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Arbeitsgruppe - Simon Arbeitsgruppe - Simon Dr. Horst Simon Institut für Anatomie and Zellbiologie III Universität Heidelberg Im Neuenheimer Feld 307 69120 Heidelberg, Germany Tel: - 49 - 6221 - 54 8342 Fax: - 49 - 6221 - 54 5605 E-Mail: [email protected] Development of the midbrain dopaminergic neurons PhD 1993 Guy`s Hospital, University of London, UK, Postdoctoral work at the Salk Institute, La Jolla, USA, Group Leader at the Department of Anatomy and Cell Biology III, IZN, since July 1999. 1 Current Research Structure of the Group Group Leader Horst Simon Graduate students Sandrine Thuret Paola Sgadó Murat-Burak Yaylaoglu Daniel Gherbassi Lavinia Alberí Lavinia Bhatt Undergraduate student Christian Scholz Technicians Gabi Döderlein Current Research The dopaminergic neurons (DA) of the substantia nigra and ventral tegmentum are located in the midbrain and are the main source of dopamine in the vertebrate central nervous system. They play a major role in the modulation of the neurons in the basal ganglia, a component of the vertebrate motor system. Parkinson?s Disease, one of the most prominent human neurodegenerative disease, is directly linked to loss of these neurons. Engrailed and the midbrain dopaminergic neurons 2 Engrailed and the midbrain dopaminergic neurons Transcription factors are DNA binding elements, which regulate the expression of other genes as activators or repressors. They play a major role in the formation and maintenance of cellular properties during development and in the adult. En-1 and En-2 are two homeodomain containing transcription factors which were originally cloned by sequence homology to the Drosophila engrailed gene. One of their main expression domain is the cluster of DA neurons in the ventral midbrain. We demonstrated recently by mouse mutant analysis that engrailed expression is essential for the proper differentiation and maintenance of the midbrain DA neurons. The DA neurons of mice deficient of En-1 and En-2 develop normally in the ventral midbrain and start to express molecules typical for their subclass, like for example tyrosine hydroxylase (see image below). Soon thereafter, they disappear at developmental stage when En-1 and En-2 are normally expressed in the wildtype. Furthermore, these studies revealed that a-Synuclein, a membrane associated protein, which was recently genetically linked to Parkinson?s Disease, is dependent on its expression in the DA neurons on the presence of these two transcription factors. Downstream targets of the engrailed genes P0 mouse brain, wildtype(right) and mutant (left) deficient of En-1 and En-2 stained against tyrosine hydroxylase delineating the midbrain dopaminergic neurons (arrow), which are absent in the mutant. The disappearance of the substantia nigra and ventral tegmentum in mutant mice for En-1 and En-2 suggests that some essential downstream targets are wrongly regulated in their expression. We applied a recently 3 Downstream targets of the engrailed genes modified differential display method to identify genes which are under regulatory control of the En-1 and En-2. Up to date, we identified several genes specifically expressed in the DA neurons and started to analyze their function by mouse mutant studies and in cell culture. Future Projects and Goals The developmental cascade which leads to the proper formation of the DA neurons in the ventral midbrain are not very well understood. Up to date, only a small amount of genes were successfully linked to their development. Furthermore, only little is known of the proteins which Parkinson's Disease acts upon. Our main emphasis will be the isolation, identification and functional characterization of genes, which are key players in this developmental cascade. The differential display identified several cDNA fragments which are likely candidate molecules. The short term aim will be to analyze these cDNA fragments and to determine their in vivo function. This analysis will include the creation of mouse knock out animals and conditional mutants if required or the use of cell culture systems. Long term goal will be to determine the regulatory elements which underlies the proper initiation of the expression of these genes and how this expression is maintained during the adult. In situ hybridization of differentially expressed genes on P0 brain sections. Each shows a distinct expression in the DA neurons. Furthermore, the laboratory tries to identify the molecules involved in guiding the axons originating from the DA neurons in the substantia nigra and ventral tegmentum to the developing basal ganglia. We will use explant cultures in 3-D collagen matrices to identify molecules with tropic or trophic activity towards the midbrain DA neurons. Since one of the therapeutical approaches to PD is the transplantation of embryonic tissue containing the DA neurons, such molecules could be administered in conjunction with embryonic tissue implants to facilitate their establishment in the basal ganglia. 4 Selected Publications Selected Publications [1] Simon, H., and Lumsden, A. (1993). Rhombomere-specific origin of the contralateral vestibulo-acoustic efferent neurons and their migration across the embryonic midline, Neuron 11, 209-20. [2] Gassmann, M., Casagranda, F., Orioli, D., Simon, H., Lai, C., Klein, R., and Lemke, G. (1995). Aberrant neural and cardiac development in mice lacking the ErbB4 neuregulin receptor, Nature 378, 390-4. [3] Simon, H., Hornbruch, A., and Lumsden, A. (1995). Independent assignment of antero-posterior and dorso-ventral positional values in the developing chick hindbrain, Curr Biol 5, 205-14. [4] Yee, K. T., Simon, H. H., Tessier-Lavigne, M., and O'Leary, D. M. (1999). Extension of long leading processes and neuronal migration in the mammalian brain directed by the chemoattractant netrin-1, Neuron 24, 607-22. 5