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The combination of gene targeting and biochip technology to study neurotrophin function during brain development

Obiettivo



The develepment of the brain is characterized by a spatiotemporally controlled sequence of cell migration, synaptogenesis and formation of topographic maps. Neurotrophins such as brain derived neurotrophic factor BDNF are likely to be instrumental during these processess, though major aspects remain to be unraveled. To elucidate the regulatory function of BDNF in these interconnected developmental processes, a multidisciplinary approach is proposed by 3 research laboratories and a small enterprise from 3 european countries employing novel strategies and highly innovative combinations of gene and microchip technologies.
Stimulation-dependent BDNF expression. Induction of long term potentiation and epileptic seizures by short term supra-physiological electrical stimulation is accompanied by increased expression of BDNF. In order to allow molecular single cell analysis within a native neuronal network under more physiological conditions, hippocampi will be organotypically cultured directly on micro electrode arrays (MEA with 60 electrodes), which allows spatially controlled and continuous electrical stimulation over weeks. This most advanced biochip will be combined with fluorescent reporter gene technology. Introduction of green fluorescent protein at the activity required exon 3 BDNF promoter will enable us to evaluate the effects of longterm electrical stimulation paradigms on regulation of neurotrophin expression under physiological and pathophysiological conditions.
BDNF-dependent granular cell migration. In the dentate gyrus/hippocampus a life-long generation and defined migration of granular cells, which is correlated with the function of the neuronal network, takes place. To analyse the impact of BDNF on cell migration, conditional BDNF knockout mice with a restricted tissue-specific gene deficiency will be generated. Analysis of gene-targeted mice and of a novel xenograft transplantion model in vitro will reveal BDNF regulation of cell migration.
BDNF-dependent topographic map formation. The topographic projections of afferents from the contralateral hippocampus and enterorhinal cortex become sequentially separated into distinct sublayers of the hippocampus. Different neurothrophin knockouts will be used to analyse the impact of neurotrophins in this process in vivo and in vitro. Histological and biochemical assays will be used to monitor the influence of neurotrophins on synaptogenesis and the expression of the synapse-associated proteins SNARES.
Since all three aspects of BDNF regulation are causally linked, the sole use of the mouse hippocampus as unifying model will facilitate to unravel the multifunctionality of BDNF.
Precompetetive measureable objectives comprise 2 novel transgenic mice strains, fluoro biochip technology, and a prototype of a MEA stimulation device. The proposal objectives of neurotrophin function during hippocampal development comply with the objectives of the programme Biotechnology, Tasks 4.1 4. 4, and 4.5.

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CORDIS classifica i progetti con EuroSciVoc, una tassonomia multilingue dei campi scientifici, attraverso un processo semi-automatico basato su tecniche NLP. Cfr.: Il Vocabolario Scientifico Europeo.

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NATURWISSENSCHAFTLICHES UND MEDIZINISCHES INSTITUT AN DER UNIVERSITAET TUEBINGEN
Contributo UE
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55,Markwiesenstr. 55
72072 TUEBINGEN
Germania

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