We have shown in the parkinsonism-inducing neurotoxin MPP(+)/MPTP model that alpha-Synuclein (alpha-Syn), a presynaptic protein causal in Parkinson's disease (PD), contributes to hyperphosphorylation of Tau (p-Tau), a protein normally linked to tauopathies, such as Alzheimer's disease (AD). Here, we investigated the kinase involved and show that the Tau-specific kinase, glycogen synthase kinase 3beta (GSK-3beta), is robustly activated in various MPP(+)/MPTP models of Parkinsonism (SH-SY5Y cotransfected cells, mesencephalic neurons, transgenic mice overexpressing alpha-Syn, and postmortem striatum of PD patients). The activation of GSK-3beta was absolutely dependent on the presence of alpha-Syn, as indexed by the absence of p-GSK-3beta in cells lacking alpha-Syn and in alpha-Syn KO mice. MPP(+) treatment induced translocation and accumulation of p-GSK-3beta in nuclei of SH-SY5Y cells and mesencephalic neurons. Through coimmunoprecipitation (co-IP), we found that alpha-Syn, pSer396/404-Tau, and p-GSK-3beta exist as a heterotrimeric complex in SH-SY5Y cells. GSK-3beta inhibitors (lithium and TDZD-8) protected against MPP(+)-induced events in SH-SY5Y cells, preventing cell death and p-GSK-3beta formation, by reversing increases in alpha-Syn accumulation and p-Tau formation. These data unveil a previously unappreciated role of alpha-Syn in the induction of p-GSK-3beta, and demonstrate the importance of this kinase in the genesis and maintenance of neurodegenerative changes associated with PD.

Human alpha-synuclein accumulates in dopaminergic neurons as intraneuronal inclusions, Lewy bodies, which are characteristic of idiopathic Parkinson's disease (PD). Here, we suggest that modulation of the functional activity of the dopamine transporter (DAT) by alpha-synuclein may be a key factor in the preferential degeneration of mesencephalic dopamine (DA)-synthesizing neurons in PD. In cotransfected Ltk-, HEK 293, and SK-N-MC cells, alpha-synuclein induced a 35% decrease in [3H]DA uptake. Biotinylated DAT levels were decreased by 40% in cotransfected cells relative to cells expressing only DAT. DAT was colocalized with alpha-synuclein in mesencephalic neurons and cotransfected Ltk- cells. Coimmunoprecipitation studies showed the existence of a complex between alpha-synuclein and DAT, in specific rat brain regions and cotransfected cells, through specific amino acid motifs of both proteins. The attenuation of DAT function by alpha-synuclein was cytoprotective, because DA-mediated oxidative stress and cell death were reduced in cotransfected cells. The neurotoxin MPP+ (1-methyl-4-phenylpyridinium), oxidative stress, or impairment of cell adhesion ablated the alpha-synuclein-mediated inhibition of DAT activity, which caused increased uptake of DA and increased biotinylated DAT levels, in both mesencephalic neurons and cotransfected cells. These studies suggest a novel normative role for alpha-synuclein in regulating DA synaptic availability and homeostasis, which is relevant to the pathophysiology of PD.

