scholarly journals Vglut2 expression in dopamine neurons contributes to post-lesional striatal reinnervation

2019 ◽  
Author(s):  
Willemieke M. Kouwenhoven ◽  
Guillaume Fortin ◽  
Anna-Maija Penttinen ◽  
Clélia Florence ◽  
Benoît Delignat-Lavaud ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Glutamate Transporter ◽  
Dorsal Striatum ◽  
Dopamine Neurons ◽  
Developmental Expression ◽  
Genetic Lineage ◽  
Axonal Arborization ◽  
Neurotoxic Lesion ◽  
6 Hydroxydopamine

ABSTRACTIn Parkinson’s disease, the most vulnerable neurons are found in the ventral tier of the substantia nigra (SN), while the adjacent dopamine (DA) neurons of the ventral tegmental area (VTA) are mostly spared. Although a significant subset of adult VTA DA neurons expresses Vglut2, a vesicular glutamate transporter, and release glutamate as a second neurotransmitter in the striatum, only very few adult SN DA neurons have this capacity. Previous work has demonstrated that lesions created by neurotoxins such as MPTP and 6-hydroxydopamine (6-OHDA) can upregulate the expression of Vglut2 in surviving DA neurons. Currently, the molecular mechanisms explaining the plasticity of Vglut2 expression in DA neurons are unknown, as are the physiological consequences for DA neuron function and survival. Here we aimed to characterize the developmental expression pattern of Vglut2 in DA neurons and the role of this transporter in post-lesional plasticity in these neurons. Using an intersectional genetic lineage-mapping approach, based on Vglut2-Cre and TH-Flpo drivers, we first found that more than 98% of DA neurons expressed Vglut2 at some point in their embryonic development. Expression of this transporter was detectable in most DA neurons until E11.5 and was found to be localized in developing axons. Moderate enhancement of VGLUT2 expression in primary DA neurons caused an increase in axonal arborization length. Compatible with a developmental role, constitutive deletion of Vglut2 caused a regional defect in TH-innervation of the dorsal striatum in E18.5 embryos. Moreover, using an in vitro neurotoxin model, we demonstrate that Vglut2 expression can be upregulated in post-lesional DA neurons by 2.5-fold, arguing that the developmental expression of Vglut2 in DA neurons can be reactivated at postnatal stages and contribute to post-lesional plasticity of dopaminergic axons. In support of this hypothesis, we find fewer mesostriatial dopaminergic projections in the striatum of conditional Vglut2 KO mice 7 weeks after a neurotoxic lesion, compared to control animals. Thus, we propose here that one of the functions of Vglut2 in adult DA neurons is to promote post-lesional recovery of meso-striatal axons.

scholarly journals Neuroprotective Potential of a Small Molecule RET Agonist in Cultured Dopamine Neurons and Hemiparkinsonian Rats

2021 ◽  
pp. 1-24
Author(s):  
Juho-Matti Renko ◽  
Arun Kumar Mahato ◽  
Tanel Visnapuu ◽  
Konsta Valkonen ◽  
Mati Karelson ◽  
...  
Keyword(s):  
Mapk Signaling ◽  
Dopamine Neurons ◽  
Dopamine Depletion ◽  
Wild Type ◽  
Disease Modifying Therapy ◽  
Immortalized Cells ◽  
Midbrain Dopamine ◽  
6 Hydroxydopamine

Background: Parkinson’s disease (PD) is a progressive neurological disorder where loss of dopamine neurons in the substantia nigra and dopamine depletion in the striatum cause characteristic motor symptoms. Currently, no treatment is able to halt the progression of PD. Glial cell line-derived neurotrophic factor (GDNF) rescues degenerating dopamine neurons both in vitro and in animal models of PD. When tested in PD patients, however, the outcomes from intracranial GDNF infusion paradigms have been inconclusive, mainly due to poor pharmacokinetic properties. Objective: We have developed drug-like small molecules, named BT compounds that activate signaling through GDNF’s receptor, the transmembrane receptor tyrosine kinase RET, both in vitro and in vivo and are able to penetrate through the blood-brain barrier. Here we evaluated the properties of BT44, a second generation RET agonist, in immortalized cells, dopamine neurons and rat 6-hydroxydopamine model of PD. Methods: We used biochemical, immunohistochemical and behavioral methods to evaluate the effects of BT44 on dopamine system in vitro and in vivo. Results: BT44 selectively activated RET and intracellular pro-survival AKT and MAPK signaling pathways in immortalized cells. In primary midbrain dopamine neurons cultured in serum-deprived conditions, BT44 promoted the survival of the neurons derived from wild-type, but not from RET knockout mice. BT44 also protected cultured wild-type dopamine neurons from MPP +-induced toxicity. In a rat 6-hydroxydopamine model of PD, BT44 reduced motor imbalance and could have protected dopaminergic fibers in the striatum. Conclusion: BT44 holds potential for further development into a novel, possibly disease-modifying therapy for PD.

