Neurotrophin-3 (NT-3) and brain-derived neurotrophic factor (BDNF) were recently shown to have biological activity in central neurons. In the present study, NT-3 and BDNF attenuated glucose deprivation-induced neuronal damage dose-dependently in rat hippocampal, septal and cortical cultures. Direct measurements of intraneuronal free calcium levels ([Ca2+]i) and manipulations of calcium influx demonstrated that NT-3 and BDNF each prevented the elevation of [Ca2+]i that mediated glucose deprivation-induced injury. Studies in cultures depleted of glia indicated a direct action of NT-3 and BDNF on neurons. Neurons pretreated with NT-3 or BDNF for 24 hr were more resistant to glutamate neurotoxicity, and showed attenuated [Ca2+]i responses to glutamate. TrkB (BDNF receptor) and trkC (NT-3 receptor) proteins were present in hippocampal, cortical and septal cultures where they were localized to neuronal cell bodies and neurites. The data demonstrate that NT-3 and BDNF can protect neurons against metabolic and excitotoxic insults, and suggest that these neurotrophins may serve [Ca2+]i-stabilizing and neuroprotective functions in the brain.