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S. C. Neuhauss, C. M. Maurer, A. Lesslauer, H. B. Schönthaler, M. Gesemann; Glutamate Transport in the Zebrafish Retina. Invest. Ophthalmol. Vis. Sci. 2009;50(13):4000.
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© ARVO (1962-2015); The Authors (2016-present)
Glutamate is the main excitatory neurotransmitter in the vertebrate central nervous system, including the retina. At the photoreceptor synapse glutamate acts both as an excitatory (Off pathway) and inhibitory (On pathway) neurotransmitter. In order to ensure precise synaptic transmission and to avoid excitotoxicity, the removal of glutamate from the synapse is crucial for function and survival of postsynaptic cells. The clearance of glutamate is achieved by proteins of the excitatory amino acid transporter (EAAT) family, which consists of 5 members in mammals. Here we describe the cloning, genomic localization, phylogenetic analysis and expression of EAAT genes in the developing zebrafish retina.
Cloning and annotation of zebrafish EAAT genes was achieved by a combination of database searches and PCR based cloning of full length transcripts. Phylogenetic trees were constructed using a variety of bioinformatic approaches. Radiation hybrid mapping was used for genomic mapping of transcripts and expression was determined by in situ hybridization using digoxigenin labelled antisense probes. Morpholino-mediated knockdown and standard electroretinography was used to elucidate functional roles of EAAT’s.
We found a total of 11 EAAT genes, representing orthologs of the 5 mammalian counterparts. These genes map to different genomic locations, thereby excluding tandem duplications. The distribution of the paralogs within the phylogenetic tree can not solely be explained by the teleost specific genome duplication but suggests a lineage specific loss of EAAT genes in the mammalian lineage. We found retinal expression for most zebrafish EAAT genes, which is consistent with the expression pattern of their respective mammalian orthologs. However, we also found orthologs being expressed in different cell types than their mammalian counterparts. The knockdown of the glial EAAT2 leads to a loss of the b-wave in the electroretinogram, consistent with a major role in glutamate clearance from the photoreceptor synapse.
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