scholarly journals Adeno-Associated Viral Delivery of GDNF Promotes Recovery of Dopaminergic Phenotype following a Unilateral 6-Hydroxydopamine Lesion

2002 ◽  
Vol 11 (3) ◽  
pp. 215-227 ◽  
Author(s):  
John Mcgrath ◽  
Elishia Lintz ◽  
Barry J. Hoffer ◽  
Greg A. Gerhardt ◽  
E. Matthew Quintero ◽  
...  
Keyword(s):  
Neurotrophic Factor ◽  
Viral Vectors ◽  
Medial Forebrain Bundle ◽  
Therapeutic Potential ◽  
Clinical Situation ◽  
Dopamine Neurons ◽  
Nigrostriatal System ◽  
Nigrostriatal Pathway ◽  
Neurotoxic Lesion ◽  
6 Hydroxydopamine

Glial cell line-derived neurotrophic factor (GDNF) is a potent neurotrophic factor for dopamine neurons that has been proposed for use in the treatment of Parkinson's disease (PD). Previous studies using viral vectors to deliver GDNF in rodent models of PD have entailed administering the virus either prior to or immediately after neurotoxin-induced lesions, when the nigrostriatal pathway is largely intact, a paradigm that does not accurately reflect the clinical situation encountered with Parkinson's patients. In this study, recombinant adeno-associated virus carrying the gene encoding GDNF (rAAV-GDNF) was administered to animals bearing a maximal lesion in the nigrostriatal system, more closely resembling fully developed PD. Rats were treated with 6-hydroxydopamine into the medial forebrain bundle and assessed by apomorphine-induced rotational behavior for 5 weeks prior to virus administration. Within 4 weeks of a single intrastriatal injection of rAAV-GDNF, unilaterally lesioned animals exhibited significant behavioral recovery, which correlated with increased expression of dopaminergic markers in the substantia nigra, the medial forebrain bundle, and the striatum. Our findings demonstrate that rAAV-GDNF is capable of rescuing adult dopaminergic neurons from near complete phenotypic loss following a neurotoxic lesion, effectively restoring a functional dopaminergic pathway and diminishing motoric deficits. These data provide further support for the therapeutic potential of rAAV-GDNF-based gene therapy in the treatment of PD.

Cordycepin Exerts Neuroprotective Effects via an Anti-Apoptotic Mechanism based on the Mitochondrial Pathway in a Rotenone-Induced Parkinsonism Rat Model

2019 ◽  
Vol 18 (8) ◽  
pp. 609-620 ◽  
Author(s):  
Xin Jiang ◽  
Pei-Chen Tang ◽  
Qin Chen ◽  
Xin Zhang ◽  
Yi-Yun Fan ◽  
...  
Keyword(s):  
Rat Model ◽  
Molecular Mechanisms ◽  
Caspase 3 ◽  
Therapeutic Potential ◽  
Neuroprotective Effect ◽  
Dopamine Neurons ◽  
Substantia Nigra Pars Compacta ◽  
Neuroprotective Effects ◽  
Cyt C

