Cochlear hair cells are the sensory receptors of the auditory system. It is well established that antibiotic drugs such as gentamicin can damage hair cells and cause hearing loss. Rescuing hair cells after ototoxic injury is an important issue in hearing recovery. Although many studies have indicated a positive effect of low-level laser therapy (LLLT) on neural cell survival, there has been no study on the effects of LLLT on cochlear hair cells. Therefore, the aim of this study was to elucidate the effects of LLLT on hair cell survival following gentamicin exposure in organotypic cultures of the cochlea of rats. The cochlea cultures were then divided into a control group (n = 8), a laser-only group (n = 8), a gentamicin-only group (n = 8) and a gentamicin plus laser group (n = 7). The control cultures were allowed to grow continuously for 11 days. The laser-only cultures were irradiated with a laser with a wavelength of 810 nm at 8 mW/cm(2) for 60 min per day (0.48 J/cm(2)) for 6 days. The gentamicin groups were exposed to 1 mM gentamicin for 48 h and allowed to recover (gentamicin-only group) or allowed to recover with daily irradiation (gentamicin plus laser group). The hair cells in all groups were stained with FM1-43 and counted every 3 days. The number of hair cells was significantly larger in the gentamicin plus laser group than in the gentamicin-only group. The number of hair cells was larger in the laser-only group than in the control group, but the difference did not reach statistical significance. These results suggest that LLLT may promote hair cell survival following gentamicin damage in the cochlea. This is the first study in the literature that has demonstrated the beneficial effect of LLLT on the recovery of cochlear hair cells.