Differential Effects of Paraquat, Rotenone, and MPTP on Cellular Bioenergetics of Undifferentiated and Differentiated Human Neuroblastoma Cells.

Paraquat (PQ), rotenone (RO), and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) are neurotoxicants that can damage human health. Exposure to these neurotoxicants has been linked to neurodegeneration, particularly Parkinson's disease. However, their mechanisms of action have not been fully elucidated, nor has the relative vulnerability of neuronal subtypes to their exposures. To address this, the current study investigated the cytotoxic effects of PQ, RO, and MPTP and their relative effects on cellular bioenergetics and oxidative stress on undifferentiated human neuroblastoma (SH-SY5Y) cells and those differentiated to dopaminergic (DA) or cholinergic (CH) phenotypes. The tested neurotoxicants were all cytotoxic to the three cell phenotypes that correlated with both concentration and exposure duration. At half-maximal effective concentrations (EC50s), there were significant reductions in cellular ATP levels and reduced activity of the mitochondrial complexes I and III, with a parallel increase in lactate production. PQ at 10 µM significantly decreased ATP production and mitochondrial complex III activity only in DA cells. RO was the most potent inhibitor of mitochondrial complex 1 and did not inhibit mitochondrial complex III even at concentrations that induced a 50% loss of cell viability. MPTP was the most potent toxicant in undifferentiated cells. All neurotoxicants significantly increased reactive oxygen species, lipid peroxidation, and nuclear expression of Nrf2, with a corresponding inhibition of the antioxidant enzymes catalase and superoxide dismutase. At a 10 µM exposure to PQ or RO, oxidative stress biomarkers were significant in DA cells. Collectively, this study underscores the importance of mitochondrial dysfunction and oxidative stress in PQ, RO, and MPTP-induced cytotoxicity and that neuronal phenotypes display differential vulnerability to these neurotoxicants.

Whole Transcriptome Analysis of Substantia Nigra in Mice with MPTP-Induced Parkinsonism Bearing Defective Glucocerebrosidase Activity.

Mutations in the GBA1 gene represent the major genetic risk factor for Parkinson's disease (PD). The lysosomal enzyme beta-glucocerebrosidase (GCase) encoded by the GBA1 gene participates in both the endolysosomal pathway and the immune response. Disruption of these mechanisms is involved in PD pathogenesis. However, molecular mechanisms of PD associated with GBA1 mutations (GBA-PD) are unknown today in particular due to the partial penetrance of GBA1 variants in PD. The modifiers of GBA1 penetrance have not been elucidated. We characterized the transcriptomic profiles of cells from the substantia nigra (SN) of mice with co-injection with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and selective inhibitor of GCase activity (conduritol-ß-epoxide, (CBE)) to mimic PD bearing GCase dysfunction (MPTP+CBE), mice treated with MPTP, mice treated with CBE and control mice treated with injection of sodium chloride (NaCl) (vehicle). Differential expression analysis, pathway enrichment analysis, and outlier detection were performed. Functional clustering of differentially represented transcripts revealed more processes associated with the functioning of neurogenesis, inflammation, apoptosis and autophagy in MPTP+CBE and MPTP mice than in vehicle mice, with a more pronounced alteration of autophagy processes in MPTP+CBE mice than in MPTP mice. The PI3K-Akt-mTOR signaling pathway may be considered a potential target for therapy in PD with GCase dysfunction.

Vitamin E Analog Trolox Attenuates MPTP-Induced Parkinson's Disease in Mice, Mitigating Oxidative Stress, Neuroinflammation, and Motor Impairment.

Trolox is a potent antioxidant and a water-soluble analog of vitamin E. It has been used in scientific studies to examine oxidative stress and its impact on biological systems. Trolox has been shown to have a neuroprotective effect against ischemia and IL-1ß-mediated neurodegeneration. In this study, we investigated the potential protective mechanisms of Trolox against a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced Parkinson's disease mouse model. Western blotting, immunofluorescence staining, and ROS/LPO assays were performed to investigate the role of trolox against neuroinflammation, the oxidative stress mediated by MPTP in the Parkinson's disease (PD) mouse model (wild-type mice (C57BL/6N), eight weeks old, average body weight 25-30 g). Our study showed that MPTP increased the expression of α-synuclein, decreased tyrosine hydroxylase (TH) and dopamine transporter (DAT) levels in the striatum and substantia nigra pars compacta (SNpc), and impaired motor function. However, Trolox treatment significantly reversed these PD-like pathologies. Furthermore, Trolox treatment reduced oxidative stress by increasing the expression of nuclear factor erythroid-2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1). Lastly, Trolox treatment inhibited the activated astrocytes (GFAP) and microglia (Iba-1), also reducing phosphorylated nuclear factor-κB, (p-NF-κB) and tumor necrosis factor-alpha (TNF-α) in the PD mouse brain. Overall, our study demonstrated that Trolox may exert neuroprotection on dopaminergic neurons against MPTP-induced oxidative stress, neuroinflammation, motor dysfunction, and neurodegeneration.

