This article presents a unique approach for the delivery of gene therapy vectors into the cochlea of the laboratory rat. Mice and guinea pigs are established in vivo models for cochlear gene therapy each of which has distinct advantages and disadvantages. The rat has some of the molecular advantages of a mouse model combined with size advantages for surgical approaches. Vector delivery via cochleostomy or injection through the round window causes concomitant sensorineural hearing loss and is therefore not suitable for studies where the change in hearing is being followed. Compared to the mouse, the rat does not demonstrate easily recognizable landmarks that allow for use of the semicircular canal as an approach to the inner ear. We analyzed sagittal and coronal temporal bone sections of Long Evans rats and identified the bony entrance of the facial nerve as a crucial landmark for canalostomy. Auditory brainstem response and distortion product otoacustic emission measurements revealed minimal differences in the hearing threshold after adenovirus vector application when large volumes of vector were infused to the inner ear. Canalostomy and infusion of adenoviral vectors also resulted in temporary balance disturbance in the rat. Immunohistochemical assessment after delivery of a green fluorescent protein expressing vector showed significant GFP expression in the cochlea. Anat Rec, 2012. © 2012 Wiley Periodicals, Inc.