title: Long-term controlled GDNF over-expression reduces dopamine transporter activity without affecting tyrosine hydroxylase expression in the rat mesostriatal system.
abstract: The dopamine (DA) transporter (DAT) is a plasma membrane glycoprotein expressed in dopaminergic (DA-) cells that takes back DA into presynaptic neurons after its release. DAT dysfunction has been involved in different neuro-psychiatric disorders including Parkinson's disease (PD). On the other hand, numerous studies support that the glial cell line-derived neurotrophic factor (GDNF) has a protective effect on DA-cells. However, studies in rodents show that prolonged GDNF over-expression may cause a tyrosine hydroxylase (TH, the limiting enzyme in DA synthesis) decline. The evidence of TH down-regulation suggests that another player in DA handling, DAT, may also be regulated by prolonged GDNF over-expression, and the possibility that this effect is induced at GDNF expression levels lower than those inducing TH down-regulation. This issue was investigated here using intrastriatal injections of a tetracycline-inducible adeno-associated viral vector expressing human GDNF cDNA (AAV-tetON-GDNF) in rats, and doxycycline (DOX; 0.01, 0.03, 0.5 and 3mg/ml) in the drinking water during 5weeks. We found that 3mg/ml DOX promotes an increase in striatal GDNF expression of 12× basal GDNF levels and both DA uptake decrease and TH down-regulation in its native and Ser40 phosphorylated forms. However, 0.5mg/ml DOX promotes a GDNF expression increase of 3× basal GDNF levels with DA uptake decrease but not TH down-regulation. The use of western-blot under non-reducing conditions, co-immunoprecipitation and in situ proximity ligation assay revealed that the DA uptake decrease is associated with the formation of DAT dimers and an increase in DAT-α-synuclein interactions, without changes in total DAT levels or its compartmental distribution. In conclusion, at appropriate GDNF transduction levels, DA uptake is regulated through DAT protein-protein interactions without interfering with DA synthesis.
authors.affiliation: Departamento de Ciencias Médicas Básicas (Anatomía), Facultad de Ciencias de la Salud (Medicina), Instituto de Tecnologías Biomédicas (ITB, CIBICAN), Universidad de La Laguna, Tenerife, Spain.
authors.forename: Pedro
authors.initials: P
authors.lastname: Barroso-Chinea
authors.affiliation: Departamento de Ciencias Médicas Básicas (Anatomía), Facultad de Ciencias de la Salud (Medicina), Instituto de Tecnologías Biomédicas (ITB, CIBICAN), Universidad de La Laguna, Tenerife, Spain.
authors.forename: Ignacio
authors.initials: I
authors.lastname: Cruz-Muros
authors.affiliation: Departamento de Ciencias Médicas Básicas (Anatomía), Facultad de Ciencias de la Salud (Medicina), Instituto de Tecnologías Biomédicas (ITB, CIBICAN), Universidad de La Laguna, Tenerife, Spain.
authors.forename: Domingo
authors.initials: D
authors.lastname: Afonso-Oramas
authors.affiliation: Departamento de Ciencias Médicas Básicas (Anatomía), Facultad de Ciencias de la Salud (Medicina), Instituto de Tecnologías Biomédicas (ITB, CIBICAN), Universidad de La Laguna, Tenerife, Spain.
authors.forename: Javier
authors.initials: J
authors.lastname: Castro-Hernández
authors.affiliation: Departamento de Ciencias Médicas Básicas (Anatomía), Facultad de Ciencias de la Salud (Medicina), Instituto de Tecnologías Biomédicas (ITB, CIBICAN), Universidad de La Laguna, Tenerife, Spain.
authors.forename: Josmar
authors.initials: J
authors.lastname: Salas-Hernández
authors.affiliation: Laboratory of Experimental Neurosurgery and Multidisciplinary Research Institute (I.R.I.B.H.M.), Free University of Brussels (ULB), Brussels, Belgium.
authors.forename: Abdelwahed
authors.initials: A
authors.lastname: Chtarto
authors.affiliation: Departamento de Ciencias Médicas Básicas (Anatomía), Facultad de Ciencias de la Salud (Medicina), Instituto de Tecnologías Biomédicas (ITB, CIBICAN), Universidad de La Laguna, Tenerife, Spain.
authors.forename: Diego
authors.initials: D
authors.lastname: Luis-Ravelo
authors.affiliation: Laboratory of Cellular and Molecular Neurotherapies, Neuroscience Research Center, Department of Clinical Neuroscience, Lausanne University Hospital, Switzerland.
authors.forename: Marie
authors.initials: M
authors.lastname: Humbert-Claude
authors.affiliation: Laboratory of Cellular and Molecular Neurotherapies, Neuroscience Research Center, Department of Clinical Neuroscience, Lausanne University Hospital, Switzerland.
authors.forename: Liliane
authors.initials: L
authors.lastname: Tenenbaum
authors.affiliation: Departamento de Ciencias Médicas Básicas (Anatomía), Facultad de Ciencias de la Salud (Medicina), Instituto de Tecnologías Biomédicas (ITB, CIBICAN), Universidad de La Laguna, Tenerife, Spain; Centro de investigación Biomédica en Red sobre enfermedades neurodegenerativas, CIBERNED, Instituto de Salud Carlos III, Spain. Electronic address: tgonhern@ull.es.
authors.forename: Tomás
authors.initials: T
authors.lastname: González-Hernández
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D003342:Corpus Striatum / Q000166:cytology / Q000378:metabolism*; D004298:Dopamine / Q000378:metabolism; D050483:Dopamine Plasma Membrane Transport Proteins / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D051100:Glial Cell Line-Derived Neurotrophic Factor / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D008026:Ligation; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D014161:Transduction, Genetic; D014316:Tritium / Q000378:metabolism; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism*; D051844:alpha-Synuclein / Q000378:metabolism
chemical_list: D050483:Dopamine Plasma Membrane Transport Proteins; D051100:Glial Cell Line-Derived Neurotrophic Factor; D051844:alpha-Synuclein; C485184:enhanced green fluorescent protein; D014316:Tritium; D049452:Green Fluorescent Proteins; D014446:Tyrosine 3-Monooxygenase; D004298:Dopamine
keywords: Dopamine transporter; GDNF; Inducible adeno-associated viral vectors; Parkinson's disease; Tyrosine hydroxylase