Α-synuclein Preformed Fibrils Research Articles

Developmental origins of parkinson's disease risk: perinatal exposure to the organochlorine pesticide dieldrin leads to sex-specific DNA modifications in critical neurodevelopmental pathways in the mouse midbrain.

Epidemiological studies show that exposure to the organochlorine pesticide dieldrin is associated with increased risk of Parkinson's disease (PD). Animal studies support a link between developmental dieldrin exposure and increased neuronal susceptibility in the α-synuclein preformed fibril (α-syn PFF) and MPTP models in adult male C57BL/6 mice. In a previous study, we showed that developmental dieldrin exposure was associated with sex-specific changes in DNA modifications within genes related to dopaminergic neuron development and maintenance at 12 weeks of age. Here, we used capture hybridization-sequencing with custom baits to interrogate DNA modifications across the entire genetic loci of the previously identified genes at multiple time points-birth, 6 weeks, 12 weeks, and 36 weeks old. We identified largely sex-specific dieldrin-induced changes in DNA modifications at each time point that annotated to pathways important for neurodevelopment, potentially related to critical steps in early neurodevelopment, dopaminergic neuron differentiation, synaptogenesis, synaptic plasticity, and glial-neuron interactions. Despite large numbers of age-specific DNA modifications, longitudinal analysis identified a small number of DMCs with dieldrin-induced deflection of epigenetic aging. The sex-specificity of these results adds to evidence that sex-specific responses to PD-related exposures may underly sex-specific differences in disease. Overall, these data support the idea that developmental dieldrin exposure leads to changes in epigenetic patterns that persist after the exposure period and disrupt critical neurodevelopmental pathways, thereby impacting risk of late life diseases, including PD.

Novel 4-triazole phenyl amide (4-TPA) molecules: Potent promoters of α-synuclein fibril disassembly

Parkinson's disease profoundly compromises patients' daily lives, and the disassembly of α-synuclein aggregates, a primary pathological factor, represents a promising therapeutic approach. In this study, we conducted a systematic screening and optimization process to identify the novel scaffold B37, a 4-triazolyl-phenylamine derivative, exhibiting a potent disassembly activity of 1.1 μM against α-synuclein preformed fibrils. Notably, B37 demonstrated significant neuroprotective effects, ameliorated autophagic dysfunction induced by preformed fibrils, mitigated oxidative stress, and restored the co-localization of preformed fibrils with lysosomes. Transmission electron microscopy corroborated its in vitro disassembly function. Pharmacokinetic profiling revealed favorable parameters with a receptible blood-brain barrier permeability. B37 emerges as a promising lead compound for further optimization, aiming to develop a highly effective agent targeting the disassembly of α-synuclein aggregates to treat neurodegenerative diseases like Parkinson's disease.

Inhibition of EHMT1/2 rescues synaptic damage and motor impairment in a PD mouse model

Epigenetic dysregulation that leads to alterations in gene expression and is suggested to be one of the key pathophysiological factors of Parkinson’s disease (PD). Here, we found that α-synuclein preformed fibrils (PFFs) induced histone H3 dimethylation at lysine 9 (H3K9me2) and increased the euchromatic histone methyltransferases EHMT1 and EHMT2, which were accompanied by neuronal synaptic damage, including loss of synapses and diminished expression levels of synaptic-related proteins. Furthermore, the levels of H3K9me2 at promoters in genes that encode the synaptic-related proteins SNAP25, PSD95, Synapsin 1 and vGLUT1 were increased in primary neurons after PFF treatment, which suggests a linkage between H3K9 dimethylation and synaptic dysfunction. Inhibition of EHMT1/2 with the specific inhibitor A-366 or shRNA suppressed histone methylation and alleviated synaptic damage in primary neurons that were treated with PFFs. In addition, the synaptic damage and motor impairment in mice that were injected with PFFs were repressed by treatment with the EHMT1/2 inhibitor A-366. Thus, our findings reveal the role of histone H3 modification by EHMT1/2 in synaptic damage and motor impairment in a PFF animal model, suggesting the involvement of epigenetic dysregulation in PD pathogenesis.

Neuroprotective effect of green tea extract (-)-epigallocatechin-3-gallate in a preformed fibril-induced mouse model of Parkinson's disease.

