The course of putrescine immunocytochemical appearance in neurons, astroglia and microglia in rat brain cultures.

Putrescine, the diamine precursor for polyamine biosynthesis, is a ubiquitous molecule normally present at low concentration in quiescent cells. During development, or after traumatic stress, putrescine concentrations are greatly increased. Here we describe the localization of putrescine by fluorescence immunocytochemistry in primary cultures of embryonic rat brain using specific antibodies. Antibodies against putrescine conjugated to keyhole limpet hemocyanin (KLH) were produced in rabbits. The antisera were adsorbed on KLH affinity columns and the specificity of the antibodies was assessed by inhibition enzyme-linked immunoassays (ELISA). The cellular localization paralleled the temporal sequence of appearance and disappearance of the different cell types in these mixed cultures. During the first 3 days after plating the antibodies were localized mainly in neurons. As the neurons disappeared the localization was mainly in the growing astroglia, and then, as astroglia reached confluence between 10 and 14 days in vitro, labeled astroglia were diminished in numbers while the number of labeled microglia was greatly increased. The subcellular localization was prominent in the perinuclear region of the cytoplasm. The results indicate that antibodies to KLH-conjugated putrescine can be used for immunocytochemical studies of changes in putrescine concentrations during development and after traumatic injuries.