Survival Of Dopamine Neurons Research Articles

Bright light exposure reduces TH-positive dopamine neurons: implications of light pollution in Parkinson's disease epidemiology

This study explores the effect of continuous exposure to bright light on neuromelanin formation and dopamine neuron survival in the substantia nigra. Twenty-one days after birth, Sprague–Dawley albino rats were divided into groups and raised under different conditions of light exposure. At the end of the irradiation period, rats were sacrificed and assayed for neuromelanin formation and number of tyrosine hydroxylase (TH)-positive neurons in the substantia nigra. The rats exposed to bright light for 20 days or 90 days showed a relatively greater number of neuromelanin-positive neurons. Surprisingly, TH-positive neurons decreased progressively in the substantia nigra reaching a significant 29% reduction after 90 days of continuous bright light exposure. This decrease was paralleled by a diminution of dopamine and its metabolite in the striatum. Remarkably, in preliminary analysis that accounted for population density, the age and race adjusted Parkinson's disease prevalence significantly correlated with average satellite-observed sky light pollution.

Glial cell-line derived neurotrophic factor (GDNF) replacement attenuates motor impairments and nigrostriatal dopamine deficits in 12-month-old mice with a partial deletion of GDNF

Glial cell-line derived neurotrophic factor (GDNF) has been established as a growth factor for the survival and maintenance of dopamine (DA) neurons. In phase I clinical trials, GDNF treatment in Parkinson's disease patients led to improved motor function and GDNF has been found to be down regulated in Parkinson's disease patients. Studies using GDNF heterozygous (Gdnf+/−) mice have demonstrated that a partial reduction of GDNF leads to an age-related accelerated decline in nigrostriatal DA system- and motor-function and increased neuro-inflammation and oxidative stress in the substantia nigra (SN). Therefore, the purpose of the current studies was to determine if GDNF replacement restores motor function and functional markers within the nigrostriatal DA system in middle-aged Gdnf+/− mice. At 11months of age, male Gdnf+/− and wildtype (WT) mice underwent bilateral intra-striatal injections of GDNF (10μg) or vehicle. Locomotor activity was assessed weekly 1–4weeks after treatment. Four weeks after treatment, their brains were processed for analysis of GDNF levels and various DAergic and oxidative stress markers. An intrastriatal injection of GDNF increased motor activity in Gdnf+/− mice to levels comparable to WT mice (1week after injection) and this effect was maintained through the 4-week time point. This increase in locomotion was accompanied by a 40% increase in striatal GDNF protein levels and SN GDNF expression in Gdnf+/− mice. Additionally, GDNF treatment significantly increased the number of tyrosine hydroxylase (TH)-positive neurons in the SN of middle-aged Gdnf+/− mice, but not WT mice, which was coupled with reduced oxidative stress in the SN. These studies further support that long-term changes related to the dysfunction of the nigrostriatal pathway are influenced by GDNF expression and add that this dysfunction appears to be responsive to GDNF treatment. Additionally, these studies suggest that long-term GDNF depletion alters the biological and behavioral responses to GDNF treatment.

Glutamate corelease promotes growth and survival of midbrain dopamine neurons.

Recent studies have proposed that glutamate corelease by mesostriatal dopamine (DA) neurons regulates behavioral activation by psychostimulants. How and when glutamate release by DA neurons might play this role remains unclear. Considering evidence for early expression of the type 2 vesicular glutamate transporter in mesencephalic DA neurons, we hypothesized that this cophenotype is particularly important during development. Using a conditional gene knock-out approach to selectively disrupt the Vglut2 gene in mouse DA neurons, we obtained in vitro and in vivo evidence for reduced growth and survival of mesencephalic DA neurons, associated with a decrease in the density of DA innervation in the nucleus accumbens, reduced activity-dependent DA release, and impaired motor behavior. These findings provide strong evidence for a functional role of the glutamatergic cophenotype in the development of mesencephalic DA neurons, opening new perspectives into the pathophysiology of neurodegenerative disorders involving the mesostriatal DA system.