Parkinson's disease (PD) is the second most common neurodegenerative disease characterized by the progressive degeneration of dopaminergic neurons in the substantia nigra (SN). The main bioactive component of green tea polyphenols (-)-epigallocatechin-3-gallate (EGCG) exerts protective effects against diseases such as neurodegenerative diseases and cancer. Therefore, this study investigated the effect of EGCG on the amelioration of neural damage in a chronic PD mouse model induced by α-synuclein preformed fibrils (α-syn-PFFs). A total of 20 C57BL/6J female mice were randomly divided into 3 groups: control group (saline, n = 6), model group (PFFs, n = 7), and prevention group (EGCG+PFFs, n = 7). A chronic PD mouse model was obtained by the administration of α-syn-PFFs by stereotaxic localization in the striatum. Behavioral tests were performed to evaluate PD-related anxiety-like behavior and motor impairments in the long-term PD progression. Tyrosine hydroxylase (TH) immuno-positive neurons and Ser129-phosphorylated α-syn (p-α-syn) were identified by immunohistochemistry. Pro-inflammatory and anti-inflammatory cytokines were measured by real-time quantitative PCR. EGCG pretreatment reduced anxiety-like behavior and motor impairments as revealed by the long-term behavioral test (2 weeks, 1 month, 3 months, and 6 months) on PD mice. EGCG also ameliorated PFF-induced degeneration of TH immuno-positive neurons and accumulation of p-α-syn in the SN and striatum at 6 months. Additionally, EGCG reduced the expression of pro-inflammatory cytokines while promoting the release of anti-inflammatory cytokines. EGCG exerts a neuroprotective effect on long-term progression of the PD model.

Integration of National Health Insurance claims data and animal models reveals fexofenadine as a promising repurposed drug for Parkinson's disease.

Parkinson's disease (PD) is a common and costly progressive neurodegenerative disease of unclear etiology. A disease-modifying approach that can directly stop or slow its progression remains a major unmet need in the treatment of PD. A clinical pharmacology-based drug repositioning strategy is a useful approach for identifying new drugs for PD. We analyzed claims data obtained from the National Health Insurance Service (NHIS), which covers a significant portion of the South Korean population, to investigate the association between antihistamines, a class of drugs commonly used to treat allergic symptoms by blocking H1 receptor, and PD in a real-world setting. Additionally, we validated this model using various animal models of PD such as the 6-hydroxydopmaine (6-OHDA), α-synuclein preformed fibrils (PFF) injection, and Caenorhabditis elegans (C. elegans) models. Finally, whole transcriptome data and Ingenuity Pathway Analysis (IPA) were used to elucidate drug mechanism pathways. We identified fexofenadine as the most promising candidate using National Health Insurance claims data in the real world. In several animal models, including the 6-OHDA, PFF injection, and C. elegans models, fexofenadine ameliorated PD-related pathologies. RNA-seq analysis and the subsequent experiments suggested that fexofenadine is effective in PD via inhibition of peripheral immune cell infiltration into the brain. Fexofenadine shows promise for the treatment of PD, identified through clinical data and validated in diverse animal models. This combined clinical and preclinical approach offers valuable insights for developing novel PD therapeutics.

Phosphorylated α-synuclein deposited in Schwann cells interacting with TLR2 mediates cell damage and induces Parkinson’s disease autonomic dysfunction

Despite the significant frequency of autonomic dysfunction (AutD) in Parkinson’s disease (PD) patients, its pathogenesis and diagnosis are challenging. Here, we aimed to further explore the mechanism of phosphorylated α-synuclein (p-α-syn) deposited in vagus nerve Schwann cells (SCs) causing SCs damage and PD AutD. 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP, 20 mg/kg) was administrated to C57BL/6 mice twice a week for 35 days. Following the final injection, locomotor functions, gastrointestinal symptoms, urine functions, and cardiovascular system functions were evaluated. Meanwhile, we examined p-α-syn deposited in vagus nerve SCs, Toll-like receptor 2 (TLR2) activation, and SCs loss using immunofluorescence, western blot, and Luxol fast blue staining. In vitro, the rat SCs line RSC96 cells were exposed to α-synuclein preformed fibril (α-syn PFF), and cell viability was detected by CCK8. Co-IP was used to identify the interaction between p-α-syn and TLR2. Furthermore, the role of TLR2 in p-α-syn-mediated SCs damage was confirmed by the administration of CU-CPT22, a specific blocker of TLR2. In vivo, apart from dyskinesia, MPTP mice exhibited constipation, urinary dysfunction, and cardiovascular failure, which were associated with the deposition of p-α-syn in vagus nerve SCs, TLR2 activation, and vagus nerve demyelination. In vitro, stimulation of α-syn PFF induced a time-dependent loss of viability, and p-α-syn deposited in RSC96 cells induced a cellular inflammatory response by interacting with TLR2, resulting in cell dysfunction and apoptosis. However, both SCs inflammatory response and cell viability were alleviated after inhibition of TLR2. Furthermore, 1 h fecal pellets and water content, the frequency of 1 h urine, blood pressure, heart rate, and heart rate variability of mice in the MPTP + CU-CPT22 group were also improved. Our results support the perspective that p-α-syn interacts with TLR2 induced SCs damage and is involved in PD AutD, which sheds fresh light on the mechanism of PD AutD and indicates a promising treatment for PD AutD targeting SCs p-α-syn/ TLR2 signaling pathway.