The transient receptor potential melastatin 2: a new therapeutical target for Parkinson's disease?

The transient receptor potential melastatin 2 is a calcium-permeable cation channel member of the TRP family. Also known as an oxidative stress-activated channel, the transient receptor potential melastatin 2 gating mechanism is dependent on reactive oxygen species. In pathological conditions, transient receptor potential melastatin 2 is overactivated, leading to a Ca2+ influx that alters cell homeostasis and promotes cell death. The role of transient receptor potential melastatin 2 in neurodegenerative diseases, including Alzheimer's disease and ischemia, has already been described and reviewed. However, data on transient receptor potential melastatin 2 involvement in Parkinson's disease pathology has emerged only in recent years and the issue lacks review studies that focus specifically on this topic. The present review aims to elucidate the role of the transient receptor potential melastatin 2 channel in Parkinson's disease by reviewing, summarizing, and discussing the in vitro, in vivo, and human studies published until August 2022. Here we describe fourteen studies that evaluated the transient receptor potential melastatin 2 channel in Parkinson's disease. The Parkinson's disease model used, transient receptor potential melastatin 2 antagonist and genetic approaches, and the main outcomes reported were discussed. The studies described transient receptor potential melastatin 2 activation and enhanced expression in different Parkinson's disease models. They also evidenced protective and restorative effects when using transient receptor potential melastatin 2 antagonists, knockout, or silencing. This review provides a literature overview and suggests where there is a need for more research. As a perspective point, this review shows evidence that supports transient receptor potential melastatin 2 as a pharmacological target for Parkinson's disease in the future.

Assay of MAO Inhibition by Chromatographic Techniques (HPLC/HPLC-MS).

Monoamine oxidase (MAO) enzymes (MAO A and B) catalyze the oxidative deamination of biogenic amines, neurotransmitters, and xenobiotic amines and contribute to the regulation of the content of these active substances in mammalian organisms. The oxidation of biogenic amines by MAO produces hydrogen peroxide (H2O2) and aldehydes that represent risk factors for oxidative injury. The inhibitors of MAO are useful as antidepressants and neuroprotective agents. Usually, the assays of MAO determine amine deamination products or measure the H2O2 released by using direct spectrophotometric or fluorimetric methods. Direct methods are more prone to interferences and can afford inaccurate results. Those limitations can be avoided by using chromatographic techniques. This work describes a chromatographic method to assay MAO A and MAO B activity by using kynuramine as a nonselective substrate and the subsequent analysis of 4-hydroxyquinoline by RP-HPLC-DAD-fluorescence and mass spectrometry (MS). Alternatively, the assay uses the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) neurotoxin as a substrate of MAO that is oxidized (bioactivated) to neurotoxic pyridinium cations which are analyzed by HPLC. These methods are applied to assess the inhibition of MAO by bioactive ß-carboline alkaloids occurring in foods, plants, and biological systems.

G-CSF reduces loss of dopaminergic neurons by inhibiting TNF-α and IL-1ß in mouse model of Parkinson's disease.