Gdnf Activates Midline Repulsion by Semaphorin3B via NCAM during Commissural Axon Guidance

The Neurotrophic factor gdnf plays diverse developmental roles, supporting survival and also acting asachemoattractant for axon and cell migration. We report that in the developing spinal cord, a focal source of gdnf is present in the floor plate (FP) where commissural axons cross the midline. Gdnf has nodirect guidance properties but switches on the responsiveness of crossing commissural growth cones to the midline repellent Semaphorin3B by suppressing calpain-mediated processing of the Sema3B signaling coreceptor Plexin-A1. Analysis of single and double mutant mouse models indicates that although gdnf is the principal trigger of Sema3B midline repulsion, it acts with another FP cue, NrCAM. Finally, genetic and invitro experiments provide evidence that this gdnf effect is RET independent and mediated by NCAM/GFRα1 signaling. This study identifies a regulator of midline crossing and reveals interplays between Semaphorin and gdnf signaling during axon guidance.

Mitofusin 2 is necessary for striatal axonal projections of midbrain dopamine neurons

Mitochondrial dysfunction is implicated in aging and degenerative disorders such as Parkinson's disease (PD). Continuous fission and fusion of mitochondria shapes their morphology and is essential to maintain oxidative phosphorylation. Loss-of-function mutations in PTEN-induced kinase1 (PINK1) or Parkin cause a recessive form of PD and have been linked to altered regulation of mitochondrial dynamics. More specifically, the E3 ubiquitin ligase Parkin has been shown to directly regulate the levels of mitofusin 1 (Mfn1) and Mfn2, two homologous outer membrane large GTPases that govern mitochondrial fusion, but it is not known whether this is of relevance for disease pathophysiology. Here, we address the importance of Mfn1 and Mfn2 in midbrain dopamine (DA) neurons in vivo by characterizing mice with DA neuron-specific knockout of Mfn1 or Mfn2. We find that Mfn1 is dispensable for DA neuron survival and motor function. In contrast, Mfn2 DA neuron-specific knockouts develop a fatal phenotype with reduced weight, locomotor disturbances and death by 7 weeks of age. Mfn2 knockout DA neurons have spherical and enlarged mitochondria with abnormal cristae and impaired respiratory chain function. Parkin does not translocate to these defective mitochondria. Surprisingly, Mfn2 DA neuron-specific knockout mice have normal numbers of midbrain DA neurons, whereas there is a severe loss of DA nerve terminals in the striatum, accompanied by depletion of striatal DA levels. These results show that Mfn2, but not Mfn1, is required for axonal projections of DA neurons in vivo.

Genetic analysis of NR4A2 gene in a large population of Han Chinese patients with Parkinson's disease

NR4A2 gene is a transcription factor crucial for differentiation and survival of midbrain dopamine (DA) neurons, and several variants have been eported to be associated with Parkinson's disease (PD) in the Caucasian population. To determine whether there is any association of NR4A2 mutation or variation with PD in the Han Chinese population, we performed a genetic analysis of all the exons and exon-intron boundaries in 689 PD patients and 672 controls from mainland China using direct sequencing analysis. We identified four novel variants and two previously reported variants. Two novel variants (exon 2 c.-20 C>G and exon 3 c.711 C>A) were only found in PD. The others (exon 2 c.-35 A>G; exon 8 c.1615 G>A; intron 4 IVS4-16 G>T; and intron 6 IVS6+18 insG) were found in both PD and controls at different frequencies. Collectively, our results suggest that NR4A2 may be a susceptibility gene for PD in the Chinese population.

Glucose Regulated Protein 78 Diminishes α-Synuclein Neurotoxicity in a Rat Model of Parkinson Disease

Accumulation of human wild-type (wt) α-synuclein (α-syn) induces neurodegeneration in humans and in experimental rodent models of Parkinson disease (PD). It also leads to endoplasmic reticulum (ER) stress and activation of the unfolded protein response (UPR). We overexpressed glucose regulated protein 78, also known as BiP (GRP78/BiP), to test the hypothesis that this ER chaperone modulates the UPR, blocks apoptosis, and promotes the survival of nigral dopamine (DA) neurons in a rat model of PD induced by elevated level of human α-syn. We determined that α-syn activates ER stress mediators associated with pancreatic ER kinase-like ER kinase (PERK) and activating transcription factor-6 (ATF6) signaling pathways as well as proaoptotic CCAAT/-enhancer-binding protein homologous protein (CHOP) in nigral DA neurons. At the same time, overexpression of GRP78/BiP diminished α-syn neurotoxicity by down regulating ER stress mediators and the level of apoptosis, promoted survival of nigral tyrosine hydroxylase (TH) positive cells and resulted in higher levels of striatal DA, while eliminating amphetamine induced behavioral asymmetry. We also detected a complex between GRP78/BiP and α-syn that may contribute to prevention of the neurotoxicity caused by α-syn. Our data suggest that the molecular chaperone GRP78/BiP plays a neuroprotective role in α-syn-induced Parkinson-like neurodegeneration.

