Hyperoside (Hyp) is a flavonoid compound isolated from a folk remedy, Rhododendron ponticum L. leaves. It has been shown to have neuroprotective effects both in vivo and in vitro. However, little is known about the effects of Hyp on the neuronal apoptosis induced by glutamate. The present study showed that Hyp significantly attenuated, in a concentration-dependent manner, the apoptosis induced by the exposure of cultured neurons to NMDA. Western blot analysis revealed that Hyp antagonized the expression of excess NR2B-containing NMDA receptors; however, it had no effect on the expression of NR2A-containing NMDA receptors. Our results demonstrate that the neuroprotective effect of Hyp owes, at least partially, to its differential modulation of NR2A- and NR2B-containing NMDA receptors.