Toward the quantification of α-synuclein aggregates with digital seed amplification assays.

The quantification and characterization of aggregated α-synuclein in clinical samples offer immense potential toward diagnosing, treating, and better understanding neurodegenerative synucleinopathies. Here, we developed digital seed amplification assays to detect single α-synuclein aggregates by partitioning the reaction into microcompartments. Using pre-formed α-synuclein fibrils as reaction seeds, we measured aggregate concentrations as low as 4 pg/mL. To improve our sensitivity, we captured aggregates on antibody-coated magnetic beads before running the amplification reaction. By first characterizing the pre-formed fibrils with transmission electron microscopy and size exclusion chromatography, we determined the specific aggregates targeted by each assay platform. Using brain tissue and cerebrospinal fluid samples collected from patients with Parkinson's Disease and multiple system atrophy, we demonstrated that the assay can detect endogenous pathological α-synuclein aggregates. Furthermore, as another application for these assays, we studied the inhibition of α-synuclein aggregation in the presence of small-molecule inhibitors and used a custom image analysis pipeline to quantify changes in aggregate growth and filament morphology.

Host-to-graft propagation of inoculated α-synuclein into transplanted human induced pluripotent stem cell-derived midbrain dopaminergic neurons

IntroductionCell therapeutic clinical trials using fetal mesencephalic tissue provided a proof-of-concept for regenerative therapy in patients with Parkinson's disease. Postmortem studies of patients with fetal grafts revealed that α-synuclein+ Lewy body (LB)-like inclusions emerged in long-term transplantation and might worsen clinical outcomes even if the grafts survived and innervated in the recipients. Various studies aimed at addressing whether host-derived α-synuclein could be transferred to the grafted neurons to assess α-synuclein+ inclusion appearance in the grafts. However, determining whether α-synuclein in the grafted neurons has been propagated from the host is difficult due to the intrinsic α-synuclein expression. MethodsWe induced midbrain dopaminergic (mDA) neurons from human induced pluripotent stem cells (hiPSCs) and transplanted them into the striatum of immunodeficient rats. The recombinant human α-synuclein preformed fibrils (PFFs) were inoculated into the cerebral cortex after transplantation of SNCA−/− hiPSC-derived mDA neural progenitors into the striatum of immunodeficient rats to evaluate the host-to-graft propagation of human α-synuclein PFFs. Additionally, we examined the incorporation of human α-synuclein PFFs into SNCA−/− hiPSC-derived mDA neurons using in vitro culture system. ResultsWe detected human α-synuclein-immunoreactivity in SNCA−/− hiPSC-derived mDA neurons that lacked endogenous α-synuclein expression in vitro. Additionally, we observed host-to-graft α-synuclein propagation into the grafted SNCA−/− hiPSC-derived mDA neurons. ConclusionWe have successfully proven that intracerebral inoculated α-synuclein PFFs are propagated and incorporated from the host into grafted SNCA−/− hiPSC-derived mDA neurons. Our results contribute toward the basic understanding of the molecular mechanisms related to LB-like α-synuclein deposit formation in grafted mDA neurons.

The Impact of Intestinal Microbiota and Toll-like Receptor 2 Signaling on α-Synuclein Pathology in Nontransgenic Mice Injected with α-Synuclein Preformed Fibrils.