CDNF Protects the Nigrostriatal Dopamine System and Promotes Recovery after MPTP Treatment in Mice

Cerebral dopamine neurotrophic factor (CDNF) is a recently discovered protein, which belongs to the evolutionarily conserved CDNF/MANF family of neurotrophic factors. The degeneration of dopamine neurons following 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treatment is well characterized, and efficacy in this model is considered a standard criterion for development of parkinsonian therapies. MPTP is a neurotoxin, which produces parkinsonian symptoms in humans and in C57/Bl6 mice. To date, there are no reports about the effects of CDNF on dopamine neuron survival or function in the MPTP rodent model, a critical gap. Therefore, we studied whether CDNF has neuroprotective and neurorestorative properties for the nigrostriatal dopamine system after MPTP injections in C57/Bl6 mice. We found that bilateral striatal CDNF injections, given 20 h before MPTP, improved horizontal and vertical motor behavior. CDNF pretreatment increased tyrosine hydroxylase (TH) immunoreactivity in the striatum and in the substantia nigra pars reticulata (SNpr), as well as the number of TH-positive cells in substantia nigra pars compacta (SNpc). Posttreatment with CDNF, given 1 week after MPTP injections, increased horizontal and vertical motor behavior of mice, as well as dopamine fiber densities in the striatum and the number of TH-positive cells in SNpc. CDNF did not alter any of the analyzed dopaminergic biomarkers or locomotor behavior in MPTP-untreated animals. We conclude that striatal CDNF administration is both neuroprotective and neurorestorative for the TH-positive cells in the nigrostriatal dopamine system in the MPTP model, which supports the development of CDNF-based treatment for Parkinson's disease.

Signaling pathways mediating the neuroprotective effects of sex steroids and SERMs in Parkinson’s disease

Studies with the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) animal model of Parkinson's disease have shown the ability of 17β-estradiol to protect the nigrostriatal dopaminergic system. This paper reviews the signaling pathways mediating the neuroprotective effect of 17β-estradiol against MPTP-induced toxicity. The mechanisms of 17β-estradiol action implicate activation of signaling pathways such as the phosphatidylinositol-3 kinase/Akt and the mitogen-activated protein kinase pathways. 17β-estradiol signaling is complex and integrates multiple interactions with signaling molecules that act to potentiate a protective effect. 17β-estradiol signaling is mediated via estrogen receptors, including GPER1, but others receptors, such as the IGF-1 receptor, are implicated in the neuroprotective effect. Glial and neuronal crosstalk is a critical factor in the maintenance of dopamine neuronal survival and in the neuroprotective action of 17β-estradiol. Compounds that stimulate GPER1 such as selective estrogen receptor modulators and phytoestrogens show neuroprotective activity and are alternatives to 17β-estradiol.

Neuroprotective effects of riluzole in early phase Parkinson's disease on clinically relevant parameters in the marmoset MPTP model

