ALDEHYDE FIXATION DIFFERENTIALLY AFFECTS DISTRIBUTION OF DIAPHORASE ACTIVITY BUT NOT OF NITRIC-OXIDE SYNTHASE IMMUNOREACTIVITY IN RAT-BRAIN

The effect of aldehyde fixation on NADPH- and NADH-dependent diaphoras e (d) histochemistry and nitric oxide synthase (NOS) immunocytochemist ry in the brain was investigated by comparing the distribution of thes e enzymes in in situ nitrocellulose blots of unfixed brain sections wi th that in aldehyde-fixed brain sections. Substitution of NADPH by NAD H yielded no gross differences in cellular distribution in the native blot, whereas in fixed sections NADH produced nonspecific staining of the entire section. In the in situ blot NADPHd histochemistry therefor e visualized general nitroblue tetrazolium reductase (NBTr) activity, which was particularly strong in hippocampal pyramidal neurons and cer ebellar Purkinje cells. Aldehyde fixation abolished the anatomical pat tern of general NBTr activity and changed the histochemical distributi on in that of the NADPHd activity associated with the distribution of NOS-I immunoreactivity (ir). Fixation intensified NADPHd histochemical staining in specific neurons, resulting in outstanding, Golgi-like st aining of these neurons in several brain regions, whereas the general NBTr activity in pyramidal and Purkinje cells disappeared. In contrast to the histochemical diaphorase distribution, the distribution of NOS -I ir on blots and in aldehyde-fixed brain sections was similar. No NO S was observed in hippocampal pyramidal and cerebellar Purkinje neuron s, In regions like cerebral and cerebellar cortex and striatum the app lied anti NOS-I serum had a higher affinity for the native protein. It is concluded that aldehydes, rather than to progressively suppress NO S-unrelated enzymes, differentially elicit NADPHd activity in some gro ups of neurons while leaving NOS-ir unaffected.