Dextromethorphan (DM) is a dextrorotatory isomer of levorphanol, a typical morphine-like opioid. When administered at supra-antitussive doses, DM produces psychotoxic and neurotoxic effects in humans. Although DM abuse has been well-documented, few studies have examined the effects of high-dose DM. The present study aimed to explore the effects of a single high dose of DM on mortality and seizure occurrence. After intraperitoneal administration with a high dose of DM (80 mg/kg), Sprague–Dawley rats showed increased seizure occurrence and intensity. Hippocampal expression levels of N-methyl-D-aspartate (NMDA) receptor subunits (GluN1 < GluN2A < GluN2B), c-Fos and pro-apoptotic factors (Bax and cleaved caspase-3) were upregulated by DM treatment; while levels of anti-apoptotic factors (Bcl-2 and Bcl-xL) were downregulated. Consistently, DM also induced ultrastructural degeneration in the hippocampus. A non-competitive NMDA receptor antagonist, MK-801, attenuated these effects of high-dose DM, whereas an opioid antagonist, naloxone, did not affect DM-induced neurotoxicity. Moreover, pretreatment with a highly specific GluN2B subunit inhibitor, traxoprodil, was selectively effective in preventing DM-induced c-Fos expression and apoptotic changes. These results suggest that high-dose DM produces convulsive behaviors by activating GluN2B/NMDA signaling that leads to pro-apoptotic changes.