PURPOSE OF THE STUDY: granulocyte-colony stimulating factor (G-CSF) is a hematopoietic growth factor existing in neutrophils, glial cells and neurons. Increasing researches discovered that G-CSF improved cell survival in neurodegenerative diseases by its anti-inflammatory effect. However, the effect of G-CSF in suppressing inflammation in Parkinson's disease (PD) remains unclear. Thus, the purpose of this study is to explored the anti-inflammatory effect of G-CSF in mouse model of PD. MATERIALS AND METHODS: G-CSF was administrated in the PD model induced by MPTP. Subsequently, the protein of tyrosine hydroxylase (TH), ionized calcium-binding adaptor molecule 1 (Iba-1) and the inflammatory cytokines including tumor necrosis factor-alpha (TNF-α) and interleukin-1 beta (IL-1ß) in the midbrain were examined. In addition, the phosphorylated mitogen-activated protein kinases (MAPK) including c-Jun NH2-terminal kinase (JNK), extracellular signal-regulated kinase (ERK) and p38 MAPK in the midbrain were investigated. RESULTS: Compared with the MPTP group, the protein of TH in the midbrain was increased, while the Iba-1 and the inflammatory factors were decreased. In addition, the expression of phosphorylated JNK (p-JNK) in the midbrain of the MPTP + G-CSF group was decreased, while the phosphorylated ERK (p-ERK) levels were elevated. CONCLUSIONS: These findings emphasize that G-CSF inhibited the degradation of DA neurons. The protective effect is associated with the reduction of the inflammatory factors caused by the inhibition of the microglial activation. Moreover, G-CSF may decrease the inflammatory factors through the decrease of P-JNK and the increase of P-ERK.

Neuroprotective Effects of Nicotinamide against MPTP-Induced Parkinson's Disease in Mice: Impact on Oxidative Stress, Neuroinflammation, Nrf2/HO-1 and TLR4 Signaling Pathways.

Nicotinamide (NAM) is the amide form of niacin and an important precursor of nicotinamide adenine dinucleotide (NAD), which is needed for energy metabolism and cellular functions. Additionally, it has shown neuroprotective properties in several neurodegenerative diseases. Herein, we sought to investigate the potential protective mechanisms of NAM in an intraperitoneal (i.p) 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced Parkinson's disease (PD) mouse model (wild-type mice (C57BL/6N), eight weeks old, average body weight 25-30 g). The study had four groups (n = 10 per group): control, MPTP (30 mg/kg i.p. for 5 days), MPTP treated with NAM (500 mg/kg, i.p for 10 days) and control treated with NAM. Our study showed that MPTP increased the expression of α-synuclein 2.5-fold, decreased tyrosine hydroxylase (TH) 0.5-fold and dopamine transporters (DAT) levels up to 0.5-fold in the striatum and substantia nigra pars compacta (SNpc), and impaired motor function. However, NAM treatment significantly reversed these PD-like pathologies. Furthermore, NAM treatment reduced oxidative stress by increasing the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) between 0.5- and 1.0-fold. Lastly, NAM treatment regulated neuroinflammation by reducing Toll-like receptor 4 (TLR-4), phosphorylated nuclear factor-κB, tumor (p-NFκB), and cyclooxygenase-2 (COX-2) levels by 0.5- to 2-fold in the PD mouse brain. Overall, these findings suggest that NAM exhibits neuroprotective properties and may be an effective therapeutic agent for PD.

Preventative effects of 1-methyl-1,2,3,4-tetrahydroisoquinoline derivatives (N-functional group loading) on MPTP-induced parkinsonism in mice.

1,2,3,4-tetrahydroisoquinoline (TIQ) is endogenously present in the human brain, and some of its derivatives are thought to contribute to the induction of Parkinson's disease (PD)-like signs in rodents and primates. In contrast, the endogenous TIQ derivative 1-methyl-TIQ (1-MeTIQ) is reported to be neuroprotective. In the present study, we compared the effects of artificially modified 1-MeTIQ derivatives (loading an N-propyl, N-propenyl, N-propargyl, or N-butynyl group) on 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD-like signs in mice. In a behavioral study, MPTP-induced bradykinesia was significantly decreased by all compounds. However, only 1-Me-N-propargyl-TIQ showed an inhibitory effect by blocking the MPTP-induced reduction in striatal dopamine content and the number of nigral tyrosine hydroxylase-positive cells. Western blot analysis showed that 1-Me-N-propargyl-TIQ and 1-Me-N-butynyl-TIQ potently prevented the MPTP-induced decrease in dopamine transporter expression, whereas 1-MeTIQ and 1-Me-N-propyl-TIQ did not. These results suggest that although loading an N-propargyl group on 1-MeTIQ clearly enhanced neuroprotective effects, other N-functional groups showed distinct pharmacological properties characteristic of their functional groups. Thus, the number of bonds and length of the N-functional group may contribute to the observed differences in effect.

MPTP-Treated Zebrafish Recapitulate 'Late-Stage' Parkinson's-like Cognitive Decline.

