A chemiluminescence method of detecting nitric oxide (NO) in combination with a microdialysis technique was employed for the real-time measurement of NO production in living rat brain. This method based on the luminol-H2O2 system has a detection limit of 1 nM, and is the most sensitive method currently available for measuring NO. We applied this new technique to rat cerebellum to record the increase of chemiluminescence intensity arising from NO production after the injection of N-methyl-D-aspartate or kainic acid around the microdialysis probe. This highly sensitive method should be useful for the direct clarification of the functions of NO in the central nervous system.