. Here, we show that L-DOPA induces locomotor hyperactivity in VD(-/-) mice and reverses the deficit in motor coordination and balance as tested with the rotarod. We report that evidence for cytosolic accumulation of dopamine in substantia nigra neurons in these mice is two-fold: firstly, there is reduced phosphorylation of tyrosine hydroxylase at the residue associated with catechol feedback inhibition; and, secondly, there are increased rates of dopamine turnover at 6, 12 and 24 months of age. These animals exhibit a progressive decline in striatal monoamine levels and rotarod performance with increasing age. However, despite these data, there was no loss of nigral dopamine neurons as estimated by quantification of tyrosine hydroxylase-immunoreactive cells in the substantia nigra pars compacta of old VD(-/-) mice (24-month-old), implying that these age-dependent manifestations may be due to senescence alone.