Immunocytochemical localization and developmental profile of neuron specific enolase (NSE) and non-neuronal enolase (NNE) in aggregating cell cultures of fetal rat brain.

In aggregating cultures, neuron specific enolase (NSE) was first detected biochemically at 3 days. NSE levels increased with time in aggregate cultures and at 48 days reached a level which was 33% of that found in adult rat brain in vivo. The level of non-neuronal enolase (NNE) was essentially identical in aggregate cultures and normal rat brain. Immunocytochemically, NSE(+) cells first appeared at 10 days in vitro. Their number increased until 20 days in culture and then remained constant. When the immunocytochemical localization of NSE and NNE was compared in vibratome sections of 25 day aggregates, all identifiable neurons were NSE(+), NNE(-) and glial cells were NSE(-), NNE(+). In 1 micron thick epon sections of 30 day aggregates NSE antiserum stained neuronal cytoplasm intensely. Comparison of NSE staining in 1 micron thick epon sections with the same cell in an adjacently cut thin section provided conclusive evidence that NSE(+) cells were neurons and NSE(-) cells were glia. These results demonstrate that the three-dimensional organization of aggregate cells provides an excellent environment for neuronal differentiation and also emphasize the advantages of this culture system for multidisciplinary studies of brain development.