Modulation of extracellular glutamate concentration in rat brain slices bycystine-glutamate exchange

1. The role of cystine-glutamate exchange in controlling the extracellular glutamate concentration in the central nervous system was examined by whole -cell clamping neurons in rat brain dices, and using their glutamate recept ors as sensors of extracellular glutamate concentration. 2. Applying cystine to cerebellar slices generated a membrane current in Pu rkinje cells which was abolished by glutamate receptor blockers. Similar cy stine-evoked currents were seen in pyramidal cells of frontal cortex slices . 3. Control experiments on non-N-methyl-D-aspartate (non-NMDA) receptors in enzymatically isolated Purkinje cells showed that cystine did not produce a current in slice Purkinje cells by directly activating glutamate receptors , nor by potentiating the action of background levels of glutamate on recep tors. Experiments on isolated salamander Muller cells showed that cystine d id not block Na+-dependent GLAST glutamate transporters (homologous to the transporters in the Bergmann glia ensheathing the Purkinje cells), nor did it block the current produced by EAAT4 and EAAC1. glutamate transporters in Purkinje cells. Thus the cystine-evoked current in Purkinje cells is not d ue to a rise in extracellular glutamate concentration caused by block of Na +-dependent uptake. 4. The dependence of cystine-evoked current on cystine concentration in sli ce Purkinje cells could be fitted by a Michaelis-Menten relation with a K-m of 250 mu M. The K-m predicted from this for cystine activating glutamate efflux is less than 140 mu M, because of the non-linear dependence on gluta mate concentration of the Purkinje cell current. The current evoked by 1 mM cystine was little affected by removal of extracellular chloride or additi on of 1 mM furosemide (frusemide), but was potentiated by 1 mM 4,4'-diisoth iocyanatostilbene-2,2'-disulfonic acid (DIDS). 5. These data suggest that external cystine generates a current in slice Pu rkinje cells by activating cystine-glutamate exchange in cells of the slice , releasing glutamate which activates non-NMDA receptors in the Purkinje ce ll membrane.