NF-kappa B is found in many neuronal cell types in different states of activity. This study aimed to define which conditions induce constitutive NF-kappa B activity in cultured hippocampal neurons using activity-specific antibody staining. In co-culture with astroglia, hippocampal neurons were devoid of activated NF-kappa B. In these co-cultures, NF-kappa B could not be activated via kainate or glutamate. In contrast, separating neurons from the glial compartment resulted in a time-dependent increase of activated neuronal NF-kappa B. In this line, activation of NF-kappa B by kainate or glutamate is very effective in freshly separated cultures, but inhibited when the cultures are reassembled after stimulation. These findings suggests that a neuronal-glial interaction may regulate gene expression via NF-kappa B.