INDUCIBLE NITRIC-OXIDE SYNTHASE ACTIVITY OF CLONED MURINE MICROGLIAL CELLS

Nitric oxide (NO) is a short-lived diffusable molecule now believed to participate in multiple physiologic functions in the CNS including neurotransmission and the maintenance of vascular tone. Previously, we reported that cell lines obtained by retroviral immortalization of tissue macrophages (Mphi) could be induced to synthesize nitrite (NO2-), a stable end product of the NO synthetic pathway. We have further characterized the induction and activity of this pathway in a panel of seven microglial clones derived from primary embryonic mouse brain cultures. Like Mphi, these clones were found to release high levels of NO2- in response to recombinant interferon-gamma (rIFN-gamma) as a priming signal together with either bacterial lipopolysaccharide (LPS) or exogenous recombinant tumor necrosis factor-alpha (rTNF-alpha). As previously demonstrated for Mphi, phagocytosis of zymosan particles during induction of enzyme activity enhanced subsequent NO2- production, which is of interest in light of the postulated phagocytic role of microglia within the CNS. Biochemical characterization of enzyme activity in intact microglial clones and in isolated cytosolic fractions indicates that the microglial NO synthase present in these murine cell clones represents the Mphi-like isotype. These findings suggest that microglial cells could represent a major source of NO within the CNS.