The present study evaluates neuroprotection in a marmoset MPTP (1-methyl-1,2,3,6-tetrahydropyridine) model representing early Parkinson's disease (PD). The anti-glutamatergic compound riluzole is used as a model compound for neuroprotection. The compound is one of the few protective compounds used in the clinic for a neurodegenerative disorder.Marmoset monkeys were randomized into three groups of six: 1) an MPTP group receiving a total MPTP dose of 7 mg/kg (4 injections over two weeks, s.c.) 2) a riluzole group receiving besides MPTP, a twice daily dose of riluzole (10 mg/kg, p.o.), starting one week before MPTP and continuing for one week after the final MPTP injection and 3) a control group receiving saline instead of MPTP and riluzole. The marmosets' Parkinsonian symptoms were scored daily and their activity level, hand-eye coordination, jumping behavior, axial turning and night sleep parameters were tested and recorded weekly. At three weeks following the last MPTP challenge, brains were dissected and dopamine levels in the striatum and the tyrosine hydroxylase (TH) expressing dopamine (DA) neurons in the substantia nigra (SN) were compared. MPTP affected all behavioral parameters and sleep architecture and induced a relatively mild (50%) decline of DA neurons in the substantia nigra (SN). Riluzole relieved the Parkinsonian signs, and improved the hand-eye coordination as well as turning ability. Moreover, riluzole prevented the impact of MPTP on sleep architecture and rapid eye movement behavioral disorder (RBD). Riluzole also increased the number of surviving DA neurons in MPTP-treated marmosets to 75%. However, riluzole did not prevent the MPTP-induced impairments on locomotor activity and jumping activity.In conclusion, reduction of excitotoxicity by riluzole appeared to be effective in reducing progressive neurodegeneration and relieved several clinically relevant PD symptoms in an animal model representing the early phase of PD.

Dopamine neurons derived from human ES cells efficiently engraft in animal models of Parkinson’s disease

SUMMARYHuman pluripotent stem cells (hPSCs) are a promising source of cells for applications in regenerative medicine. Directed differentiation of hPSCs into specialized cells such as spinal motoneurons1 or midbrain dopamine (DA) neurons2 has been achieved. However, the effective use of hPSCs for cell therapy has lagged behind. While mouse PSC-derived DA neurons have shown efficacy in models of Parkinson’s disease (PD)3, 4, DA neurons from human PSCs generally display poor in vivo performance5. There are also considerable safety concerns for hPSCs related to their potential for teratoma formation or neural overgrowth6, 7Here we present a novel floor plate-based strategy for the derivation of human DA neurons that efficiently engraft in vivo, suggesting that past failures were due to incomplete specification rather than a specific vulnerability of the cells. Midbrain floor plate precursors are derived from hPSCs in 11 days following exposure to small molecule activators of sonic hedgehog (SHH) and canonical WNT signaling. Engraftable midbrain DA neurons are obtained by day 25 and can be maintained in vitro for several months. Extensive molecular profiling, biochemical and electrophysiological data define developmental progression and confirm identity of hPSC-derived midbrain DA neurons. In vivo survival and function is demonstrated in PD models using three host species. Long-term engraftment in 6-OHDA-lesioned mice and rats demonstrates robust survival of midbrain DA neurons, complete restoration of amphetamine-induced rotation behavior and improvements in tests of forelimb use and akinesia. Finally, scalability is demonstrated by transplantation into Parkinsonian monkeys. Excellent DA neuron survival, function and lack of neural overgrowth in the three animal models indicate promise for the development of cell based therapies in PD.

Regulation of complex I by Engrailed is complex too

Engrailed, a homeobox transcription factor that is crucial for neuronal development, is now shown to regulate mitochondrial complex I and to be critical for the survival, protection and physiology of adult dopamine neurons.

Dual immunofluorescence study of citrullinated proteins in Parkinson diseased substantia nigra

Deimination is a post-translational modification of proteins in which selected arginine amino acids are enzymatically converted to citrullines. Using dual-color immunofluorescence and an established monoclonal antibody (F95) against peptidyl-citrulline moieties, the present study is the first to compare immunohistochemical staining patterns for deiminated proteins in human substantia nigra (SN) from patients with Parkinson disease (PD) versus similar control specimens supplied by the Harvard Brain Bank. In control SN sections, many tyrosine hydroxylase (TH)-immunoreactive dopamine neurons were seen surrounded either by small fibers immunoreactive for deiminated proteins, or large reactive astrocytes, co-localized with glial fibrillary acidic protein (GFAP). However, in SN specimens from PD patients, immunoreactivity for deiminated proteins was also demonstrated within the cytoplasm of many surviving dopamine neurons that were also immunoreactive for TH, but this staining was not specifically restricted to Lewy bodies. Although the identity of neuronal deiminated proteins in these SN dopamine neurons is unknown, the present study provides evidence that the anatomical expression of these proteins in PD is altered and thus suggests that deimination may be involved in the pathophysiology of this disease.