The zebrafish is a promising model species in biomedical research, including neurotoxicology and neuroactive drug screening. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) evokes degeneration of dopaminergic neurons and is commonly used to model Parkinson's disease (PD) in laboratory animals, including zebrafish. However, cognitive phenotypes in MPTP-evoked experimental PD models remain poorly understood. Here, we established an LD50 (292 mg/kg) for intraperitoneal MPTP administration in adult zebrafish, and report impaired spatial working memory (poorer spontaneous alternation in the Y-maze) in a PD model utilizing fish treated with 200 µg of this agent. In addition to conventional behavioral analyses, we also employed artificial intelligence (AI)-based approaches to independently and without bias characterize MPTP effects on zebrafish behavior during the Y-maze test. These analyses yielded a distinct cluster for 200-µg MPTP (vs. other) groups, suggesting that high-dose MPTP produced distinct, computationally detectable patterns of zebrafish swimming. Collectively, these findings support MPTP treatment in adult zebrafish as a late-stage experimental PD model with overt cognitive phenotypes.

Tauroursodeoxycholic acid (TUDCA) is neuroprotective in a chronic mouse model of Parkinson's disease.

OBJECTIVE: Parkinson's disease (PD) is a progressive motor disease of unknown etiology. Although neuroprotective ability of endogenous bile acid, tauroursodeoxycholic acid (TUDCA), shown in various diseases, including an acute model of PD,the potential therapeutic role of TUDCA in progressive models of PD that exhibit all aspects of PD has not been elucidated. In the present study, mice were assigned to one of four treatment groups: (1) Probenecid (PROB); (2) TUDCA, (3) MPTP + PROB (MPTPp); and (3) TUDCA + MPTPp. Methods: Markers for dopaminergic function, neuroinflammation, oxidative stress and autophagy were assessed using high performance liquid chromatography (HPLC), immunohistochemistry (IHC) and western blot (WB) methods. Locomotion was measured before and after treatments. Results: MPTPp decreased the expression of dopamine transporters (DAT) and tyrosine hydroxylase (TH), indicating dopaminergic damage, and induced microglial and astroglial activation as demonstrated by IHC analysis. MPTPp also decreased DA and its metabolites as demonstrated by HPLC analysis. Further, MPTPp-induced protein oxidation; increased LAMP-1 expression indicated autophagy and the promotion of alpha-synuclein (α-SYN) aggregation. Discussion: Pretreatment with TUDCA protected against dopaminergic neuronal damage, prevented the microglial and astroglial activation, as well as the DA and DOPAC reductions caused by MPTPp. TUDCA by itself did not produce any significant change, with data similar to the negative control group. Pretreatment with TUDCA prevented protein oxidation and autophagy, in addition to inhibiting α-SYN aggregation. Although TUDCA pretreatment did not significantly affect locomotion, only acute treatment effects were measured, indicating more extensive assessments may be necessary to reveal potential therapeutic effects on behavior. Together, these results suggest that autophagy may be involved in the progression of PD and that TUDCA may attenuate these effects. The efficacy of TUDCA as a novel therapy in patients with PD clearly warrants further study.

Generation of Mitochondrial Toxin Rodent Models of Parkinson's Disease Using 6-OHDA , MPTP , and Rotenone.

Several animal models are employed to discover novel treatments for the symptoms of Parkinson's disease (PD). PD models can be divided into two models: neurotoxin models and genetic models. Among neurotoxins to produce PD models, 6-hydroxydopamine (6-OHDA), 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), and rotenone, which inhibit the mitochondrial complex I, are widely used. Animal models of PD using these neurotoxins are also known as mitochondrial toxin models. Here this chapter describes the preparation of these mitochondrial toxin models.

Korean red ginseng suppresses 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced inflammation in the substantia nigra and colon.

