P2Y₁receptor 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.

BDNF, via truncated TrkB receptor, modulates GlyT1 and GlyT2 in astrocytes

“…To clarify if the decrease could be attributed to the impairment of GlyT insertion pathways in the plasma membrane or the enhancement of the endocytic cell machinery, well known pharmacological tools were used: monensin, that blocks GlyT recycling back to the membrane without influencing endocytosis (Mollenhauer et al, 1990) and dynasore, which inhibits the dynamin/clathrin-dependent endocytosis (Macia et al, 2006). The intracellular Ca 21 chelator, BAPTA-AM, which affects Ca 21 signaling in astrocytes (Jacob et al, 2014), did not prevent the BDNF effect. Dynasore inhibits the dynamin/clathrin-dependent endocytosis, but other types of endocytosis have already been described in astrocytes, namely an endocytic pathway independent of clathrin and dynamin but tightly regulated by intracellular Ca 21 (Jiang and Chen, 2009).…”

Reduced Expression of P2Y2 Receptor and Acetylcholinesterase at Neuromuscular Junction of P2Y1 Receptor Knock-out Mice

“…ATP is co-stored with other neurotransmitters, such as acetylcholine (ACh), in a ratio of about 1:5, in which ATP is found to stabilize ACh and co-released from the pre-synaptic vesicles in vertebrate neuromuscular junction (nmj). Upon pre-synaptic neural stimulation, ATP is released and triggers a series of signal transduction, which cross-talks with other co-released neurotransmitter-stimulated signaling pathways, and results in regulating post-synaptic gene expressions (Noguchi et al 2013;Yuan et al 2013;Jacob et al 2014;Yu et al 2014).…”

Hippocampal GABAergic transmission: a new target for adenosine control of excitability

“…) GABA transport into cultured astrocytes, the main difference in both studies being the removal (Jacob et al . ) or not (Cristóvão‐Ferreira et al . ) of microglia, which is known to release purines.…”

“…) or facilitate (Jacob et al . ) GABA transport into cultured astrocytes, the main difference in both studies being the removal (Jacob et al . ) or not (Cristóvão‐Ferreira et al .…”

“…This suggests that slight changes in the adenosine concentrations in the extracellular space will determine the direction of the influence of adenosine on GABA transport into astrocytes. Accordingly, it has been shown that extracellular addition of adenosine deaminase, which degrades extracellular adenosine, can either inhibit (Crist ovão-Ferreira et al 2013) or facilitate (Jacob et al 2014) GABA transport into cultured astrocytes, the main difference in both studies being the removal (Jacob et al 2014) or not (Crist ovão-Ferreira et al 2013) of microglia, which is known to release purines.…”