Abstract: The glial-derived neurotrophic protein S100β has been implicated in the development and maintenance of the nervous system. S100β has also been postulated to play a role in mechanisms of neuropathology because of its specific localization and selective overexpression in Alzheimer's disease. However, the exact relationship between S100β overexpression and neurodegeneration is unclear. Recent data have demonstrated that treatment of cultured rat astrocytes with high concentrations of S100β results in a potent activation of inducible nitric oxide synthase (iNOS) and a subsequent generation of nitric oxide (NO), which can lead to astrocytic cell death. To investigate whether S100β-induced NO release from astrocytes might influence neurons, we studied S100β effects on neuroblastoma B104 cells or primary hippocampal neurons co-cultured with astrocytes. We found that S100β treatment of astrocyte-neuron co-cultures resulted in neuronal cell death by both necrosis and apoptosis. Neuronal cell death induced by S100β required the presence of astrocytes and depended on activation of iNOS. Cell death correlated with the levels of NO and was blocked by a specific NOS inhibitor. Our data support the idea that overexpression of S100β may be an exacerbating factor in the neurodegeneration of Alzheimer's disease.