Parkinson's disease (PD) is a neurodegenerative disease involving dopaminergic neuronal death in the substantia nigra (SN); recent studies have shown that interactions between gut and brain play a critical role in the pathogenesis of PD. In this study, the anti-inflammatory effect of Korean red ginseng (KRG) and the changes in gut microbiota were evaluated in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD mouse model. Male nine-week-old C57BL/6 mice were injected intraperitoneally with 30 mg/kg of MPTP at 24-h intervals for 5 days. Two hours after the daily MPTP injection, the mice were orally administered 100 mg/kg of KRG, which continued for 7 days beyond the MPTP injections, for a total of 12 consecutive days. Eight days after the final KRG administration, the pole and rotarod tests were performed and brain and colon samples of the mice were collected. Dopaminergic neuronal death, activation of microglia and astrocytes, α-synuclein and expressions of inflammatory cytokines and disruption of tight junction were evaluated. In addition, 16S ribosomal RNA gene sequencing of mouse fecal samples was performed to investigate microbiome changes. KRG treatment prevented MPTP-induced behavioral impairment, dopaminergic neuronal death, activation of microglia and astrocytes in the nigrostriatal pathway, disruption of tight junction and the increase in α-synuclein, interleukin-1ß and tumor necrosis factor-α expression in the colon. The 16S rRNA sequencing revealed that MPTP altered the number of bacterial species and their relative abundances, which were partially suppressed by KRG treatment. Especially, KRG suppressed the abundance of the inflammation-related phylum Verrucomicrobia and genera Ruminococcus and Akkermansia (especially Akkermansia muciniphila), and elevated the abundance of Eubacterium, which produces the anti-inflammatory substances. These findings suggest that KRG prevents MPTP-induced dopaminergic neuronal death, activation of microglia and astrocytes, and accumulation of α-synuclein in the SN, and the regulation of inflammation-related factors in the colon may influence the effect.

Mitochondrial E3 Ubiquitin Ligase Parkin: Relationships with Other Causal Proteins in Familial Parkinson's Disease and Its Substrate-Involved Mouse Experimental Models.

Parkinson's disease (PD) is a common neurodegenerative disorder. Recent identification of genes linked to familial forms of PD has revealed that post-translational modifications, such as phosphorylation and ubiquitination of proteins, are key factors in disease pathogenesis. In PD, E3 ubiquitin ligase Parkin and the serine/threonine-protein kinase PTEN-induced kinase 1 (PINK1) mediate the mitophagy pathway for mitochondrial quality control via phosphorylation and ubiquitination of their substrates. In this review, we first focus on well-characterized PINK1 phosphorylation motifs. Second, we describe our findings concerning relationships between Parkin and HtrA2/Omi, a protein involved in familial PD. Third, we describe our findings regarding inhibitory PAS (Per/Arnt/Sim) domain protein (IPAS), a member of PINK1 and Parkin substrates, involved in neurodegeneration during PD. IPAS is a dual-function protein involved in transcriptional repression of hypoxic responses and the pro-apoptotic activities.

A pilot study on assessment of locomotor behavior using a video tracking system in minipigs.

Pigs are often selected for large animal models including for neuroscience and behavioral research, because their anatomy and biochemistry are similar to those of humans. However, behavioral assessments, in combination with objective long-term monitoring, is difficult. In this study, we introduced an automated video tracking system which was previously used in rodent studies, for use with pig models. Locomotor behaviors (total distance, number of zone transitions, and velocity) were evaluated and their changes were validated by different 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) administration methods and dosing regimens. Three minipigs (23-29 kg) received subcutaneous or intravenous MPTP, either 1 or 3 times per week. Immediately after MPTP injection, the minipigs remained in a corner and exhibited reduced trajectory. In addition, the total distance travelled, number of zone transitions, and velocity were greatly reduced at every MPTP administration in all the minipigs, accompanying to increased resting time. However, the MPTP-induced symptoms were reversed when MPTP administration was terminated. In conclusion, this automated video-tracking system was able to monitor long-term locomotor activity and differentiate detailed alterations in large animals. It has the advantages of being easy to use, higher resolution, less effort, and more delicate tracking. Additionally, as our method can be applied to the animals' home pen, no habituation is needed.

Bergenin Ameliorates MPTP-Induced Parkinson's Disease by Activating PI3K/Akt Signaling Pathway.

BACKGROUND: Parkinson's disease (PD) is the second most prevalent progressive neurodegenerative disease, second only to Alzheimer's disease, with motor disorders and cognitive impairment. Bergenin (Berg), extracted from the herb of Saxifrage stolonifera Curt. (Hu-Er-Cao), has anti-tumor, anti-inflammation, anti-oxidative stress, and neuroprotective properties. OBJECTIVE: In this study, we wanted to investigate the effects of Berg on PD and the underlying mechanisms. METHODS: 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) was used to introduce PD symptoms in mice. The expression levels of tyrosine hydroxylase, dopamine, and Iba-1 were examined. The levels of a series of inflammatory mediators were measured by qPCR. In addition, the PI3K/Akt signaling pathway was investigated to illustrate the underlying mechanism. In vitro, PC12 cells subjected to lipopolysaccharide (LPS) were treated with Berg. RESULTS: We found that MPTP injection introduced motor deficits, apoptosis of neurons and inflammation, as well as inhibited the PI3K/Akt signaling pathway. However, Berg treatment suppressed the MPTP-induced alterations. In vitro, Berg attenuated the cytotoxic effects on PC12 cells induced by the culture supernatants derived from LPS-induced microglial cells. CONCLUSION: Berg attenuated the PD symptoms via activating the PI3K/Akt signaling pathway in vivo and in vitro.