Background: Cordycepin (Cor), one of the major bioactive components of the traditional Chinese medicine Cordyceps militaris, has been used in clinical practice for several years. However, its neuroprotective effect remains unknown. Aim: The purpose of the study was to evaluate the neuroprotective effects of Cor using a rotenoneinduced Parkinson’s Disease (PD) rat model and to delineate the possible associated molecular mechanisms. Methods: In vivo, behavioural tests were performed based on the 10-point scale and grid tests. Levels of dopamine and its metabolites in the striatum and the numbers of TH-positive neurons in the Substantia Nigra pars compacta (SNpc) were investigated by high-performance liquid chromatography with electrochemical detection and immunohistochemical staining, respectively. In vitro, cell apoptosis rates and Mitochondrial Membrane Potential (MMP) were analysed by flow cytometry and the mRNA and protein levels of Bax, Bcl-2, Bcl-xL, Cytochrome c (Cyt-c), and caspase-3 were determined by quantitative real-time PCR and western blotting. Results: Showed that Cor significantly improved dyskinesia, increased the numbers of TH-positive neurons in the SNpc, and maintained levels of dopamine and its metabolites in the striatum in rotenone- induced PD rats. We also found that apoptosis was suppressed and the loss of MMP was reversed with Cor treatment. Furthermore, Cor markedly down-regulated the expression of Bax, upregulated Bcl-2 and Bcl-xL, inhibited the activation of caspase-3, and decreased the release of Cyt-c from the mitochondria to the cytoplasm, as compared to those in the rotenone-treated group. Conclusion: Therefore, Cor protected dopamine neurons against rotenone-induced apoptosis by improving mitochondrial dysfunction in a PD model, demonstrating its therapeutic potential for this disease.

scholarly journals Cellular Programming and Reprogramming: Sculpting Cell Fate for the Production of Dopamine Neurons for Cell Therapy

2012 ◽  
Vol 2012 ◽  
pp. 1-17 ◽  
Author(s):  
Julio C. Aguila ◽  
Eva Hedlund ◽  
Rosario Sanchez-Pernaute
Keyword(s):  
Stem Cells ◽  
Cell Fate ◽  
Pluripotent Stem Cells ◽  
Dorsal Striatum ◽  
Dopamine Neurons ◽  
Cell Fates ◽  
Intrinsic Factors ◽  
Neuronal Identity ◽  
The Right

Pluripotent stem cells are regarded as a promising cell source to obtain human dopamine neurons in sufficient amounts and purity for cell replacement therapy. Importantly, the success of clinical applications depends on our ability to steer pluripotent stem cells towards the right neuronal identity. In Parkinson disease, the loss of dopamine neurons is more pronounced in the ventrolateral population that projects to the sensorimotor striatum. Because synapses are highly specific, only neurons with this precise identity will contribute, upon transplantation, to the synaptic reconstruction of the dorsal striatum. Thus, understanding the developmental cell program of the mesostriatal dopamine neurons is critical for the identification of the extrinsic signals and cell-intrinsic factors that instruct and, ultimately, determine cell identity. Here, we review how extrinsic signals and transcription factors act together during development to shape midbrain cell fates. Further, we discuss how these same factors can be appliedin vitroto induce, select, and reprogram cells to the mesostriatal dopamine fate.

scholarly journals GDNF receptor agonist supports dopamine neurons in vitro and protects their function in animal model of Parkinson’s

2019 ◽  
Author(s):  
Arun Kumar Mahato ◽  
Juho-Matti Renko ◽  
Jaakko Kopra ◽  
Tanel Visnapuu ◽  
Ilari Korhonen ◽  
...  
Keyword(s):  
Intracellular Signaling ◽  
Dopamine Neurons ◽  
Signaling Cascades ◽  
Pharmacokinetic Properties ◽  
Nigrostriatal Dopamine ◽  
6 Hydroxydopamine ◽  
Nigrostriatal Dopamine Neurons ◽  
Excellent Tool

AbstractMotor symptoms of Parkinson’s disease (PD) are caused by degeneration and progressive loss of nigrostriatal dopamine neurons. Currently no cure for this disease is available. Existing drugs alleviate PD symptoms, but fail to halt neurodegeneration. Glial cell line-derived neurotrophic factor (GDNF) is able to protect and repair dopamine neurons in vitro and in animal models of PD, but its clinical use is complicated by pharmacokinetic properties. In the present study we demonstrate the ability of a small molecule agonist of GDNF receptor RET to support the survival of cultured dopamine neurons only when they express GDNF receptors. In addition, BT13 activates intracellular signaling cascades in vivo, stimulates release of dopamine and protect the function of dopaminergic neurons in a 6-hydroxydopamine (6-OHDA) rat model of PD. In contrast to GDNF, BT13 is able to penetrate through the blood-brain-barrier. Thus, BT13 serves as an excellent tool compound for the development of novel disease-modifying treatments against PD.