Parkinson’s disease [PD], a progressive neurodegenerative disease, results in abnormal accumulation of insoluble alpha-synuclein [α-Syn] in dopaminergic neurons. Here we examined tauopathic changes and the α-Syn/p-GSK-3β/proteasome pathway in postmortem striata and inferior frontal gyri [IFG] from patients with PD and PD with dementia [PDD]. In both PD and PDD, α-Syn levels were high, especially the insoluble form of this protein; in PDD, insoluble α-Syn levels were persistently higher than PD across both brain regions. Levels of p-GSK-3β phosphorylated at Tyr 216, which hyperphosphorylates Tau to produce toxic pathological forms of p-Tau, were higher in striata of both PD and PDD compared to controls, but were unaltered in IFG. While proteasomal activity was unchanged in striatum of PD and PDD, such activity was diminished in the IFG of both PD and PDD. A decrease in 19S subunit of the proteasomes was seen in IFG of PDD, while lower levels of 20S subunits were seen in striatum and IFG of both PD and PDD patients. Parkin levels were similar in PD and PDD, suggesting lack of involvement of this protein. Most interestingly, tauopathic changes were noted only in striatum of PD and PDD, with increased hyperphosphorylation seen at Ser262 and Ser396/404; increases in Ser202 levels were seen only in PD but not in PDD striatum. We were unable to detect any tauopathy in IFG in either PD or PDD despite increased levels of α-Syn, and decreased proteasomal activity, and is probably due to lack of increase in p-GSK-3β in IFG. Unlike Alzheimer’s disease where tauopathy is more globally observed in diverse brain regions, our data demonstrates restricted expression of tauopathy in brains of PD and PDD, probably limited to dopaminergic neurons of the nigrostriatal region.

alpha-Synuclein is a key component of the pathological process of neurodegeneration in Parkinson's disease. Although its contributions to normal physiological conditions remain elusive, converging observations suggest that a primary function of this protein in dopaminergic neurons may be the regulation of dopamine content and synaptic tone at the synapse. We review here cumulative evidence that demonstrates the participation of alpha-synuclein in the life cycle of dopamine from its synthesis, storage, release, and reuptake. The regulatory role of alpha-synuclein on dopamine metabolism is assessed by discussing the experimental evidence supporting each of these observations in the healthy physiological maintenance of dopaminergic neurons, as well as showing how disruption of these events can initiate the observed neurotoxicity of alpha-synuclein and the genesis of the degenerative processes associated with Parkinson's disease.

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Many neurodegenerative diseases associated with functional Tau dysregulation, including Alzheimer's disease (AD) and other tauopathies, also show alpha-synuclein (alpha-Syn) pathology, a protein associated with Parkinson's disease (PD) pathology. Here we show that treatment of primary mesencephalic neurons (48 h) or subchronic treatment of wild-type (WT) mice with the Parkinsonism-inducing neurotoxin MPP+/MPTP, results in selective dose-dependent hyperphosphorylation of Tau at Ser396/404 (PHF-1-reactive Tau, p-Tau), with no changes in pSer202 but with nonspecific increases in pSer262 levels. The presence of alpha-Syn was absolutely mandatory to observe MPP+/MPTP-induced increases in p-Tau levels, since no alterations in p-Tau were seen in transfected cells not expressing alpha-Syn or in alpha-Syn-/- mice. MPP+/MPTP also induced a significant accumulation of alpha-Syn in both mesencephalic neurons and in WT mice striatum. MPTP/MPP+ lead to differential alterations in p-Tau and alpha-Syn levels in a cytoskeleton-bound, vs. a soluble, cytoskeleton-free fraction, inducing their coimmunoprecipitation in the cytoskeleton-free fraction and neuronal soma. Subchronic MPTP exposure increased sarkosyl-insoluble p-Tau in striatum of WT but not alpha-Syn-/- mice. These studies describe a novel mechanism for MPTP neurotoxicity, namely a MPTP-inducible, strictly alpha-Syn-dependent, increased formation of PHF-1-reactive Tau, suggesting convergent overlapping pathways in the genesis of clinically divergent diseases such as AD and PD.