Loss of mitochondrial complex I activity potentiates dopamine neuron death induced by microtubule dysfunction in a Parkinson’s disease model

Mitochondrial complex I dysfunction is regarded as underlying dopamine neuron death in Parkinson's disease models. However, inactivation of the Ndufs4 gene, which compromises complex I activity, does not affect the survival of dopamine neurons in culture or in the substantia nigra pars compacta of 5-wk-old mice. Treatment with piericidin A, a complex I inhibitor, does not induce selective dopamine neuron death in either Ndufs4(+/+) or Ndufs4(-/-) mesencephalic cultures. In contrast, rotenone, another complex I inhibitor, causes selective toxicity to dopamine neurons, and Ndufs4 inactivation potentiates this toxicity. We identify microtubule depolymerization and the accumulation of cytosolic dopamine and reactive oxygen species as alternative mechanisms underlying rotenone-induced dopamine neuron death. Enhanced rotenone toxicity to dopamine neurons from Ndufs4 knockout mice may involve enhanced dopamine synthesis caused by the accumulation of nicotinamide adenine dinucleotide reduced. Our results suggest that the combination of disrupting microtubule dynamics and inhibiting complex I, either by mutations or exposure to toxicants, may be a risk factor for Parkinson's disease.

The transcription factor NURR1 exerts concentration-dependent effects on target genes mediating distinct biological processes

The transcription factor NURR1 plays a pivotal role in the development and maintenance of neurotransmitter phenotype in midbrain dopamine neurons. Conversely, decreased NURR1 expression is associated with a number of dopamine-related CNS disorders, including Parkinson’s disease and drug addiction. In order to better understand the nature of NURR1-responsive genes and their potential roles in dopamine neuron differentiation and survival, we used a human neural cellular background (SK-N-AS cells) in which to generate a number of stable clonal lines with graded NURR1 gene expression that approximated that seen in DA cell-rich human substantia nigra. Gene expression profiling data from these NURR1-expressing clonal lines were validated by quantitative RT-PCR and subjected to bioinformatic analyses. The present study identified a large number of NURR1-responsive genes and demonstrated the potential importance of concentration-dependent NURR1 effects in the differential regulation of distinct NURR1 target genes and biological pathways. These data support the promise of NURR1-based CNS therapeutics for the neuroprotection and/or functional restoration of DA neurons.

Effects of brain-derived neurotrophic factor on dopaminergic function and motor behavior during aging

Brain-derived neurotrophic factor (BDNF) is critical in synaptic plasticity and in the survival and function of midbrain dopamine neurons. In this study, we assessed the effects of a partial genetic deletion of BDNF on motor function and dopamine (DA) neurotransmitter measures by comparing Bdnf(+/-) with wildtype mice (WT) at different ages. Bdnf(+/-) and WT mice had similar body weights until 12 months of age; however, at 21 months, Bdnf(+/-) mice were significantly heavier than WT mice. Horizontal and vertical motor activity was reduced for Bdnf(+/-) compared to WT mice, but was not influenced by age. Performance on an accelerating rotarod declined with age for both genotypes and was exacerbated for Bdnf(+/-) mice. Body weight did not correlate with any of the three behavioral measures studied. Dopamine neurotransmitter markers indicated no genotypic difference in striatal tyrosine hydroxylase, DA transporter (DAT) or vesicular monoamine transporter 2 (VMAT2) immunoreactivity at any age. However, DA transport via DAT (starting at 12 months) and VMAT2 (starting at 3 months) as well as KCl-stimulated DA release were reduced in Bdnf(+/-) mice and declined with age suggesting an increasingly important role for BDNF in the release and uptake of DA with the aging process. These findings suggest that a BDNF expression deficit becomes more critical to dopaminergic dynamics and related behavioral activities with increasing age.