scholarly journals Senescent Cell-Secreted Netrin-1 Modulates Aging-Related Disorders by Recruiting Sympathetic Fibers

2020 ◽  
Vol 12 ◽  
Author(s):  
Ai Qing Yu ◽  
Jie Wang ◽  
Xiao Jia Zhou ◽  
Ke Yu Chen ◽  
You De Cao ◽  
...  
Keyword(s):  
Cellular Senescence ◽  
Molecular Mechanisms ◽  
Human Colon ◽  
Senescent Cell ◽  
Aged Mouse ◽  
Age Related ◽  
Mouse Tissues ◽  
6 Hydroxydopamine

Cellular senescence is implicated in several lines of aging-related disorders. However, the potential molecular mechanisms by which cellular senescence modulates age-related pathologies remain largely unexplored. Herein, we report that the density of sympathetic fibers (SFs) is significantly elevated in naturally aged mouse tissues and human colon adenoma tissues compared to the SFs densities in the corresponding young mouse tissues and human non-lesion colon tissues. A dorsal root ganglion (DRG)-human diploid fibroblast coculture assay revealed that senescent cells promote the outgrowth of SFs, indicating that the senescent cells induce recruitment of SFs in vitro. Additionally, subcutaneous transplantation of 2BS fibroblasts in nude mice shows that transplanted senescent 2BS fibroblasts promote SFs infiltration. Intra-articular senolytic molecular injection can reduce SFs density and inhibit SFs infiltration caused by senescent cells in osteoarthritis (OA), suggesting senescent cells promote the infiltration of SFs in vivo in aged tissues. Notably, the elevated level of SFs contributes to impaired cognitive function in naturally aged mice, which can be reversed by treatment with propranolol hydrochloride, a non-selective β receptor blocker that inhibits sympathetic nerve activity (SNA) by blocking non-selective β receptors. Additionally, 6-hydroxydopamine (6-OHDA)-induced sympathectomy improved hepatic sympathetic overactivity mediated hepatic steatosis in high fat diet (HFD)-fed APOE knockout mice (APOE−/− mice) by reducing hepatic SNA. Taken together, this study concludes that senescent cell-secreted netrin-1 mediated SFs outgrowth and infiltration, which contributes to aging-related disorders, suggesting that clearing senescent cells or inhibiting SNA is a promising therapeutic strategy for improving sympathetic nervous system (SNS) hyperactivity-induced aging-related pathologies.

scholarly journals A Novel Family of Serine/Threonine Kinases Participating in Spermiogenesis

1997 ◽  
Vol 139 (7) ◽  
pp. 1851-1859 ◽  
Author(s):  
Peter Kueng ◽  
Zariana Nikolova ◽  
Valentin Djonov ◽  
Andrew Hemphill ◽  
Valeria Rohrbach ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Serine Phosphorylation ◽  
Developmental Expression ◽  
Seminiferous Tubules ◽  
Serine Kinase ◽  
Related Family ◽  
Maturation In Vitro ◽  
Threonine Kinases ◽  
Sexually Mature

The molecular mechanisms regulating the spectacular cytodifferentiation observed during spermiogenesis are poorly understood. We have recently identified a murine testis-specific serine kinase (tssk) 1, constituting a novel subfamily of serine/threonine kinases. Using low stringency screening we have isolated and molecularly characterized a second closely related family member, tssk 2, which is probably the orthologue of the human DGS-G gene. Expression of tssk 1 and tssk 2 was limited to the testis of sexually mature males. Immunohistochemical staining localized both kinases to the cytoplasm of late spermatids and to structures resembling residual bodies. tssk 1 and tssk 2 were absent in released sperms in the lumen of the seminiferous tubules and the epididymis, demonstrating a tight window of expression restricted to the last stages of spermatid maturation. In vitro kinase assays of immunoprecipitates containing either tssk 1 or tssk 2 revealed no autophosphorylation of the kinases, however, they led to serine phosphorylation of a coprecipitating protein of ∼65 kD. A search for interacting proteins using the yeast two-hybrid system with tssk 1 and tssk 2 cDNA as baits and a prey cDNA library from mouse testis, led to the isolation of a novel cDNA, interacting specifically with both tssk 1 and tssk 2, and encoding the coprecipitated 65-kD protein phosphorylated by both kinases. Interestingly, expression of the interacting clone was also testis specific and paralleled the developmental expression observed for the kinases themselves. These results represent the first demonstration of the involvement of a distinct kinase family, the tssk serine/threonine kinases, together with a substrate in the cytodifferentiation of late spermatids to sperms.

