A light gray mouse against an orange background listens to tiny headphones
Researchers restored hearing in mice with a gene therapy delivered via the cerebrospinal fluid.
© Adobe Stock, Margo_Alexa

To hear a sound, auditory signals travel through a person’s ear, where they are transformed into vibrations that swirl through fluid in the cochlea, a spiral, snail-like structure in the inner ear. This fluid movement pushes on hair cells that turn the physical sensations into electrical signals, which stimulate the brain. While elegant, this complex sensory system can break down, leading to hearing impairment in over a billion people worldwide.1

Hearing loss is often due to dysfunction or death of the cochlear cells and interacting neurons from environmental and/or genetic factors.2 While medical devices help restore auditory function, they cannot completely restore hearing; hearing aids merely amplify sound waves, while people with cochlear implants struggle to understand speech against background noise and fully appreciate music.Researcher have explored gene therapies to permanently reverse genetic-based hearing loss and have seen success in neonatal mouse models.Maiken Nedergaard, a professor and the co-director of the Center for Translational Neuromedicine at both the University of Rochester and the University of Copenhagen, and Barbara Canlon, a professor of hearing physiology at the Karolinska Institute, discovered a novel gene therapy delivery method that sends viral vectors to the inner ear of adult mice via the cerebrospinal fluid (CSF).

Nedergaard is well-known for discovering the glymphatic system—the network by which the central nervous system clears itself of waste proteins through CSF transport.6 She wondered if CSF could reach the inner ear through this system. By injecting mice with contrast agent in the cisterna magna—a CSF-filled space at the skull’s base—and performing magnetic resonance imaging (MRI), Nedergaard found that the fluid dispersed via a structure called the cochlear aqueduct rather than the traditional glymphatic transport channels. “Within minutes, you see that if you inject any contrast agent, it enters the inner ear,” said Nedergaard. 

Nedergaard shared her finding with Canlon, who became excited by the prospect of using this route to deliver functional copies of genes responsible for genetic hearing loss. While researchers have recently delivered gene therapy by injection to the inner ear,7 Nedergaard and Canlon saw promise in using this alternative, and likely safer, method to bring viral vectors in contact with the cochlear cells that need repairing. 

An MRI image of the mouse brain, taken from below, showing the inner ear in green, arteries in pink, and ventricles in blue.
Magnetic resonance imaging of the mouse brain from below shows the inner ear (green), CSF-filled ventricles (blue), and arteries (pink). The cisterna magna connects to the subarachnoid space, allowing fluid to flow from these structures to the inner ear via the cochlear aqueduct.
Yuki Mori

“The inner ear is almost not accessible, it's embedded deep within very heavy cranial structure,” said Nedergaard. In humans, the inner ears are mature at birth,4 and to effectively deliver therapeutics or cochlear implants, clinicians surgically breech the cochlea, which imparts its own risks. “If you go surgically, you now have to disrupt the cochlea, which is the exact same structure that you're trying to restore,” said Tiffany Peng Hwa, a neurotologist at the University of Pennsylvania who was not involved in this study. “Going through the cerebrospinal fluid, through the spinal cord [in humans], it allows you an access point that is fairly direct…and you don't have to damage the ear.” In humans, such injections could occur via lumbar puncture, which is a quick and routine procedure. 

Nedergaard and Canlon’s research team aimed to restore hearing in mice with a defective vesicular glutamate transporter-3 (VGLUT3) protein. These mice are deaf because their inner ear hair cells do not release enough glutamate to transmit auditory signals to the brain.8 The researchers delivered an adeno-associated viral (AAV) vector containing a functional copy of the gene by injection into the cisterna magna. 

Two weeks later, the researchers placed electrode on the mice’s heads to measure auditory responses. They found that they had restored hearing for all except the highest sound frequencies. The injections also reinstated VGLUT3 expression in inner ear hair cells. “It's basically a proof of principle that you can almost noninvasively cure deafness,” said Nedergaard.

While these results are promising, according to Doug Epstein, who researches the genetic basis of hearing loss at the University of Pennsylvania and was not involved in this study, one limitation is that the viral vector was not specifically targeted to inner ear hair cells. “They did not detect very many cells that were infected outside of the inner ear, but there was no system in place to control the gene expression through the virus other than [its] tropism,” said Epstein. In contrast to delivery in mice, a previous study showed that injecting AAV vectors into the CSF of nonhuman primates caused gene expression in various brain regions and other organs, including the liver and spleen.9 “You don't want to put genes into the wrong cell types and have them be compromised because of that,” said Epstein.

To further develop this technique for reversing hearing loss, the researchers will need to assess its safety profile in nonhuman primates and specify the vector’s target. “The next step would be to develop viruses that can transfect the inner ear without infecting any other tissue,” said Nedergaard. “That would both increase efficiency and have less side transfection.”


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