Cisplatin (CDDP) is a widely used chemotherapeutic agent that is highly ototoxic. Animal studies and clinical trials have shown that thiosulfates can protect against platinum-induced ototoxicity. This study investigated a new model for CDDP ototoxicity in the rat, and tested the potential chemoprotective effect of administering N-acetylcysteine (NAC) before giving CDDP. Long Evans rats were treated with CDDP 6 mg/kg delivered to the aorta via a retrograde right external carotid artery infusion, 15 min after intravenous (IV) infusion of saline (n=8) or NAC 400 mg/kg (n=8), such that the vertebral arteries were perfused. Subsequent groups were similarly treated with NAC 30 min before (n=7) and 4 h after (n=7) CDDP. Auditory brainstem response (ABR) thresholds were tested at 4–20 kHz, 7 days after treatment and compared to baseline ABR values. The NAC-treated rats exhibited no significant change from baseline values at all time intervals, while the saline-treated rats showed marked ototoxicity, especially at higher frequencies. Furthermore, the rats treated with NAC 15 min before CDDP exhibited less overall toxicity to CDDP, as evidenced in weight loss 7 days post-treatment (mean for saline=−39.63 g; mean for NAC=−21.13 g; p=0.0084). These data show that treatment with NAC can prevent CDDP-induced ototoxicity in rats.