Intestinal microbiota and Toll-like receptor 2 (TLR2), which can bind lipoteichoic acid produced by microbiota, might contribute to the pathogenesis of Parkinson's disease (PD), which is characterized by α-synuclein accumulation. Although the contribution of intestinal microbiota and TLR2 to PD pathology was validated in genetic PD models, evidence suggests that the effects of TLR2 signaling on proteinopathy might depend on the presence of a genetic etiology. We examined the impact of intestinal microbiota and TLR2 signaling on α-synuclein pathology in a nontransgenic mouse model of sporadic PD. While an α-synuclein preformed fibrils injection successfully reproduced PD pathology by inducing accumulation of α-synuclein aggregates, microglial activation and increased TLR2 expression in the brains of nontransgenic mice, antibiotic-induced reduction in the density of intestinal microbiota and TLR2 knockout had small impact on these changes. These findings, which are in contrast to those reported in transgenic mice harboring transgene encoding α-synuclein, indicate that the contribution of intestinal microbiota and TLR2 signaling to α-synuclein pathogenesis might be influenced by the presence of a genetic etiology. Additionally, these findings suggest that integrating insights from this experimental model and genetic models would further advance our understanding of the molecular mechanisms underlying sporadic PD.

Interspecies chimerism with human embryonic stem cells generates functional human dopamine neurons at low efficiency

Interspecies chimeras offer great potential for regenerative medicine and the creation of human disease models. Whether human pluripotent stem cell-derived neurons in an interspecies chimera can differentiate into functional neurons and integrate into host neural circuity is not known. Here, we show, using Engrailed 1 (En1) as a development niche, that human naive-like embryonic stem cells (ESCs) can incorporate into embryonic and adult mouse brains. Human-derived neurons including tyrosine hydroxylase (TH)+ neurons integrate into the mouse brain at low efficiency. These TH+ neurons have electrophysiologic properties consistent with their human origin. In addition, these human-derived neurons in the mouse brain accumulate pathologic phosphorylated α-synuclein in response to α-synuclein preformed fibrils. Optimization of human/mouse chimeras could be used to study human neuronal differentiation and human brain disorders.

6PPD-quinone exposure induces neuronal mitochondrial dysfunction to exacerbate Lewy neurites formation induced by α-synuclein preformed fibrils seeding

The emerging toxicant N-(1,3-dimethylbutyl)-N′-phenyl-p-phenylenediamine quinone (6PPD-Q) is of wide concern due to its ubiquitous occurrence and high toxicity. Despite regular human exposure, limited evidence exists about its presence in the body and potential health risks. Herein, we analyzed cerebrospinal fluid (CSF) samples from Parkinson's disease (PD) patients and controls. The CSF levels of 6PPD-Q were twice as high in PD patients compared to controls. Immunostaining assays performed with primary dopaminergic neurons confirm that 6PPD-Q at environmentally relevant concentrations can exacerbate the formation of Lewy neurites induced by α-synuclein preformed fibrils (α-syn PFF). Assessment of cellular respiration reveals a considerable decrease in neuronal spare respiratory and ATP-linked respiration, potentially due to changes in mitochondrial membrane potential. Moreover, 6PPD-Q-induced mitochondrial impairment correlates with an upsurge in mitochondrial reactive oxygen species (mROS), and Mito-TEMPO-driven scavenging of mROS can lessen the amount of pathologic phospho-serine 129 α-synuclein. Untargeted metabolomics provides supporting evidence for the connection between 6PPD-Q exposure and changes in neuronal metabolite profiles. In-depth targeted metabolomics further unveils an overall reduction in glycolysis metabolite pool and fluctuations in the quantity of TCA cycle intermediates. Given its potentially harmful attributes, the presence of 6PPD-Q in human brain could potentially be a risk factor for PD.

MLKL deficiency alleviates neuroinflammation and motor deficits in the α-synuclein transgenic mouse model of Parkinson’s disease

