Canalostomy as a Surgical Approach for Cochlear Gene Therapy in the Rat

Abstract

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.

Cochlear gene therapy with ancestral AAV in adult mice: complete transduction of inner hair cells without cochlear dysfunction

“…Others have injected GFP-expressing AAV vectors into the normal adult ear, as in the present study, to evaluate the effectiveness of different delivery routes or viral serotypes in either mice16172024, rat25 or guinea pig38. Although the most successful of these prior studies reports near 100% transfection of IHCs in the base of the cochlea after round window injection20, most report significantly less than that, e.g.…”

“…~30%17 and ~80%38, and all report only 0–20% transduction in the cochlear apex172038. Similarly, although four prior studies have looked at virus delivery via a posterior canal route22232425, none convincingly demonstrated a high targeting efficiency for hair cells throughout the cochlear spiral.…”

“…These approaches include: (1) a cochleostomy into either scala tympani or scala media1718224142, (2) injection through the round window membrane5172043, (3) or injection onto the surface of the round window, with or without manipulations to increase penetration38444546. The approach to the posterior canal differs in that the access is not through the middle ear22232425.…”

“…This approach does not require opening the middle ear, and prior studies have reported minimal threshold shifts2223242547. However, prior studies have reported that transduced cells mainly localize in the vestibular system222324, and the canalostomy approach has not previously been used for cochlear gene therapy.…”

“…In the adult, the sensory epithelia of the cochlear and vestibular systems are encased in the hardest bone in the body, and, as highly sensitive mechanosensors, can be easily damaged by mechanical manipulations such as drilling or fluid injections. The three commonly used routes for drug/virus delivery into the inner ear are 1) via the round window membrane, at the base of the cochlear spiral, where it faces the middle ear space, 2) via a hole drilled through the cochlear capsule into the cochlear fluid spaces near the round window, or 3) through a hole drilled into the posterior semicircular canal where it runs near the external surface of the skull1617182022232425. In our experience in animal models, injection through the semicircular canals is the least traumatic to the cochlea.…”