Microinjection through the round window membrane has been found to represent a method for vector delivery in intracochlear gene transfer in animal models but breaches the round window membrane, making it necessary to evaluate animals for possible postinjection hearing loss. In the present study healthy guinea pigs were evaluated for their baseline click auditory brainstem response (ABR) thresholds. Each animal was then injected with saline via the round window membrane. After 1 week auditory function was evaluated by click ABR. Animals with increased ABR thresholds were retested at 4 weeks. Animals with 1-week postoperative ABRs similar to baseline were not retested. Results showed that postoperative ABR thresholds in five animals (71%) remained unchanged from baseline, while two animals had increases of 20-25 dB in ABRs after 1 week but recovered baseline ABRs after 4 weeks. The mean baseline ABR threshold was 25.7 dB and was 27.9 dB after 1 week after injection. The difference between preoperative and 1-week postoperative averages was not significant (P = 0.707). In this preliminary study saline microinjection through the round window membrane did not cause permanent hearing loss in the guinea pigs tested, and any damage caused by microinjection appeared to be reversible.