Rapid increase of glial glutamate uptake via blockade of the protein kinase A pathway.
Glutamate is the main excitatory neurotransmitter in the vertebrate central nervous system. Removal of this transmitter from the synaptic cleft by glial and neuronal transporter systems plays an important role in terminating glutamatergic neurotransmission. The effects of different activators and blockers of PKA and PKC on glutamate uptake were studied in primary glial cells cultivated from the rat cortex using the patch-clamp recording technique and immunocytochemical methods. GF 109203X enhances glutamate-induced membrane currents in a concentration- and time-dependent manner. After pre-application for 40 s the maximal transport capacity was increased by 30-80%. The estimated Km-value of the transport system did not change after drug application and the enhanced glutamate uptake was reversible within a few minutes upon washout. Activators and blockers of the PKC pathway did not affect glutamate uptake, whereas H89, a selective blocker of PKA, mimicked the effects of GF 109203X, indicating involvement of the protein kinase A pathway. The GF 109203X-induced increase in transport capacity is likely to be mediated by GLAST since the GLT-1 selective blocker dihydrokainate was unable to block basal or stimulated glutamate uptake. Furthermore, the increase in transport activity may well be based on an increase in cell surface expression of the transporter protein since preincubation with cytochalasin-B, a protein that blocks actin polymerization, almost completely abolished the effect of GF 109203X and H89. These results indicate that GF 109203X and H89 enhance glial glutamate uptake via blockade of the PKA. The described effect may affect glutamatergic neurotransmission by reducing the glutamate concentration in the synaptic cleft.
Adolph
Oliver
O
Department of Anesthesiology, University Hospital of Ulm, Steinhoevelstrasse 9, 89075 Ulm, Germany. oliver.adolph@uniklinik-ulm.de
Köster
Sarah
S
Räth
Monika
M
Georgieff
Michael
M
Weigt
Henry U
HU
Engele
Jürgen
J
Senftleben
Uwe
U
Föhr
Karl J
KJ
D027322:Amino Acid Transport System X-AG / Q000502:physiology; D000818:Animals; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors*; D004305:Dose-Response Relationship, Drug; D004553:Electric Conductivity; D004791:Enzyme Inhibitors / Q000008:administration & dosage / Q000494:pharmacology*; D027341:Excitatory Amino Acid Transporter 1 / Q000378:metabolism; D018698:Glutamic Acid / Q000378:metabolism*; D007211:Indoles / Q000008:administration & dosage / Q000494:pharmacology*; D007546:Isoquinolines / Q000494:pharmacology; D008301:Maleimides / Q000008:administration & dosage / Q000494:pharmacology*; D009457:Neuroglia / Q000187:drug effects / Q000378:metabolism*; D009994:Osmolar Concentration; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051381:Rats; D013449:Sulfonamides / Q000494:pharmacology; D013997:Time Factors; D014018:Tissue Distribution
D027322:Amino Acid Transport System X-AG; D004791:Enzyme Inhibitors; D027341:Excitatory Amino Acid Transporter 1; D007211:Indoles; D007546:Isoquinolines; D008301:Maleimides; D047428:Protein Kinase Inhibitors; C493392:Slc1a3 protein, rat; D013449:Sulfonamides; D018698:Glutamic Acid; D017868:Cyclic AMP-Dependent Protein Kinases; C070515:bisindolylmaleimide I; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide