Neurotrophin-3 (NT-3) is one of the most potent stimulators for survival of auditory sensory neurons. Viral transfer of neurotrophins into auditory neurons may offer a route to provide a permanent supply of the growth factor and guarantee their long-term survival. Herpes simplex virus type 1 (HSV-1)-based vectors have demonstrated their effectiveness to transfer genes into peripheral sensory neurons. In the present report, we have produced a HSV-1-based amplicon vector expressing NT-3. This vector efficiently infects isolated auditory neurons and stimulates their survival during distinct developmental stages of the inner ear. Therefore, this vector may present a unique entry point to develop therapies preventing or treating hearing impairment caused by the degeneration of auditory neurons.