The natriuretic effect of DA-1 agonists is less in the spontaneously hypertensive rat (SHR) than its normotensive control, the Wistar-Kyoto rat (WKY). To determine a mechanism of the decreased effect of DA-1 agonists on sodium transport, DA-1 receptors in renal proximal convoluted tubule (PCI) were studied by radioligand binding and by adenylate cyclase (AC) determinations. Specific binding of '25I-SCH 23982 (defined by 10 MM SCH 23390, a DA-1 antagonist) was concentration dependent, saturable, and stereoselective. The dissociation constant, maximum receptor density, and DA-1 antagonist inhibition constant were similar in SHR and WKY. The apparent molecular weight of the DA-1 receptor determined by the photoaffinity D1 probe '25I-MAB was also similar in WKY and SHR. However, DA-1 agonists competed more effectively for specific '25I-SCH 23982 binding sites in WKY than in SHR.Basal as well as forskolin, parathyroid hormone, GTP and Gpp(NH)p-stimulated-AC activities were similar. In contrast DA-1 agonists (fenoldopam, SKF 38393, SND 911C12) stimulated AC activity to a lesser extent in SHR. GTP and Gpp(NH)p enhanced the ability of DA-1 agonists to stimulate AC activity in WKY but not in SHR. These data suggest a defect in the DA-1 receptor-second messenger coupling mechanism in the PCI of the SHR.

Although clinically distinct diseases, tauopathies and synucleinopathies share common genesis and mechanisms, leading to overlapping degenerative changes within neurons. In human postmortem striatum of Parkinson's disease [PD] and PD with dementia, we have recently described elevated levels of tauopathy, indexed as increased hyperphosphorylated Tau [p-Tau]. Here we assessed tauopathy in striatum of a transgenic animal model of PD, overexpressing human α-synuclein under the PDGF promoter. At 11 months of age, large and progressive increases in p-Tau in transgenic mice, hyperphosphorylated at sites reminiscent of Alzheimer's disease, were noted, along with elevated levels of α-synuclein and p-GSK-3β, a major kinase involved in hyperphosphorylation of Tau. Differential Triton X-100 extraction of striata showed the presence of aggregated α-Syn in the Tg mice, along with p-Tau and p-GSK-3β, which was also confirmed through immunohistochemistry. After p-Tau formation, both Tau and MAP1 dissociated from the cytoskeleton, consistent with diminished ability of these cytoskeleton-binding proteins to bind microtubules. Increases in free tubulin and actin were also noted, indicative of cytoskeleton remodeling and destabilization. In vivo magnetic resonance imaging of the transgenic animals showed a reduction in brain volume of transgenic mice indicating substantial atrophy. From immunohistochemical studies, α-synuclein, p-Tau and p-GSK-3β were found to be overexpressed and co-localized in large inclusion bodies, reminiscent of Lewy bodies. The elevated state of tauopathy seen in these PDGF-α-synuclein mice provides further confirmation that Parkinson's may be a tauopathic disease. KeywordsAlzheimer's disease; neurodegeneration; synucleinopathies; tauopathies Sporadic Parkinson's disease [PD] is a progressive neurodegenerative disease of unknown etiology [Pollanen et al, 1993], resulting in loss of motor function and degeneration of dopaminergic neurons [Pollanen et al, 1993;Jakes et al, 1994;Forno, 1996;Spillantini et al, 1998;Corti et al 2005]. Alpha-synuclein [α-Syn], a presynaptic protein, is causal in the genesis of PD [Forno, 1996;Spillantini et al, 1998], and gene duplication and triplication of α-Syn are found in sporadic and early onset forms of PD [Singleton et al, 2003] NIH Public Access Author ManuscriptEur J Neurosci. Author manuscript; available in PMC 2012 May 1. A53T and E46K] in the gene are linked to autosomal dominant familial forms of the disease [Polymeropoulos et al, 1997;Kruger et al, 1998]. Although a soluble protein, under pathological conditions, α-Syn becomes insoluble, self-aggregates and accumulates into intra-neuronal inclusion bodies [Spillantini et al, 1998;Masliah et al, 2000;El-Agnaf et al, 1998;Gosavi et al, 2002]. Similar to α-Syn, Tau is also a highly soluble protein that becomes insoluble by pathological hyperphosphorylation, leading to tauopathies, including Alzheimer's disease (AD), [Dickson et al, 2002;Esper et al, 2007;Gasparini et al, 2007;Wadia & Lang, 2007;Gong & Iqba...