Background: Dopamine-synthesizing (dopaminergic, DA) neurons in the ventral midbrain (VM) constitute a pivotal neuronal population controlling motor behaviors, cognitive and affective brain functions, which generation critically relies on the activation of Wingless-type MMTV integration site (Wnt)/β-catenin pathway in their progenitors. In Parkinson's disease, DA cell bodies within the substantia nigra pars compacta (SNpc) progressively degenerate, with causes and mechanisms poorly understood. Emerging evidence suggests that Wnt signaling via Frizzled (Fzd) receptors may play a role in different degenerative states, but little is known about Wnt signaling in the adult midbrain. Using in vitro and in vivo model systems of DA degeneration, along with functional studies in both intact and SN lesioned mice, we herein highlight an intrinsic Wnt1/Fzd-1/β-catenin tone critically contributing to the survival and protection of adult midbrain DA neurons. Results: In vitro experiments identifie Fzd-1 receptor expression at a mRNA and protein levels in dopamine transporter (DAT) expressing neurons, and demonstrate the ability of exogenous Wnt1 to exert robust neuroprotective effects against Caspase-3 activation, the loss of tyrosine hydroxylase-positive (TH⁺) neurons and [³H] dopamine uptake induced by different DA-specific insults, including serum and growth factor deprivation, 6-hydroxydopamine and MPTP/MPP⁺. Co-culture of DA neurons with midbrain astrocytes phenocopies Wnt1 neuroprotective effects, whereas RNA interference-mediated knockdown of Wnt1 in midbrain astrocytes markedly reduces astrocyte-induced TH⁺ neuroprotection. Likewise, silencing β-catenin mRNA or knocking down Fzd-1 receptor expression in mesencephalic neurons counteract astrocyte-induced TH⁺ neuroprotection. In vivo experiments document Fzd-1 co-localization with TH⁺ neurons within the intact SNpc and blockade of Fzd/β-catenin signaling by unilateral infusion of a Fzd/β-catenin antagonist within the SN induces reactive astrocytosis and acutely inhibits TH⁺ neuron survival in ipsilateral SNpc, an effect efficiently prevented by pharmacological activation of β-catenin signaling within the SNpc. Conclusion: These results defining a novel Wnt1/Fzd-1/β-catenin astrocyte-DA autoprotective loop provide a new mechanistic inside into the regulation of pro-survival processes, with potentially relevant consequences for drug design or drug action in Parkinson's disease.

A common hallmark of age‐dependent neurodegenerative diseases is an impairment of adult neurogenesis. Wingless‐type mouse mammary tumor virus integration site (Wnt)/β‐catenin (WβC) signalling is a vital pathway for dopaminergic (DAergic) neurogenesis and an essential signalling system during embryonic development and aging, the most critical risk factor for Parkinson's disease (PD). To date, there is no known cause or cure for PD. Here we focus on the potential to reawaken the impaired neurogenic niches to rejuvenate and repair the aged PD brain. Specifically, we highlight WβC‐signalling in the plasticity of the subventricular zone (SVZ), the largest germinal region in the mature brain innervated by nigrostriatal DAergic terminals, and the mesencephalic aqueduct‐periventricular region (Aq‐PVR) Wnt‐sensitive niche, which is in proximity to the SNpc and harbors neural stem progenitor cells (NSCs) with DAergic potential. The hallmark of the WβC pathway is the cytosolic accumulation of β‐catenin, which enters the nucleus and associates with T cell factor/lymphoid enhancer binding factor (TCF/LEF) transcription factors, leading to the transcription of Wnt target genes. Here, we underscore the dynamic interplay between DAergic innervation and astroglial‐derived factors regulating WβC‐dependent transcription of key genes orchestrating NSC proliferation, survival, migration and differentiation. Aging, inflammation and oxidative stress synergize with neurotoxin exposure in “turning off” the WβC neurogenic switch via down‐regulation of the nuclear factor erythroid‐2‐related factor 2/Wnt‐regulated signalosome, a key player in the maintenance of antioxidant self‐defense mechanisms and NSC homeostasis. Harnessing WβC‐signalling in the aged PD brain can thus restore neurogenesis, rejuvenate the microenvironment, and promote neurorescue and regeneration.