Preliminary evidence that polymorphisms in dopamine-related transcription factors LMX1A, LMX1B and PITX3 are associated with schizophrenia

The early development of dopaminergic pathways has been attributed importance for the aetiology of schizophrenia. Several transcription factors are involved in the survival and maturation of dopamine neurons, including LMX1A, LMX1B and PITX3. The possibility that polymorphisms in these genes may influence the development and/or the maintenance of dopaminergic neurons prompted us to investigate if five single nucleotide polymorphisms (SNPs) previously linked to Parkinson's disease are associated with this disorder. Preliminary evidence that genetic variation in LMX1A (rs6668493, rs4657411), LMX1B (rs10987386) and PITX3 (rs4919621) may increase the risk of developing schizophrenia is presented.

DJ‐1, PINK1, and their effects on mitochondrial pathways

Genetic forms of parkinsonism are interesting for two particular reasons. First, finding a gene identifies a cause for a disease that would otherwise be unexplained. Second, finding several genes for the same disorder allows us to reconstruct molecular pathways that, in the example of Parkinson's disease, are be associated with the survival of dopamine neurons in the substantia nigra. Two rare causes of parkinsonism, DJ-1 and PINK1, are associated with mitochondria. This organelle has long been linked with Parkinson's disease, and recent results are starting to show how mutations impact mitochondrial function. In this short review, I will discuss how we can use some of this information to understand why it is that neurons become dysfunctional in PD.

Unexpected Lack of Hypersensitivity in LRRK2 Knock-Out Mice to MPTP (1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine)

Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are the most common known cause of Parkinson's disease (PD). Whether loss of LRRK2 function accounts for neurodegeneration of dopamine neurons in PD is not known, nor is it known whether LRRK2 kinase activity modulates the susceptibility of dopamine (DA) neurons to the selective dopaminergic toxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). To better understand the role of LRRK2 in DA neuronal survival and its role in the susceptibility of DA neurons to MPTP, we generated LRRK2 knock-out (KO) mice lacking the kinase domain of LRRK2. Here, we show that LRRK2 KO mice are viable and have no major abnormalities and live to adulthood. The dopaminergic system is normal in LRRK2 KO mice as assessed via HPLC for DA and its metabolites and via stereologic assessment of DA neuron number in young and aged mice. Importantly, there is no significant difference in the susceptibility of LRRK2 KO and wild-type mice to MPTP. These results suggest that LRRK2 plays little if any role in the development and survival of DA neurons under physiologic conditions. Thus, PD due to LRRK2 mutations are likely not due to a loss of function. Moreover, LRRK2 is not required for the susceptibility of DA neurons to MPTP.

A Sporadic Parkinson Disease Model via Silencing of the Ubiquitin-Proteasome/E3 Ligase Component SKP1A

The aim of this study was to develop a new model of sporadic Parkinson disease (PD) based on silencing of the SKP1A gene, a component of the ubiquitin-proteasome/E3 ligase complex, Skp1, Cullin 1, F-box protein, which was found to be highly decreased in the substantia nigra of sporadic PD patients. Initially, an embryonic mouse substantia nigra-derived cell line (SN4741 cells) was infected with short hairpin RNA lentiviruses encoding the murine transcript of the SKP1A gene or with scrambled vector. SKP1A silencing resulted in increased susceptibility to neuronal damages induced by the parkinsonism-inducing neurotoxin 1-methyl-4-phenylpyridinium ion and serum starvation, in parallel with a decline in the expression of the dopaminergic markers, dopamine transporter and vesicular monoamine transporter-2. SKP1A-deficient cells presented a delay in completion of the cell cycle and the inability to arrest at the G(0)/G(1) phase when induced to differentiate. Instead, the cells progressed through S phase, developing rounded aggregates with characteristics of aggresomes including immunoreactivity for gamma-tubulin, alpha-synuclein, ubiquitin, tyrosine hydroxylase, Hsc-70 (70-kDa heat shock cognate protein), and proteasome subunit, and culminating in a lethal phenotype. Conversely, stably enforced expression of wild type SKP1A duplicated the survival index of naïve SN4741 cells under proteasomal inhibition injury, suggesting a new structural role of SKP1 in dopaminergic neuronal function, besides its E3 ligase activity. These results link, for the first time, SKP1 to dopamine neuronal function and survival, suggesting an essential role in sporadic PD. In summary, this new model has reproduced to a significant extent the molecular alterations described in sporadic PD at the cellular level, implicating Skp1 as a potential modifier in sporadic PD neurodegeneration.