ETHANOL-INDUCED CL- FLUX IN RAT CEREBELLAR GRANULE CELLS AS MEASURED BY A FLUORESCENT-PROBE

Cl- fluxes through the GABA(A) receptor gated ion channels in cultured rat cerebellar granule cells were measured using the chloride-sensitive fluorescent probe SPQ (6-methoxy-N-(3-sulphopropyl)quinolinium) incorporated into the cells. The fluorescence of SPQ is quenched by Cl- ions. The cells were bathed in a low Cl- medium so that the Cl- gradient was directed outward. Ethanol increased the SPQ fluorescence indicating a decrease in intracellular Cl- due to Cl- efflux. Picrotoxin inhibited the effect at low concentrations of ethanol (< 50 mM) in a concentration dependent manner. The effects of ethanol were potentiated at low concentrations (< 10-mu-M) of gamma-aminobutyric acid (GABA), but inhibited at higher concentrations (0.3-2.0 mM). The results support the hypothesis that ethanol may act via the GABA(A) receptor gated ion channel. The results also suggest that SPQ is a suitable probe for measuring GABA(A) receptor-coupled Cl- fluxes through the GABA(A) receptor-gated channels in living cells.