Ultrastructural demonstration of dehydrogenases in rat cerebral cortex

Techniques for the ultrastructural demonstration of dehydrogenases in cerebral cortex are described. The best fixation for good fine structural preservation and retention of LDH and NADH-diphorase was obtained by perfusion with a misture of formaldehyde and glutaraldehyde and for SDH by perfusion with formaldehyde. Comparison of incubation conditions showed that consistent results were obtained using enzyme markers NBT and DS-NBT for LDH and NADH-diaphorase: DS-NBT was more satisfactory than NBT and BSPT for SDH. Penetration of incubation media was improved by Triton X-100: DMSO and ultrasonic treatment were less effective. The techniques enabled the first electron cytochemical demonstration of dehydrogenases in different elements of prefixed cerebral cortex. Ultrastructural sites of enzyme activities were localized within cristae and inter-membrane spaces of mitochondria in nerve cell cytoplasm and its processes, oligodendrocytes and astrocytes. Authenticity of the ultrastructural sites was confirmed by four different control experiments.