Time course of in vitro expression of NADPH-diaphorase in cultured rat brain neurons: comparison with in vivo expression

Abstract

The time course of NADPH-diaphorase expression was examined in primary cultures of rat central nervous system and in embryonic or neonatal rat brains using a histochemical method. In cerebral and brainstem cultures from 17-day-old embryonic rats, neuronal cells moderately expressing NADPH-diaphorase were first detected on about the 5th to 7th day of culturing. Both the density of positive cells and the staining intensity increased with age of cultures. The density of positive cells, calculated as a percent of the total number of cells, increased up to day 21 in cultures from both the cerebrum and the brainstem, indicating that NADPH-diaphorase is preferentially expressed in neurons with longer viability. On the other hand, virtually no intensely positive cells were detectable in cerebellar cultures at any period examined up to 21 days. In the in vivo study, moderately stained NADPH-diaphorase-positive neurons were first detected, mainly in the laterodorsal-pedunculopontine tegmental nuclei complex and partly in the striatum, in 16-day-old embryonic rat brain. At 2 days postnatal, intensely stained neurons were detectable in the cerebral cortex as well as in the tegmental nuclei complex and the striatum, indicating some delay in the in vitro, as compared to the in vivo, expression of neuronal NADPH-diaphorase.