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(Investigative Ophthalmology and Visual Science. 2004;45:1442-1449.)
© 2004 by The Association for Research in Vision and Ophthalmology, Inc.
DOI:  10.1167/iovs.03-0062
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Retinal Colocalization and In Vitro Interaction of the Glutamate Receptor EAAT3 and the Serum- and Glucocorticoid-Inducible Kinase SGK1

Roman Schniepp,1 Konrad Kohler,2 Thomas Ladewig,2 Elke Guenther,2 Guido Henke,1 Monica Palmada,1 Christoph Boehmer,1 Jeffrey D. Rothstein,3 Stefan Bröer,4 and Florian Lang1

1From the Department of Physiology I, University of Tübingen, Germany; the 2Department of Experimental Ophthalmology, University Eye Hospital, Tübingen, Germany; the 3Department of Neurology, John Hopkins University School of Medicine, Baltimore, Maryland; and the 4School of Biochemistry and Molecular Biology, Australian National University, Canberra, Australia.

PURPOSE. The serum- and glucocorticoid-inducible kinase SGK1 regulates several epithelial channels and transporters, the related protein kinase B (PKB) regulates glucose transport. SGK1 is expressed in the brain and could thus regulate glial and/or neuronal transport processes. The present study explores whether SGK1 is expressed in the retina and whether it regulates EAAT3, a Na+-coupled glutamate transporter. EAAT3 is expressed in retinal ganglion cells and accomplishes the clearance of glutamate from synaptic clefts.

METHODS. Immunohistochemistry was performed to test for retinal SGK1 expression. For functional analysis, cRNA encoding EAAT3 was injected into Xenopus oocytes with or without additional injection of wild-type SGK1, constitutively active S422DSGK1, inactive K127NSGK1, and/or constitutively active T308D,S473DPKB. Glutamate induced current (IGLU) was taken as a measure for transport.

RESULTS. SGK1 is indeed expressed in several retinal cells including retinal ganglion cells where it is colocalized with EAAT3. In EAAT3-expressing Xenopus oocytes, glutamate-induced current was stimulated by coexpression of wild-type SGK1, constitutively active S422DSGK1, and constitutively active T308D,S473DPKB, but not by inactive K127NSGK1.

CONCLUSIONS. SGK1 and EAAT3 are coexpressed in retinal neurons, and SGK1 serves to stimulate EAAT3. This function is shared by protein kinase B (PKB). The experiments reveal a novel mechanism regulating EAAT3, which may be essential for the function of the retinal ganglion cells.




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