Parkinson’s disease (PD), one of the most devastating neurodegenerative brain disorders, is characterized by the progressive loss of dopaminergic neurons in the substantia nigra (SN) and deposits of α-synuclein aggregates. Currently, pharmacological interventions for PD remain inadequate. The cell necroptosis executor protein MLKL (Mixed-lineage kinase domain-like) is involved in various diseases, including inflammatory bowel disease and neurodegenerative diseases; however, its precise role in PD remains unclear. Here, we investigated the neuroprotective role of MLKL inhibition or ablation against primary neuronal cells and human iPSC-derived midbrain organoids induced by toxic α-Synuclein preformed fibrils (PFFs). Using a mouse model (Tg-Mlkl−/−) generated by crossbreeding the SNCA A53T synuclein transgenic mice with MLKL knockout (KO)mice, we assessed the impact of MLKL deficiency on the progression of Parkinsonian traits. Our findings demonstrate that Tg-Mlkl−/− mice exhibited a significant improvement in motor symptoms and reduced phosphorylated α-synuclein expression compared to the classic A53T transgenic mice. Furthermore, MLKL deficiency alleviated tyrosine hydroxylase (TH)-positive neuron loss and attenuated neuroinflammation by inhibiting the activation of microglia and astrocytes. Single-cell RNA-seq (scRNA-seq) analysis of the SN of Tg-Mlkl−/− mice revealed a unique cell type-specific transcriptome profile, including downregulated prostaglandin D synthase (PTGDS) expression, indicating reduced microglial cells and dampened neuron death. Thus, MLKL represents a critical therapeutic target for reducing neuroinflammation and preventing motor deficits in PD.

Enhanced mTORC1 signaling and protein synthesis in pathologic α-synuclein cellular and animal models of Parkinson's disease.

Pathologic α-synuclein plays an important role in the pathogenesis of α-synucleinopathies such as Parkinson's disease (PD). Disruption of proteostasis is thought to be central to pathologic α-synuclein toxicity; however, the molecular mechanism of this deregulation is poorly understood. Complementary proteomic approaches in cellular and animal models of PD were used to identify and characterize the pathologic α-synuclein interactome. We report that the highest biological processes that interacted with pathologic α-synuclein in mice included RNA processing and translation initiation. Regulation of catabolic processes that include autophagy were also identified. Pathologic α-synuclein was found to bind with the tuberous sclerosis protein 2 (TSC2) and to trigger the activation of the mammalian target of rapamycin (mTOR) complex 1 (mTORC1), which augmented mRNA translation and protein synthesis, leading to neurodegeneration. Genetic and pharmacologic inhibition of mTOR and protein synthesis rescued the dopamine neuron loss, behavioral deficits, and aberrant biochemical signaling in the α-synuclein preformed fibril mouse model and Drosophila transgenic models of pathologic α-synuclein-induced degeneration. Pathologic α-synuclein furthermore led to a destabilization of the TSC1-TSC2 complex, which plays an important role in mTORC1 activity. Constitutive overexpression of TSC2 rescued motor deficits and neuropathology in α-synuclein flies. Biochemical examination of PD postmortem brain tissues also suggested deregulated mTORC1 signaling. These findings establish a connection between mRNA translation deregulation and mTORC1 pathway activation that is induced by pathologic α-synuclein in cellular and animal models of PD.

α-Synuclein pathology from the body to the brain: so many seeds so close to the central soil.

α-Synuclein is a protein that mainly exists in the presynaptic terminals. Abnormal folding and accumulation of α-synuclein are found in several neurodegenerative diseases, including Parkinson's disease. Aggregated and highly phosphorylated α-synuclein constitutes the main component of Lewy bodies in the brain, the pathological hallmark of Parkinson's disease. For decades, much attention has been focused on the accumulation of α-synuclein in the brain parenchyma rather than considering Parkinson's disease as a systemic disease. Recent evidence demonstrates that, at least in some patients, the initial α-synuclein pathology originates in the peripheral organs and spreads to the brain. Injection of α-synuclein preformed fibrils into the gastrointestinal tract triggers the gut-to-brain propagation of α-synuclein pathology. However, whether α-synuclein pathology can occur spontaneously in peripheral organs independent of exogenous α-synuclein preformed fibrils or pathological α-synuclein leakage from the central nervous system remains under investigation. In this review, we aimed to summarize the role of peripheral α-synuclein pathology in the pathogenesis of Parkinson's disease. We also discuss the pathways by which α-synuclein pathology spreads from the body to the brain.

Synaptotagmin-11 regulates immune functions of microglia invivo.

