A great deal of our understanding of nitric oxide (NO) functioning as a unique biological messenger has derived from studies of its biosynthetic enzyme nitric oxide synthase (NOS). 1,2 Three distinct isoforms of NOS have been cloned: neuronal NOS (nNOS), inducible NOS, and endothelial NOS. nNOS was first purified from rat and porcine cerebellum. 3'4 Its primary amino acid sequence is broadly divided into a reductase domain at the COOH terminus, an oxidative domain at the NH2 terminus, and several cofactor recognition domains, including a nicotinamide adenine dinucleotide phosphate (NADPH)-binding site. 5 The enzyme is found constitutively in discrete groups of neurons. 6 To delineate the distribution of nNOS-containing neurons, it is imperative that there are sensitive and reliable detection techniques for the visualization of nNOS.

This article describes methods that have been successfully applied specifically for nNOS. With appropriate modifications, these techniques can be used for the visualizations of other isoforms of NOS.