Nitric oxide synthase in the brain: light and electron microscopical findings based on the NADPH-diaphorase reaction

The recent discovery of the identify of nitric oxide synthase with the reduced nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) has powerfully stimulated the anatomical localization of sites of nitric oxide synthesis in the nervous system. In the present study the widely used light microscopical technique for NADPH-d staining was adapted to the electron microscopical level by applying the tetrazolium salt 2-(2'-benzothiazolyl)-5-styryl-3-(4'-phthalhydrazidyl)tetrazolium chloride (BSPT) which produces an electron-dense reaction product, BSPT-formazan. Predominantly membranes of the endoplasmic reticulum were stained. Apart from singular heavily labeled neurons, a majority of nerve cells, light microscopically "unstained", shows sporadically formazan deposits, and, likewise, but regionally different, a few astroglial cells. Lesions induced by the glutamate agonists quinolinic acid and alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) display surviving neurons, which are predominantly stained for NADPH-d. Astroglial cells within lesioned areas exhibit increased amounts of reaction product, apparently as a consequence of enzyme induction.