Membrane trafficking pathways mediate key microglial activities such as cell migration, cytokine secretion, and phagocytosis. However, the underlying molecular mechanism remains poorly understood. Previously, we found that synaptotagmin-11 (Syt11), a non-Ca2+ -binding Syt associated with Parkinson's disease (PD) and schizophrenia, inhibits cytokine release and phagocytosis in primary microglia. Here we reported the invivo function of Syt11 in microglial immune responses using an inducible microglia-specific Syt11-conditional-knockout (cKO) mouse strain. Syt11-cKO resulted in activation of microglia and elevated mRNA levels of IL-6, TNF-α, IL-1β, and iNOS in various brain regions under both resting state and LPS-induced acute inflammation state in adult mice. In a PD mouse model generated by microinjection of preformed α-synuclein fibrils into the striatum, a reduced number of microglia migrated toward the injection sites and an enhanced phagocytosis of α-synuclein fibrils by microglia were found in Syt11-cKO mice. To understand the molecular mechanism of Syt11 function, we identified its direct binding proteins vps10p-tail-interactor-1a (vti1a) and vti1b. The linker domain of Syt11 interacted with both proteins and a peptide derived from it competitively inhibited the interaction of Syt11 with vti1a/vti1b invitro and in cells. Importantly, application of this peptide induced more cytokine secretion in wild-type microglia upon LPS treatment, phenocopying defects in Syt11 knockdown cells. Altogether, we propose that Syt11 inhibits microglial activation invivo and regulates cytokine secretion through interactions with vti1a and vti1b.

Formation of templated inclusions in a forebrain α-synuclein mouse model is independent of LRRK2

Leucine-rich repeat kinase 2 (LRRK2) and α-synuclein share enigmatic roles in the pathobiology of Parkinson's disease (PD). LRRK2 mutations are a common genetic cause of PD which, in addition to neurodegeneration, often present with abnormal deposits of α-synuclein in the form of Lewy-related pathology. As Lewy-related pathology is a prominent neuropathologic finding in sporadic PD, the relationship between LRRK2 and α-synuclein has garnered considerable interest. However, whether and how LRRK2 might influence the accumulation of Lewy-related pathology remains poorly understood. Through stereotactic injection of mouse α-synuclein pre-formed fibrils (PFF), we modeled the spread of Lewy-related pathology within forebrain regions where LRRK2 is most highly expressed. The impact of LRRK2 genotype on the formation of α-synuclein inclusions was evaluated at 1-month post-injection. Neither deletion of LRRK2 nor G2019S LRRK2 knockin appreciably altered the burden of α-synuclein pathology at this early timepoint. These observations fail to provide support for a robust pathophysiologic interaction between LRRK2 and α-synuclein in the forebrain in vivo. There was, however, a modest reduction in microglial activation induced by PFF delivery in the hippocampus of LRRK2 knockout mice, suggesting that LRRK2 may contribute to α-synuclein-induced neuroinflammation. Collectively, our data indicate that the pathological accumulation of α-synuclein in the mouse forebrain is largely independent of LRRK2.

Modelling α-synuclein processing in primary patient cells for pharmacological intervention

Aim: Parkinson’s disease (PD) is a complex, chronic neurodegenerative disorder with predominately sporadic etiology. Intricate genetic-environmental interactions lead to the hallmarks of the disease: degeneration of dopaminergic neurons and the deposition of α-synuclein aggregates. The aim of this study was to establish a novel primary patient cell model as an in vitro screen to study α-synuclein processing for drug screening. Methods: Primary patient olfactory neuroepithelial-derived cells (ONS) were exposed to α-synuclein and examined for cytotoxicity, processing, and solubility over 48 h. Epigallocatechin gallate (EGCG), which is known to destabilise α-synuclein fibrils, was used to investigate the solubilisation of α-synuclein in the model system. Results: Exposure to 0.1 μmol/L α-synuclein preformed fibrils was not toxic to ONS over 48 h. ONS processing of α-synuclein was observed to be different in PD cells by their increased accumulation in the cytoplasm. Processing deficits in the PD ONS were confirmed by immunoblotting with an increase in sodium dodecyl sulfate (SDS)-insoluble α-synuclein after 48 h. Conclusions: The data has illustrated the utility of primary patient ONS as a model system to understand the processing of α-synuclein. Considerable differences in α-synuclein processing were identified in PD ONS. Furthermore, the data suggests that primary patient ONS are a viable in vitro drug screening platform for α-synuclein pathology in PD.

