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Normal Nigrostriatal Innervation but Dopamine Dysfunction in Mice Carrying Hypomorphic Tyrosine Hydroxylase Alleles
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
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2003 (English)In: Journal of Neuroscience Research, ISSN 0360-4012, E-ISSN 1097-4547, Vol. 72, no 4, 444-453 p.Article in journal (Refereed) Published
Abstract [en]

We investigated the use of the mouse tyrosine hydroxylase (TH) gene to drive knock-in constructs in catecholaminergic neurons. Two targeting constructs representing truncated forms of either of the BMP receptors ALK-2 or BMPR-II preceded by an internal ribosome entry site (IRES) were introduced into the 3' untranslated region of TH. An frt-flanked neomycin-resistance (neo(r)) cassette was placed in the 3' end of the targeting constructs. Mice homozygous for the knock-in alleles showed various degrees of hypokinetic behavior, depending mainly on whether the neo(r) cassette was removed. In situ hybridization and immunohistochemistry showed that TH mRNA and protein were variously down-regulated in these mouse strains. Reduced levels of dopamine and noradrenalin were found in several brain areas. However, number and morphology of neurons in substantia nigra and their projections to striatum appeared normal in the neo(r)-positive TH hypomorphic mice as examined by markers for L-aromatic amino acid decarboxylase and the dopamine transporter. Elimination of the neo(r) cassette from the knock-in alleles partially restored TH and dopamine levels. The present neo(r)-positive TH hypomorphic mice show that nigrostriatal innervation develops independently of TH and should find use as a model for conditions of reduced catecholamine synthesis, as seen in, for example, L-dihydroxyphenylalanine-responsive dystonia/infantile parkinsonism.

Place, publisher, year, edition, pages
2003. Vol. 72, no 4, 444-453 p.
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-91750DOI: 10.1002/jnr.10606PubMedID: 12704806OAI: oai:DiVA.org:uu-91750DiVA: diva2:164582
Note

De två första författarna delar första författarskapet.

Available from: 2004-04-21 Created: 2004-04-21 Last updated: 2017-12-14Bibliographically approved
In thesis
1. Role of Bone Morphogenetic Proteins for Catecholaminergic Neurons in Vivo: Use of the Tyrosine Hydroxylase Locus for Cell-Specific inactivation of Signal Transduction
Open this publication in new window or tab >>Role of Bone Morphogenetic Proteins for Catecholaminergic Neurons in Vivo: Use of the Tyrosine Hydroxylase Locus for Cell-Specific inactivation of Signal Transduction
2004 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Members of the Transforming Growth factor-β (TGF-β) superfamily and its subclass Bone Morphogenetic Proteins (BMP) play important roles for nervous system development.

In order to study the BMP role for catecholaminergic neurons in vivo, we generated three knock-in mice, expressing the transgenes specifically in the targeting cells.

Two genetic modifications result in expression of dominant negative (dn) BMP receptors (BMPRII and ALK2). The tissue-specific expression was achieved by the transgene insertion into 3’- untranslated region of the endogenous gene for tyrosine hydroxylase (TH), the first enzyme in catecholamine biosynthesis. An Internal Ribosome Entry site (IRES) preceded inserted cDNAs, allowing for functional bicistronic mRNA production. While almost no defects in Th-IRES-dnALK2, the Th-IRES-dnBMPRII mouse demonstrated declined levels of catecholamines, including dopamine in the striatum. Losses of midbrain dopaminergic neurons (MDN) might cause the effect. Additionally, intermediate lines of these mice, preserving a neo-cassette, oriented opposite to the locus transcription, demonstrate dramatic decrease of catecholamine level, hence, represent models for rare catecholamine-deficiency diseases, including L-DOPA-responsive dystonia.

The third mouse, expressing in the same way Cre-recombinase (Th-IRES-Cre), represents a tool for catecholaminergic cell-limited deletion of any gene, which has to be flanked by loxP sites. Besides TH-positive areas, unexpected sites of Cre-recombination were identified, indicating regions of transient TH expression. Surprising recombination in oocytes opens a possibility to use our mouse as a general Cre-deletor.