Luteolin-7-O-glucoside protects dopaminergic neurons by activating estrogen-receptor-mediated signaling pathway in MPTP-induced mice.

BACKGROUND: Parkinson's disease (PD) is a neurodegenerative disorder that is characterized by the degeneration of dopaminergic neurons in substantia nigra (SN). Accumulating evidences implicate the beneficial role of estrogen in the therapy of PD. METHODS: In the present study, the protective function of luteolin-7-O-glucoside (LUT-7G), a natural flavonoid, was investigated in 1-methyl-4-phenylpyridinium (MPP+) treated SH-SY5Y cells and 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) induced mice. RESULTS: Pre-treatment of LUT-7G increased the viability and reduced the apoptosis of SH-SY5Y cells treated by MPP+. At molecular level, the Bcl-2/Bax ratio was increased, while the expression of cleaved caspase 3 was markedly lessened. Moreover, LUT-7G increased the expression of estrogen receptor (ER), ERα and ERß, and enhanced the activation of ERK1/2/STAT3/c-Fos that could be abolished by ER antagonists. Furthermore, in vivo experiment indicated that pre-treatment of LUT-7G improved the bradykinesia, and enhanced the muscle strength as well as the balancing capacity of mice treated with MPTP. And LUT-7G prevented the injury of TH positive cells in substantia nigra and increased TH positive nerve fibers in striatum. In addition, pre-treatment of LUT-7G also significantly diminished the MPTP-induced gliosis in substantia nigra. CONCLUSIONS: LUT-7G effectively protected dopaminergic neurons against MPP+ or MPTP-induced toxicity, probably by activating the ER-mediated signaling pathway. Our findings explore the therapeutic potential of LUT-7G for PD therapy.

Cordycepin mitigates MPTP-induced Parkinson's disease through inhibiting TLR/NF-κB signaling pathway.

Parkinson's disease (PD) is a neurodegenerative disease with movement disorder. PD is characterized by the loss of dopaminergic (DA) neurons in the substantia nigra. Cordycepin, a small molecule extracted from cordyceps sinensis, has neuroprotective, anti-inflammatory, antioxidant and anti-tumor properties. In this study, we explored its possible beneficial effects on PD. PD rat models and cell models were established via 1­methyl­4­phenyl­1,2,3,6­tetrahydropyridine (MPTP) injection and LPS treatment respectively, and cordycepin was administered. The motor functions of rats were examined, and the tyrosine hydroxylase (TH)-positive DA neurons and Iba1-positive microglia were detected by immunohistochemical and immunofluorescence staining. The expression levels of inflammatory and oxidative stress-related factors were also measured in vivo and in vitro. In addition, the TLR/NF-κB pathway was investigated to explore the mechanism. We found that in vivo, MPTP injection introduced motor disorders, the loss of DA neurons and the activation of TLR/NF-κB signaling pathway. Cordycepin treatment alleviated these MPTP-induced changes. In vitro, the results were confirmed in Lipopolysaccharide (LPS)-induced cells. Moreover, cordycepin mitigated the cytotoxic effects on PC12 cells produced by microglia. In conclusion, cordycepin alleviated PD symptoms by inhibiting TLR/NF-κB signaling pathway in vivo and in vitro.

Dorsal hypothalamic dopaminergic neurons play an inhibitory role in the hypothalamic-pituitary-adrenal axis via activation of D2R in mice.