In Parkinson's disease (PD), neurogenesis is impaired in the subventricular zone (SVZ) of postmortem human PD brains, in primate nonhuman and rodent models of PD. The vital role ofWingless-type MMTV integration site (Wnt)/β-cateninsignaling in the modulation of neurogenesis, neuroprotection, and synaptic plasticity coupled to our recent findings uncovering an active role for inflammation andWnt/β-cateninsignaling in MPTP-induced loss and repair of nigrostriatal dopaminergic (DAergic) neurons prompted us to study the impact of neuroinflammation and theWnt/β-cateninpathway in the response of SVZ neuroprogenitors (NPCs) in MPTP-treated mice.In vivoexperiments, using bromodeoxyuridine and cell-specific markers, andex vivotime course analyses documented an inverse correlation between the reduced proliferation of NPCs and the generation of new neuroblasts with the phase of maximal exacerbation of microglia reaction, whereas a shift in the microglia proinflammatory phenotype correlated with a progressive NPC recovery.Ex vivoandin vitroexperiments using microglia–NPC coculture paradigms pointed to NADPH-oxidase (gpPHOX⁹¹), a major source of microglial ROS, and reactive nitrogen species as candidate inhibitors of NPC neurogenic potential via the activation of glycogen synthase 3 (pGSK-3β^Tyr216), leading to loss of β-catenin, a chief downstream transcriptional effector. Accordingly, MPTP/MPP⁺(1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) caused β-catenin downregulation and pGSK-3β^Tyr216overexpression, whereas manipulation ofWnt/β-cateninsignaling with RNA interference-mediatedGSK-3β knockdown or GSK-3β antagonism reversed MPTP-induced neurogenic impairmentex vivo/in vitroorin vivo. Reciprocally, pharmacological modulation of inflammation prevented β-catenindownregulation and restored neurogenesis, suggesting the possibility to modulate this endogenous system with potential consequences for DAergic neuroprotection and self-repair.

Wnt/β‐catenin signaling is required for specification and neurogenesis of midbrain dopaminergic (mDA) neurons, the pivotal neuronal population that degenerates in Parkinson's disease (PD), and in 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP) mouse model of PD. Wnt/β‐catenin signaling plays a vital role in adult neurogenesis but whether it might engage DA neurogenesis/neurorepair in the affected PD brain is yet unresolved. Recently, the adult midbrain aqueduct periventricular regions (Aq‐PVRs) were shown to harbor multipotent clonogenic neural stem/progenitor cells (mNPCs) with DA potential in vitro, but restrictive mechanisms in vivo are believed to limit their DA regenerative capacity. Using in vitro mNPC culture systems we herein demonstrate that aging is one most critical factor restricting mNPC neurogenic potential via dysregulation of Wnt/β‐catenin signaling. Coculture paradigms between young/aged (Y/A) mNPCs and Y/A astrocytes identified glial age and a decline of glial‐derived factors including Wnts as key determinants of impaired neurogenic potential, whereas Wnt activation regimens efficiently reversed the diminished proliferative, neuronal, and DA differentiation potential of A‐mNPCs. Next, in vivo studies in wild (Wt) and transgenic β‐catenin reporter mice uncovered Wnt/β‐catenin signaling activation and remarkable astrocyte remodeling of Aq‐PVR in response to MPTP‐induced DA neuron death. Spatio‐temporal analyses unveiled β‐catenin signaling in predopaminergic (Nurr1⁺/TH⁻) and imperiled or rescuing DAT⁺ neurons during MPTP‐induced DA neuron injury and self‐repair. Aging inhibited Wnt signaling, whereas β‐catenin activation in situ with a specific GSK‐3β antagonist promoted a significant degree of DA neurorestoration associated with reversal of motor deficit, with implications for neurorestorative approaches in PD. Stem Cells 2014;32:2147–2163