Excitotoxicity has been implicated in neurological disorders. This study investigated the neuroprotective effect of the extract from Rhizoma Atractylodis macrocephalae on excitotoxicity‐induced neuronal apoptosis in primary cultured cerebral cortical neurons. Excitotoxicity was induced by exposure of cortical neurons to glutamate. Neuronal apoptosis and the protective effect of Rhizoma Atractylodis macrocephalae extract were examined by multi‐indices including cell viability assay, morphological features, DNA fragmentation and flow cytometric analysis. After exposure of cultured neurons to glutamate for 24 h, the neurons exhibited marked apoptotic‐like death. Co‐treatment of the neurons with glutamate and Rhizoma Atractylodis macrocephalae extract significantly elevated the cell viability, and reduced the number of apoptotic cells. These results demonstrate that Rhizoma Atractylodis macrocephalae is an effective neuroprotective agent against glutamate‐induced excitotoxicity and may have therapeutic potential in excitotoxicity‐mediated diseases. Copyright © 2011 John Wiley & Sons, Ltd.