AIM: The present study investigated the effects of dorsal hypothalamic dopamine (dh-DA) neurons on activation of hypothalamic-pituitary-adrenal (HPA) axis in adult male mice. METHODS: Tyrosine hydroxylase-labelled DA neurons, DA content, c-Fos immune-positive (c-Fos+) cells and CRH expression in paraventricular nuclei (PVN), serum CORT and ACTH were examined at 4-, 8-, and 12-hour after a signal injection of MPTP (20 mg kg-1 ) respectively. RESULTS: The dh-DA neurons and DA content in PVN at 4-hour post-MPTP were reduced with recovery at 12-hour post-MPTP, while decline of nigrostriatal DA neurons and DA content in striatum started from 12-hour post-MPTP. Number of c-Fos+ cells, and CORT/ACTH levels increased at 4-hour post-MPTP, followed by recovery at 12-hour post-MPTP. The CRH mRNA was elevated at 4-hour post-MPTP, and sustained for over 12 hours. At 2-hour post-MPTP, PVN-injection of D2R agonist quinpirole corrected the increases in c-Fos+ cells, CORT/ACTH and CRH mRNA, but D1R agonist SKF38393 did not. PVN-injection of D2R antagonist L-sulpiride alone caused increases in c-Fos+ cells, CORT/ACTH and CRH mRNA. Similarly, PVN-injection of CB1R agonist WIN552,12 prevented the increases in c-Fos+ cells and CORT/ACTH rather than CRH mRNA, which were blocked by CB1R antagonist AM251. Levels of PKA and CREB phosphorylation in PVN were increased at 4-hour post-MPTP, which were blocked by quinpirole, but not WIN552,12. PKA inhibitor H89 corrected the increase of CRH mRNA at 8-hour post-MPTP. CONCLUSION: The activation of dh-DA neurons regulates negatively HPA axis through targeting D2Rs of CRH neurons to enhance endocannabinoid release and inhibit PKA-CREB pathway.

Calycosin attenuates MPTP-induced Parkinson's disease by suppressing the activation of TLR/NF-κB and MAPK pathways.

Parkinson is the second common neurodegenerative disease. The characteristics of Parkinson's disease (PD) are the dopamin neurons loss caused by neuroinflammation responses. C alycosin, an isoflavone phytoestrogen isolated from Astragalus membranaceus, has multiple pharmacological activities, such as anti-inflammation, anti-tumor, and neuroprotective effects. However, it is unknown whether calycosin can mitigate PD symptoms. This study aims to explore whether calycosin can alleviate PD symptoms and the underlying mechanisms. PD was induced in mice by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) injection, and calycosin was given intracerebroventricularly to these mice. A cell model of nerve inflammation was established by BV2 microglia cells injected with lipopolysaccharide (LPS). The motor states were evaluated by stepping, whisker, and cylinder experiments. The states of dopaminergic neurons and microglia were detected by immunostainning of tyrosine hydroxylase and cluster of differentiation molecule 11b (CD11b). The expression levels of inflammatory factors were detected by qPCR. Toll-like receptor (TLR)/nuclear factor kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) pathways were investigated by western blot. We found that calycosin treatment mitigated the behavioral dysfunctions and inflammatory responses in MPTP-induced PD mice. The TLR/NF-κB and MAPK pathways in MPTP-induced PD mice were inhibited by calycosin treatment, which was coincident with experiments in LPS-induced BV2 cells. Above all, calycosin mitigates PD symptoms through TLR/NF-κB and MAPK pathways in mice and cell lines.

Relationship Between the Anxious Symptoms and the Neurotransmitter in Parkinson's Mice with Different Dosages of MPTP

ABSTRACT This study aims to explore the relationship between the anxious symptoms and the impairment of 5-hydroxytryptamine system in PD mice induced by different dosages of MPTP. The mice from the three model groups, the low-dose, dose and high-dose group, took longer time in the dark box than those in the control group (P<0.05). However, no statistically significant differences were found among the model groups. The number of open arm entry (OE) and the open arm time (OT) were significant lower in the model group than those in the control group in the elevated plus-maze test (P＜0.05). The percentage of OE in modle group was significantly lower compared with the control group (P＜0.05). The concentrations of striatum DA, HVA, 5-HT, and 5-HIAA were significantly reduced in the three model groups compared to the control group (P＜0.05). The 5-HT concentrations of high-dose group was significantly lower than those of the control group in the prefrontal cortex (P＜0.05). Anxiety symptoms were appeared in the three model groups of early PD mice, but no difference existed among these groups. The 5-hydroxytryptamine system was damaged after MPTP injection, which could lead to anxiety. However, the impairment of 5-hydroxytryptamine system induced by MPTP was dose-independent.