During brain ischaemia a release of glutamate into the extracellular space, largely by reversal of uptake by glutamate transporters (Rossi et al. 2000) and independent of action potentials, triggers neuronal death. The rundown of ion gradients in ischaemia is also expected to reverse uptake by GABA transporters. GABA release and consequent GABA_A receptor activation could be neuroprotective because of the hyperpolarization it may produce, but might contribute to neuronal death by activating a Cl^– influx which facilitates water influx and cell swelling.

We whole-cell clamped CA1 pyramidal cells at 32-34 °C in hippocampal slices of P12 rats (killed humanely according to Home Office regulations), simulated ischaemia chemically (Rossi et al. 2000), and used the cells’ GABA_A receptors to sense the extracellular GABA concentration. The intracellular solution had a chloride concentration giving E_Cl = 0 mV. As reported by Rossi et al. (2000), after 6-7 min ischaemia, a large inward current which decayed to a maintained plateau was recorded at -30 mV, which in part reflects glutamate release.

When using an internal solution providing slow calcium buffering (containing 5 mM EGTA and 0.5 mM CaCl₂), applying GABA (1 mM) or the GABA_A receptor blocker GABAzine (10 µM) during the ischaemia-induced current plateau evoked no change in current, whereas with strong calcium buffering (10 mM BAPTA and 0 mM CaCl₂) GABA evoked an extra inward current and GABAzine suppressed approximately one-third of the plateau current. This suggests that with the EGTA-based internal solution the rise of [Ca]_i occurring during ischaemia leads to desensitization of GABA_A receptors (Stelzer & Shi, 1994; Harata et al. 1997), but when [Ca]_i is kept low with the BAPTA-based internal the whole-cell clamped neuron can be used to sense ischaemia-evoked GABA release. With the BAPTA-based internal solution, the GABAzine-blocked current during the ischaemic plateau was not significantly affected by TTX (1 µM, P = 0.47 by 2-tailed t test, 6 cell pairs), but the presence of SKF-89976a (100 µM) to block the neuronal GABA transporter GAT-1 reduced the GABAzine-suppressed current by 70 % (P = 0.0034, 8 cell pairs).

These data suggest that there is a significant elevation of extracellular GABA concentration during ischaemia, and that much of the GABA release occurs by reversed uptake.

This work was supported by The Wellcome Trust, a Wolfson-Royal Society award, a Burroughs-Wellcome Fellowship (D.J.R.), and the 4 year PhD Programme in Neuroscience at UCL (N.J.A.).