AMPA-gated glutamate receptors with an inwardly rectifying current-voltage (I-V) relationship and substantial Ca2+ permeability are expressed in a population of cultured rat hippocampal neurones (type II neurones). The inward rectification of these AMPA receptors was gradually lost in cell-free membrane patches. The I-V relationship of the AMPA receptors displayed a slight outward rectification in most patches 10 min after excision. This loss of inward rectification was not accompanied by a change in the Ca2+ permeability. The inward rectification was maintained by applying physiological concentrations of spermine to the cytoplasmic side of patch membranes. These results indicate that some cytoplasmic factor mediates inward rectification in the Ca(2+)-permeable AMPA receptors, and that a candidate for this substance is spermine.