title: P2Y1 receptor inhibits GABA transport through a calcium signalling-dependent mechanism in rat cortical astrocytes.
abstract: Astrocytes express a variety of purinergic (P2) receptors, involved in astrocytic communication through fast increases in [Ca(2+) ]i . Of these, the metabotropic ATP receptors (P2Y) regulate cytoplasmic Ca(2+) levels through the PLC-PKC pathway. GABA transporters are a substrate for a number of Ca(2+) -related kinases, raising the possibility that calcium signalling in astrocytes impact the control of extracellular levels of the major inhibitory transmitter in the brain. To access this possibility we tested the influence of P2Y receptors upon GABA transport into astrocytes. Mature primary cortical astroglial-enriched cultures expressed functional P2Y receptors, as evaluated through Ca(2+) imaging, being P2Y1 the predominant P2Y receptor subtype. ATP (100 μM, for 1 min) caused an inhibition of GABA transport through either GAT-1 or GAT-3 transporters, decreasing the Vmax kinetic constant. ATP-induced inhibition of GATs activity was still evident in the presence of adenosine deaminase, precluding an adenosine-mediated effect. This, was mimicked by a specific agonist for the P2Y1,12,13 receptor (2-MeSADP). The effect of 2-MeSADP on GABA transport was blocked by the P2 (PPADS) and P2Y1 selective (MRS2179) receptor antagonists, as well as by the PLC inhibitor (U73122). 2-MeSADP failed to inhibit GABA transport in astrocytes where intracellular calcium had been chelated (BAPTA-AM) or where calcium stores were depleted (α-cyclopiazonic acid, CPA). In conclusion, P2Y1 receptors in astrocytes inhibit GABA transport through a mechanism dependent of P2Y1 -mediated calcium signalling, suggesting that astrocytic calcium signalling, which occurs as a consequence of neuronal firing, may operate a negative feedback loop to enhance extracellular levels of GABA.
authors.affiliation: Institute of Pharmacology and Neurosciences, Faculty of Medicine, University of Lisbon, Lisbon, Portugal; Neurosciences Unit, Institute of Molecular Medicine University of Lisbon, Lisbon, Portugal.
authors.forename: Pedro F
authors.initials: PF
authors.lastname: Jacob
authors.forename: Sandra H
authors.initials: SH
authors.lastname: Vaz
authors.forename: Joaquim A
authors.initials: JA
authors.lastname: Ribeiro
authors.forename: Ana M
authors.initials: AM
authors.lastname: Sebastião
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001253:Astrocytes / Q000187:drug effects / Q000502:physiology*; D001693:Biological Transport, Active / Q000187:drug effects / Q000502:physiology*; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000187:drug effects / Q000502:physiology*; D050485:GABA Plasma Membrane Transport Proteins / Q000378:metabolism; D054801:Phosphoinositide Phospholipase C / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism; D017208:Rats, Wistar; D058472:Receptors, Purinergic P2Y1 / Q000378:metabolism*; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
chemical_list: D050485:GABA Plasma Membrane Transport Proteins; D058472:Receptors, Purinergic P2Y1; C493417:Slc6a11 protein, rat; D005680:gamma-Aminobutyric Acid; D000255:Adenosine Triphosphate; D011493:Protein Kinase C; D054801:Phosphoinositide Phospholipase C; D002118:Calcium
keywords: ATP; GABA transporters; P2Y1 receptors; astrocytes; calcium signalling