Activation of P2X(7) receptors decreases glutamate uptake and glutamine synthetase activity in RBA-2 astrocytes via distinct mechanisms.
Glutamate clearance by astrocytes is critical for controlling excitatory neurotransmission and ATP is an important mediator for neuron-astrocyte interaction. However, the effect of ATP on glutamate clearance has never been examined. Here we report that treatment of RBA-2 cells, a type-2-like astrocyte cell line, with ATP and the P2X(7) receptor selective agonist 3'-O-(4-benzoylbenzoyl) adenosine 5'-triphosphate (BzATP) decreased the Na+-dependent [3H]glutamate uptake within minutes. Mechanistic studies revealed that the decreases were augmented by removal of extracellular Mg2+ or Ca2+, and was restored by P2X7 selective antagonist , periodate-oxidized 2',3'-dialdehyde ATP (oATP), indicating that the decreases were mediated through P2X(7) receptors. Furthermore, stimulation of P2X7 receptors for 2 h inhibited both activity and protein expression of glutamine synthetase (GS), and oATP abolished the inhibition. In addition, removal of extracellular Ca(2+) and inhibition of protein kinase C (PKC) restored the ATP-decreased GS expression but failed to restore the P2X(7)-decreased [3H]glutamate uptake. Therefore, P2X7-mediated intracellular signals play a role in the down-regulation of GS activity/expression. Activation of P2X7 receptors stimulated increases in intracellular Na+ concentration ([Na+](i)) suggesting that the P2X(7)-induced increases in [Na+](i) may affect the local Na+ gradient and decrease the Na+-dependent [3H]glutamate uptake. These findings demonstrate that the P2X7-mediated decreases in glutamate uptake and glutamine synthesis were mediated through distinct mechanisms in these cells.
Lo
Jun-Chih
JC
Institute of Neuroscience, National Yang Ming University, Taipei, Taiwan, Republic of China.
Huang
Wei-Chi
WC
Chou
Yun-Chia
YC
Tseng
Chun-Hsien
CH
Lee
Wei-Li
WL
Sun
Synthia H
SH
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D027322:Amino Acid Transport System X-AG / Q000235:genetics / Q000378:metabolism; D000818:Animals; D000831:Animals, Newborn; D001253:Astrocytes / Q000187:drug effects / Q000378:metabolism*; D001583:Benzoxazoles / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology; D004347:Drug Interactions; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005974:Glutamate-Ammonia Ligase / Q000378:metabolism*; D018698:Glutamic Acid / Q000378:metabolism*; D011806:Quinolinium Compounds / Q000378:metabolism; D051381:Rats; D018048:Receptors, Purinergic P2 / Q000502:physiology*; D058486:Receptors, Purinergic P2X7
D027322:Amino Acid Transport System X-AG; D001583:Benzoxazoles; D004791:Enzyme Inhibitors; D011806:Quinolinium Compounds; D018048:Receptors, Purinergic P2; D058486:Receptors, Purinergic P2X7; C089813:YO-PRO 1; D018698:Glutamic Acid; C033901:3'-O-(4-benzoyl)benzoyladenosine 5'-triphosphate; D000255:Adenosine Triphosphate; D005974:Glutamate-Ammonia Ligase; D002118:Calcium