P2X7Receptor-Mediated Release of Excitatory Amino Acids from Astrocytes

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Abstract
Astrocyte glutamate release can modulate synaptic activity and participate in brain intercellular signaling. P2X7receptors form large ion channels when activated by ATP or other ligands. Here we show that P2X7 receptors provide a route for excitatory amino acid release from astrocytes. Studies were performed using murine cortical astrocyte cultures. ATP produced an inward current in patch-clamped astrocytes with properties characteristic of P2X7 receptor activation: the current was amplified in low divalent cation medium, blocked by pyridoxal phosphate-6-azophenyl-2′,4′-disulfonic acid (PPADS), and more potently activated by 3′-O-(4-benzoyl)benzoyl ATP (BzATP) than by ATP itself. Measurement of current reversal potentials showed the relative BzATP-induced permeabilities to different substrates to be Na+, 1 > Cl−, 0.34 >N-methyl-d-glucamine, 0.27 >l-glutamate, 0.15 ≈ d-aspartate, 0.16. Astrocytes exposed to BzATP also became permeable to Lucifer yellow, indicating a large channel opening. Release of l-glutamate and d-aspartate through P2X7 channels was confirmed using radiolabeled tracers. As with the inward current, release of glutamate and d-aspartate was induced by BzATP more potently than ATP, amplified in Ca2+/Mg2+-free medium, and blocked by PPADS or oxidized ATP. Efflux through P2X7channels is a previously unrecognized route of ligand-stimulated, nonvesicular astrocyte glutamate release.