The effect of carbamazepine (CBZ) on N-methyl-D-aspartate (NMDA)-stimulated CA++ influx in rat cerebellar granule cells was studied by use of fura-2 microfluorometry. CBZ inhibited the rise in intracellular free Ca++ concentration ([Ca++]i) induced by NMDA and glycine in a rapid reversible and concentration-dependent manner. CBZ's inhibition of the [Ca++]i increase was noncompetitive with respect to NMDA, glycine and the facilitatory neurosteroid pregnenolone sulfate. The degree of inhibition of the NMDA response produced by CBZ increased with increasing concentrations of extracellular KCl. Excluding non-NMDA receptor-mediated contributions to Ca++ influx, depolarization by 50 mM KCl resulted in a 20-fold decrease (from 723 to 33 microM) in the IC50 for CBZ blockade of the NMDA response. Thus, significant blockade of NMDA receptor responses in cerebellar granule cells can occur at concentrations of CBZ within the therapeutic range under conditions believed to accompany seizures. Moreover, the common toxic side effects of CBZ, which include signs of cerebellar dysfunction, may occur as a result of CBZ blockade of the NMDA receptors of cerebellar granule cells.