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February 29, 2008

Zebrafish provide useful screening tool for genes, compounds that protect against hearing loss

UW Health Sciences/UW Medicine

In a study published in the Feb. 29 issue of the journal PLoS Genetics, researchers at the University of Washington have developed a research method that relies on a zebrafish’s lateral line—the faint line running down each side of a fish that enables it to sense its surroundings—to quickly screen for genes and chemical compounds that protect against hearing loss from some medications.

When people are exposed to some antibiotics and chemotherapy agents, the sensory structures in the inner ear, called hair cells, can be irreversibly damaged, resulting in hearing loss and balance problems. Such medications are called ototoxic. People vary widely in their susceptibility to these agents, as well as to damage caused by other chemical agents, loud sounds and aging.

To find out why this is so, UW researchers Dr. Edwin Rubel, a professor in the departments of otolaryngology, physiology and biophysics, and Dr. David Raible, professor of biological structure, developed a screening strategy that uses hair cells in the lateral line of zebrafish larvae to signal how hair cells in a person’s inner ear might respond under similar conditions. Hair cells are named for small bristly extensions, or stereocilia, jutting from their tops. Movement of fluid, which may be triggered by sound vibrations in the inner ear or changes in water pressure in the fish’s environment, causes the stereocilia to tilt to one side, generating an electrical impulse that travels to the brain.

“One of the pluses about working with zebrafish is that, like other fish, they produce hundreds of offspring. We can look at lots of animals and we can look at many hair cells per animal, which means that we can get good quantitative data,” Raible said.

The researchers first set out to identify genes that may be involved in how hair cells respond to ototoxic medicines. Using a chemical that causes random mutations in zebrafish, the researchers bred various fish families, with each family exhibiting a different set of mutations. They then exposed five-day-old larval offspring to the drug neomycin, a type of antibiotic that damages these hair cells, as well as those in the human inner ear. The larvae were stained to determine if the hair cells were still intact. Fish that were resistant to damage were quickly identified, as were those that were especially vulnerable.

Using genetic techniques, the group then examined the larvae’s DNA, searching for segments that were closely tied to the desired property. They initially focused on five mutations—each located on different genes—that, when inherited from each parent, routinely protected against hair cell damage. Further examination and gene identification of one gene revealed that it corresponds to a gene that is also found in other vertebrates, including humans. Identification of the other four mutations is progressing. Another five mutations were identified that offer protection under more complex genetic conditions.

Next, the researchers investigated whether they could identify chemical compounds that protect hair cells against ototoxic medicines. Using the same screening technique—exposing five-day-old zebrafish larvae to neomycin and later applying special stains to the hair cells—they screened more than 10,000 compounds and narrowed them down to two similar chemicals that provide robust protection of hair cells against the neomycin. One of the compounds was later found to protect hair cells from a mouse’s inner ear against the drug, indicating that the same compound may be protective for other mammals as well.

The authors suggest that their research technique, which combines chemical screening with traditional genetic approaches, offers a fast and efficient way to identify potential drugs and drug targets that may one day provide therapies for people with hearing loss and balance disorders.

The study was funded in part by the National Institute on Deafness and Other Communication Disorders (NIDCD), one of the National Institutes of Health. Other sponsors include the American Academy of Otolaryngology—Head and Neck Surgery Foundation, the University of Washington’s Virginia Merrill Bloedel Hearing Research Center, and the University of Washington Royalty Research Fund.