Desensitization of NMDA receptors was studied at the single-channel level using outside-out patches taken from rat hippocampus CA neurons maintained short term in culture. The amount of desensitization that accumulated as a function of time during the application of NMDA (2.5 microM) was measured by the response to the rapid application of NMDA at a high concentration (100 microM). Records were sorted into two classes: those in which no openings were detected during the application of 2.5 microM NMDA, and those in which some channel openings occurred. Analysis of these data reveals that some desensitization accumulated when no channels opened, but that more desensitization developed when channels opened. The observations indicate that channels can pass from the closed state to the desensitized state, but that desensitization occurs more rapidly from the open state.