To detect the release of nitric oxide (NO) in brain tissue, an electrochemical microprobe was developed. The output current of the probe correlated linearly with the NO concentration at the tip, and the sensitivity of the probe was between 3.5 and 106 pA per 1 #M change in NO concentration. This probe showed no sensitivity to oxygen or to oxidized derivatives of NO. The NO release from sodium nitroprusside solution was successfully detected by the probe. An NO probe inserted into the molecular layer of a rat cerebellar slice detected a response corresponding to 8-58 nM of NO concentration following electrical stimulation of the white matter. This response was blocked reversibly by tetrodotoxin (1 /.tM) and was attenuated in the medium containing hemoglobin (1 or 10/.tM). The dependence of the response amplitude on the voltage at the cathode in the probe was the same as that of the NO-induced probe current. These results ensure that the NO probes developed in this study effectively detect the endogenous NO release in brain tissue.
Recently, an endothelium-derived relaxing factor (EDRF) has been identified as nitric oxide (NO) 20,21. NO release occurs not only from endothelial cells but also from cells in various tissues including that of the brain 3,10,13,16,17,19,22. Cerebellar cells liberate NO in response to excitatory amino acids 3,to which activate a calmodulin-dependent NO synthetase 4. It is likely that the NO release is induced normally as a consequence of synaptic transmission.
The measurement of NO release is usually done with bioassays based on physiological effects of NO such as the relaxation of blood vessels 20,21, the facilitation of guanylate cyclase activity 3,10,12,16,19,22 or the inhibition of platelet aggregation 22. The measurement of citrulline, a byproduct of NO synthesis, is also used 3,16,19. Direct chemical or physical measurements such as detection of chemiluminescence 9,19.20 could be fraught with uncertainty due to the contamination of nitrite ions (NO2), which are produced by the rapid oxidation of NO.
To measure PO 2 in neural tissue, a miniaturized oxygen electrode has been developed 23,24. This electrode is essentially a glass pipette whose opening is sealed with a thin rubber membrane. Only a gas of low molecular weight (such as 0 2) readily diffuses