EFFLUX OF GAMMA-AMINOBUTYRIC-ACID CAUSED BY CHANGES IN ION CONCENTRATIONS AND CELL SWELLING SIMULATING THE EFFECT OF CEREBRAL-ISCHEMIA

The relationships among ischaemic GABA efflux from brain tissue and ex tracellular and intracellular concentrations of sodium, chloride and p otassium ions were investigated by means of 1) transverse hippocampal slices from rat and 2) functional expression of a high affinity GABA t ransporter in Xenopus oocytes. Brain slices were incubated for 20 min in medium where extracellular sodium and chloride were substituted wit h impermeant ions. Isethionate (Iseth) substitution for chloride gener ated a 7-fold increase in GABA efflux. Choline (Chol) but not N-methyl -D-glucamine (NMDG) substitution for sodium likewise increased GABA ef flux. Reducing the osmolarity of the medium by decreasing both sodium and chloride concentrations (Hyp) increased GABA efflux 3-fold. This r elease was blocked by mannitol (Man). Blocking sodium channels with 1 mu M of tetrodotoxin (TTX) also increased the release 3-fold. Energy d eprivation (ED) increased the GABA release 50-fold. ED/Iseth left the release unchanged, ED/Chol increased the GABA efflux by 23%, whereas E D/NMDG reduced the release by 41%;. Adding mannitol did not block the ED-evoked release, whereas TTX reduced it by 52%. Release of preloaded [H-3]-GABA from oocytes expressing the GAT-1 GABA transporter was the n examined. Depolarisation by current injection or 100 mM extracellula r K+ did not increase GABA release. Sodium chloride injection, however , caused membrane depolarisation and a 100-fold increased GABA efflux from the oocytes. This release was blacked when the osmolarity was inc reased extracellularly by adding mannitol. These results show that 1) TTX releases GABA from brain tissue but blocks release during ED, 2) t he high affinity GABA carrier must be altered in order to reverse, 3) ischaemic GABA release is sodium independent, and is modulated by larg e cations, 4) mannitol blocks the reversal of high affinity carriers i n oocytes, but the release from brain slices during ED is unaffected. Taken together, the results suggest that ischaemic release of GABA fro m brain tissue does not occur by means of reversed high affinity carri ers alone, but rather that it is controlled by more complex mechanisms .