Effect of hyperosmotic conditions on the expression of the betaine-GABA-transporter (BGT-1) in cultured mouse astrocytes.
The adaptation of cells to hyperosmotic conditions involves accumulation of organic osmolytes to achieve osmotic equilibrium and maintenance of cell volume. The Na+ and Cl(-)-coupled betaine/GABA transporter, designated BGT-1, is responsible for the cellular accumulation of betaine and has been proposed to play a role in osmoregulation in the brain. BGT-1 is also called GAT2 (GABA transporter 2) when referring to the mouse transporter homologue. Using Western Blotting the expression of the mouse GAT2 protein was investigated in astrocyte primary cultures exposed to a growth medium made hyperosmotic (353+/-2.5 mosmol/kg) by adding sodium chloride. A polyclonal anti-BGT-1 antibody revealed the presence of two characteristic bands at 69 and 138 kDa. When astrocytes were grown for 24 h under hyperosmotic conditions GAT2 protein was up-regulated 2-4-fold compared to the level of the isotonic control. Furthermore, the expected dimer of GAT2 was also up-regulated after 24 h under the hyperosmotic conditions. The [3H]GABA uptake was examined in the hyperosmotic treated astrocytes, and characterized using different selective GABA transport inhibitors. The up-regulation of GAT2 protein was not affecting total GABA uptake but the hyperosmotic condition did change total GABA uptake possibly involving GAT1. Immunocytochemical studies revealed cell membrane localization of GAT2 throughout astroglial processes. Taken together, these results indicate that astroglial GAT2 expression and function may be regulated by hyperosmolarity in cultured mouse astrocytes, suggesting a role of GAT2 in osmoregulation in neural cells.
Olsen
Mads
M
Department of Pharmacology, The Danish University of Pharmaceutical Sciences, Universitetsparken 2, Copenhagen, DK-2100, Denmark.
Sarup
Alan
A
Larsson
Orla M
OM
Schousboe
Arne
A
D000818:Animals; D001253:Astrocytes / Q000378:metabolism*; D015153:Blotting, Western; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D003470:Culture Media; D050485:GABA Plasma Membrane Transport Proteins; D007150:Immunohistochemistry; D051379:Mice; D009994:Osmolar Concentration; D005680:gamma-Aminobutyric Acid / Q000378:metabolism
D002352:Carrier Proteins; D003470:Culture Media; D050485:GABA Plasma Membrane Transport Proteins; C072207:betaine plasma membrane transport proteins; D005680:gamma-Aminobutyric Acid