scholarly journals Ventral tegmental area astrocytes orchestrate avoidance and approach behavior

2018 ◽  
Author(s):  
J. A. Gomez ◽  
J. M. Perkins ◽  
G. M. Beaudoin ◽  
N. B. Cook ◽  
S. A. Quraishi ◽  
...  
Keyword(s):  
Ventral Tegmental Area ◽  
Glutamate Transporter ◽  
Dopamine Neurons ◽  
Inhibitory Neurons ◽  
Approach Behavior ◽  
Gaba Neurons ◽  
Avoidance Behaviors ◽  
Glutamatergic Signaling ◽  
Ventral Tegmental

AbstractThe ventral tegmental area (VTA) is a heterogeneous midbrain structure, containing neurons and astrocytes, that coordinates approach and avoidance behaviors by integrating activity from numerous afferents. Within neuron-astrocyte networks, astrocytes control signals from distinct afferents in a circuit-specific manner, but whether this capacity scales up to drive motivated behavior has been undetermined. Using genetic and optical dissection strategies in vitro and during behavior we report that VTA astrocytes tune glutamatergic signaling selectively on local inhibitory neurons to drive a functional circuit for learned avoidance. In this circuit, VTA astrocytes facilitate excitation of local GABA neurons to increase inhibition of dopamine neurons. The increased inhibition of dopamine neurons elicits real-time and learned avoidance behavior that is sufficient to impede expression of learned preference for reward. Despite the large number of functions performed by astrocytes, loss of one glutamate transporter (GLT-1) from VTA astrocytes selectively blocks these avoidance behaviors and spares preference for reward. Thus, VTA astrocytes selectively regulate excitation of local GABA neurons to drive a distinct learned avoidance circuit that opposes learned approach behavior.

High resolution mapping of nucleic acid containing complexes in vitro and in situ

1996 ◽  
Vol 54 ◽  
pp. 48-49
Author(s):  
D. P. Bazett-Jones ◽  
M. J. Hendzel
Keyword(s):  
Nucleic Acid ◽  
High Resolution ◽  
Transcription Initiation ◽  
Molecular Mechanisms ◽  
Heavy Atom ◽  
Regulation Of Transcription ◽  
High Exposure ◽  
Resolution Mapping ◽  
Tata Sequence

Structural analysis of combinations of nucleosomes and transcription factors on promoter and enhancer elements is necessary in order to understand the molecular mechanisms responsible for the regulation of transcription initiation. Such complexes are often not amenable to study by high resolution crystallographic techniques. We have been applying electron spectroscopic imaging (ESI) to specific problems in molecular biology related to transcription regulation. There are several advantages that this technique offers in studies of nucleoprotein complexes. First, an intermediate level of spatial resolution can be achieved because heavy atom contrast agents are not necessary. Second, mass and stoichiometric relationships of protein and nucleic acid can be estimated by phosphorus detection, an element in much higher proportions in nucleic acid than protein. Third, wrapping or bending of the DNA by the protein constituents can be observed by phosphorus mapping of the complexes. Even when ESI is used with high exposure of electrons to the specimen, important macromolecular information may be provided. For example, an image of the TATA binding protein (TBP) bound to DNA is shown in the Figure (top panel). It can be seen that the protein distorts the DNA away from itself and much of its mass sits off the DNA helix axis. Moreover, phosphorus and mass estimates demonstrate whether one or two TBP molecules interact with this particular promoter TATA sequence.

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