ACTIVATION OF GLUTAMATE RECEPTORS AND GLUTAMATE UPTAKE IN IDENTIFIED MACROGLIAL CELLS IN RAT CEREBELLAR CULTURES

1. Patch-clamp methods have been used to examine the action of excitatory amino acids on three types of glial cell in cultures of rat cerebellum, namely type-1-like astrocytes, type-2 astrocytes and oligodendrocytes. In addition we have examined glutamate sensitivity of the precursor cell (the O-2A progenitor) that gives rise to type-2 astrocytes and oligodendrocytes. 2. Glutamate (30-mu-M), quisqualate (3-100-mu-M), (S)-alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA, 10-30-mu-M) and kainate (10-500-mu-M) were applied to cerebellar type-2 astrocytes examined under whole-cell voltage clamp. Each of these agonists induced inward currents in cells held at negative membrane potentials. The currents reversed direction near O mV holding potential. N-Methyl-D-aspartate (NMDA, 30-100-mu-M) or aspartate (30-mu-M) in the presence of glycine (1-mu-M) did not evoke any whole-cell current changes in type-2 astrocytes. 3. The distribution of glutamate receptors in type-2 astrocytes was mapped with single- or double-barrelled ionophoretic pipettes containing quisqualate or kainate. Application of these agonists (current pulses 100 ms, 50-100 nA) to cells held at -60 mV evoked inward currents of 20-120 pA in the cell soma and 10-80 pA in the processes. Responses could also be obtained at the extremities of processes (approximately 60-mu-m from the soma). 4. Quisqualate or kainate (at 30-mu-m) applied to O-2A progenitor cells from rat cerebellum or optic nerve induced whole-cell currents (quisqualate 20-30 pA; kainate 20-50 pA, holding potential, V(h) = -60 mV) that reversed near 0 mV. In common with type-2 astrocytes, the progenitor cells did not respond to NMDA (30-mu-M). 5. Type-1-like astrocytes produced large inward currents to glutamate (30-mu-M). These currents remained inward-going at holding potentials as positive as + 80 mV and were not accompanied by any apparent noise increase. This result can be explained by the presence of an electrogenic glutamate uptake carrier. In cells kept up to 4 days in vitro, quisqualate, kainate and NMDA each failed to produce any whole-cell current changes, indicating the absence of receptors in type-1-like astrocytes at this stage in culture. Furthermore the glutamate uptake currents in type-1-like astrocytes were inhibited when external Na+ was replaced by Li+, although Li+ was found to pass through the glutamate channel in type-2 astrocytes. 6. Oligodendrocytes in cerebellar cultures did not respond to glutamate, quisqualate or kainate indicating a lack both of a detectable electrogenic glutamate uptake mechanism and of glutamate receptor ion channels in these cells. 7. We conclude that, of the various macroglial cells, only the type-2 astrocyte and its progenitor cell posses 'fast' glutamate receptor channels in short-term (< 4 days) cultures. Detectable uptake currents were confined to type-1-like astrocytes. However, in older cultures (> 7 days) type-1-like astrocytes also developed glutamate receptor channels, in addition to their uptake currents. The possible involvment of glial glutamate receptors and glutamate uptake in neuronal-glial interaction is considered.