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Department of Pharmacology, University College London, UK.
1. Patch-clamp methods have been used to characterize GABA-and glycine-activated channels and spontaneous synaptic currents in granule cells in thin cerebellar slices from 7- to 20-day-old rats. 2. All granule cells responded to 10 microM GABA, while approximately 60% responded to 100 microM glycine. With repeated against application, whole-cell responses to GABA, but not those to glycine, declined over a period of minutes unless the pipette solution contained Mg-ATP. 3. Whole-cell concentration-response curves gave EC50 values at 45.2 and 99.6 microM and Hill slopes of 0.94 and 2.6 for GABA and glycine, respectively. At saturating concentrations, currents evoked by GABA were fivefold larger than those evoked by glycine. 4. Whole-cell current-voltage (I-V) relationships of GABA- and glycine-activated currents reversed close to the predicted Cl- equilibrium potential. Partial replacement of intracellular Cl- with F- shifted the GABA reversal potential to a more negative value. 'Instantaneous' I-V relationships produced by ionophoretic application of GABA were linear, while 'steady-state' I-V relationships produced by ramp changes in potential showed outward rectification. For glycine, 'steady-state' I-V plots were linear. 5. Responses to GABA were blocked by the GABAA receptor antagonists bicuculline (15 microM), SR-95531 (10 microM) and picrotoxinin (100 microM) while responses to glycine were selectively blocked by strychnine (200 nM), indicating the presence of two separate receptor types. 6. In outside-out membrane patches, GABA opened channels with conductances of 16 and 28 pS. The proportion of openings to each of the conductances varied between patches, possibly indicating the activation of two distinct channel types. Glycine-activated single-channel currents had conductances of 32, 55 and 104 pS. Single-channel I-V relationships were linear. 7. Spontaneous synaptic currents with a rapid rise time and biexponential decay were present in more than half of the cells examined. These currents were eliminated by bicuculline (15 microM) or SR-95331 (10 microM) and were greatly reduced in frequency by tetrodotoxin (TTX; 300 nM), suggesting that they were mediated by GABA and arose from spontaneous activity in Golgi interneurones. In granule cells where this spontaneous synaptic activity was apparent, glycine and low concentrations of GABA increased the frequency of the synaptic currents.(ABSTRACT TRUNCATED AT 400 WORDS)
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