Using TH-IRES-Cre mouse we generated tissue-specific knockout mice for two BMP signal transducers: Smad1 and Smad4 (also crucial for TGF-β). While no phenotype in Smad1 knockout, TH-IRES-Cre/Smad4 mouse revealed several defects including decreased level of striatal dopamine.

These results demonstrate a positive role of BMPs for MDN fate in vivo. Generated mice represent a tool-box for comprehensive study of the BMP function in catecholaminergic neurons. This study is of potential interest for understanding some aspects of Parkinson’s disease.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2004. 74 p.
Series
Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 0282-7476 ; 1350
Keyword
Neurosciences, Tyrosine Hydroxylase, Bone Morphogenetic Proteins (BMP), Transforming Growth Factor-β (TGF-β), tissue-specific knockout, Parkinson's disease, Neurovetenskap
National Category
Neurology
Identifiers
urn:nbn:se:uu:diva-4258 (URN)91-554-5964-1 (ISBN)
Public defence
2004-05-13, B21, BMC, Husargatan 3, Uppsala, 10:15
Opponent
Supervisors
Available from: 2004-04-21 Created: 2004-04-21Bibliographically approved
2. Experimental Studies of BMP Signalling in Neuronal Cells
Open this publication in new window or tab >>Experimental Studies of BMP Signalling in Neuronal Cells
2003 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The developing nervous system depends largely on extracellular cues to shape its complex network of neurons. Classically, neurotrophins are known to be important mediators in this process. More recently, Bone Morphogenetic Proteins (BMPs), belonging to the Transforming Growth Factor beta (TGFβ) superfamily of secreted cytokines, have been shown to exert a wide range of effects, such as cellular growth, differentiation, survival and apoptosis, both in the developing and adult nervous system. They signal via serine/threonine kinase receptor essentially to the Smad pathway, which carries the signal to the nucleus where the transcription of target genes is regulated.

This thesis investigates the functions of BMPs in the nervous system, using a set of different models. Firstly, a targeted deletion of GDF10 (BMP3b) in the mouse was established to evaluate the role of this growth/differentiation factor in the hippocampal formation, a brain area known to be involved in memory processing. Other members of the TGFβ superfamily likely compensate for the lack of GDF10, since no detectable alterations in hippocampal function or gene transcription profile have been found. Secondly, a mouse model was set up, with the aim to study impaired BMP-signalling in dopaminergic neurons. The tyrosine hydroxylase (TH) locus was used to drive the expression of dominant negative BMP receptors by means of bicistronic mRNAs. TH is the rate-limiting enzyme in the biosynthesis of catecholamine and the mice described, show a graded decrease of TH-activity resulting in severe to mild dopamine deficiency. The contribution of the dominant negative BMP receptors to the phenotype is however secondary to the apparent TH hypomorphism. The final theme of this thesis is the potentiating effects of BMPs on neurotrophin-induced neurite outgrowth as studied in explanted ganglia from chick embryos and in the rat phaeochromocytoma cell line PC12. A number of pharmacological inhibitors of intracellular signalling kinases were applied to the cultures in order to reveal the contribution of different pathways to the enhanced neurite outgrowth. We made the unexpected finding that inhibition of MEK signalling mimicked the potentiating effects of BMP stimulation in the chick system. The underlying mechanisms for the synergistic effects, however, are still an enigma.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2003. 75 p.
Series
Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 0282-7476 ; 1259
Keyword
Neurosciences, Bone Morphogenetic Protein (BMP), Growth Differentiation Factor 10 (GDF10), Aktivne-receptor Like Kinase 2 (ALK2), Tyrosine Hydroxylase (TH), Neurotrophic Factor, Neurite Outgrowth, Hippocampus, Catecholamine, PC12, Sympathetic Ganglia, Substantia Nigra (SN), Gene Targeting, Neurovetenskap
National Category
Neurology
Research subject
Developmental Neurosciences
Identifiers
urn:nbn:se:uu:diva-3398 (URN)91-554-5624-3 (ISBN)
Public defence
2003-05-16, B21, BMC, Uppsala, 09:15
Opponent
Supervisors
Available from: 2003-04-24 Created: 2003-04-24 Last updated: 2013-03-22Bibliographically approved

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