Dopaminergic neurodegeneration in the substantia nigra is associated with olfactory dysfunction in mice models of Parkinson’s disease

Olfactory dysfunction represents a prodromal stage in Parkinson’s disease (PD). However, the mechanisms underlying hyposmia are not specified yet. In this study, we first observed an early olfactory dysfunction in mice with intragastric rotenone administration, consistent with dopaminergic neurons loss and α-synuclein pathology in the olfactory bulb. However, a much severer olfactory dysfunction was observed without severer pathology in olfactory bulb when the loss of dopaminergic neurons in the substantia nigra occurred. Then, we established the mice models by intrastriatal α-synuclein preformed fibrils injection and demonstrated the performance in the olfactory discrimination test was correlated to the loss of dopaminergic neurons in the substantia nigra, without any changes in the olfactory bulb analyzed by RNA-sequence. In mice with intranasal ferric ammonium citrate administration, we observed olfactory dysfunction when dopaminergic neurodegeneration in substantia nigra occurred and was restored when dopaminergic neurons were rescued. Finally we demonstrated that chemogenetic inhibition of dopaminergic neurons in the substantia nigra was sufficient to cause hyposmia and motor incoordination. Taken together, this study shows a direct relationship between nigral dopaminergic neurodegeneration and olfactory dysfunction in PD models and put forward the understandings that olfactory dysfunction represents the early stage of neurodegeneration in PD progression.

α-Synuclein propagation leads to synaptic abnormalities in the cortex through microglial synapse phagocytosis

The major neuropathologic feature of Parkinson’s disease is the presence of widespread intracellular inclusions of α-synuclein known as Lewy bodies. Evidence suggests that these misfolded protein inclusions spread through the brain with disease progression. Changes in synaptic function precede neurodegeneration, and this extracellular α-synuclein can affect synaptic transmission. However, whether and how the spreading of α-synuclein aggregates modulates synaptic function before neuronal loss remains unknown. In the present study, we investigated the effect of intrastriatal injection of α-synuclein preformed fibrils (PFFs) on synaptic activity in the somatosensory cortex using a combination of whole-cell patch-clamp electrophysiology, histology, and Golgi-Cox staining. Intrastriatal PFF injection was followed by formation of phosphorylated α-synuclein inclusions in layer 5 of the somatosensory cortex, leading to a decrease in synapse density, dendritic spines, and spontaneous excitatory post-synaptic currents, without apparent neuronal loss. Additionally, three-dimensional reconstruction of microglia using confocal imaging showed an increase in the engulfment of synapses. Collectively, our data indicate that propagation of α-synuclein through neural networks causes abnormalities in synaptic structure and dynamics prior to neuronal loss.

Developmental exposure to the Parkinson's disease-associated organochlorine pesticide dieldrin alters dopamine neurotransmission in α-synuclein pre-formed fibril (PFF)-injected mice.

Parkinson's disease (PD) is the fastest-growing neurological disease worldwide, with increases outpacing aging and occurring most rapidly in recently industrialized areas, suggesting a role of environmental factors. Epidemiological, post-mortem, and mechanistic studies suggest that persistent organic pollutants, including the organochlorine pesticide dieldrin, increase PD risk. In mice, developmental dieldrin exposure causes male-specific exacerbation of neuronal susceptibility to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and synucleinopathy. Specifically, in the α-synuclein (α-syn) pre-formed fibril (PFF) model, exposure leads to increased deficits in striatal dopamine (DA) turnover and motor deficits on the challenging beam. Here, we hypothesized that alterations in DA handling contribute to the observed changes and assessed vesicular monoamine transporter 2 (VMAT2) function and DA release in this dieldrin/PFF 2-hit model. Female C57BL/6 mice were exposed to 0.3 mg/kg dieldrin or vehicle every 3 days by feeding, starting at 8 weeks of age and continuing throughout breeding, gestation, and lactation. Male offspring from independent litters underwent unilateral, intrastriatal injections of α-syn PFFs at 12 weeks of age, and vesicular 3H-DA uptake assays and fast-scan cyclic voltammetry were performed 4 months post-PFF injection. Dieldrin-induced an increase in DA release in striatal slices in PFF-injected animals, but no change in VMAT2 activity. These results suggest that developmental dieldrin exposure increases a compensatory response to synucleinopathy-triggered striatal DA loss. These findings are consistent with silent neurotoxicity, where developmental exposure to dieldrin primes the nigrostriatal striatal system to have an exacerbated response to synucleinopathy in the absence of observable changes in typical markers of nigrostriatal dysfunction and degeneration.