(Image caption: The hair cells of mice missing just Hey2 are neatly lined up in four rows (left) while those missing Hey1 and Hey2 are disorganized (right). The cells’ hairlike protrusions (pink) can be misoriented, too. Credit: Angelika Doetzlhofer)

Hey1 and Hey2 ensure inner ear ‘hair cells’ are made at the right time, in the right place

Two Johns Hopkins neuroscientists have discovered the “molecular brakes” that time the generation of important cells in the inner ear cochleas of mice. These “hair cells” translate sound waves into electrical signals that are carried to the brain and are interpreted as sounds. If the arrangement of the cells is disordered, hearing is impaired.

A summary of the research will be published in The Journal of Neuroscience on Sept. 16.

"The proteins Hey1 and Hey2 act as brakes to prevent hair cell generation until the time is right," says Angelika Doetzlhofer, Ph.D., an assistant professor of neuroscience. "Without them, the hair cells end up disorganized and dysfunctional."

The cochlea is a coiled, fluid-filled structure bordered by a flexible membrane that vibrates when sound waves hit it. This vibration is passed through the fluid in the cochlea and sensed by specialized hair cells that line the tissue in four precise rows. Their name comes from the cells’ hairlike protrusions that detect movement of the cochlear fluid and create electrical signals that relay the sound to the brain.

During development, “parent cells” within the cochlea gradually differentiate into hair cells in a precise sequence, starting with the cells at the base of the cochlea and progressing toward its tip. The signaling protein Sonic Hedgehog was known to be released by nearby nerve cells in a time- and space-dependent pattern that matches that of hair cell differentiation. But the mechanism of Sonic Hedgehog’s action was unclear.

Doetzlhofer and postdoctoral fellow Ana Benito Gonzalez bred mice whose inner ear cells were missing Hey1 and Hey2, two genes known to be active in the parent cells but turned off in hair cells. They found that, without those genes, the cells were generated too early and were abnormally patterned: Rows of hair cells were either too many or too few, and their hairlike protrusions were often deformed and pointing in the wrong direction.

"While these mice didn’t live long enough for us to test their hearing, we know from other studies that mice with disorganized hair cell patterns have serious hearing problems," says Doetzlhofer.

Further experiments demonstrated the role of Sonic Hedgehog in regulating the two key genes.

"Hey1 and Hey2 stop the parent cells from turning into hair cells until the time is right," explains Doetzlhofer. "Sonic Hedgehog applies those ‘brakes,’ then slowly releases pressure on them as the cochlea develops. If the brakes stop working, the hair cells are generated too early and end up misaligned."

She adds that Sonic Hedgehog, Hey1 and Hey2 are found in many other parent cell types throughout the developing nervous system and may play similar roles in timing the generation of other cell types.

Single tone alerts brain to complete sound pattern

The processing of sound in the brain is more advanced than previously thought. When we hear a tone, our brain temporarily strengthens that tone but also any tones separated from it by one or more octaves. A research team from Utrecht and Nijmegen published an article on the subject in the journal PNAS on 2 September.

We hear with our brain. The cochlea picks up sound vibrations but the signals produced as a result are processed by the brain, using known patterns. If, for example, you briefly hear a weak tone, your hearing focuses on that tone and suppresses any frequencies around it. This makes it easier to notice any relevant sounds in your surroundings. The present research has shown that this ‘auditory attention filter’ is much more complex than believed until now: frequencies that have an octave relationship with the target tone are also heard better.

John van Opstal, professor of Biophysics at Radboud University: ‘This test proves that the brain prepares for a more extensive pattern of tones, even if the person just hears a single test tone or if he has a tone in mind. These extra tones in the pattern were not sounded during the experiment, but the brain complements the information received from the cochlea. This is scientifically interesting. Audiology, for example, at present places great emphasis on the cochlea.’

Octave relationship
The subjects undergoing the experiment did not have an easy time. For an hour they listened to unstructured noise containing very soft tones that they had to detect. Every few seconds they were presented with a tone of 1000 Hz, the cue. Then during one of two time intervals, a very quiet, short second tone was sounded. The subject had to indicate in which of the two intervals they had heard the second tone. It became apparent that tones having an octave relationship with the cue were all heard better, and those around the cue were heard less well. An octave is a well-known term in music, indicating the distance between two tones, the frequencies of which have a 2-to-1 relationship. 

Voice
Van Opstal: ‘We wanted to gather data on the auditory attention filter around the target tone. When we made the range larger than other researchers had done previously, more peaks suddenly appeared. This was a complete surprise to us. One possible explanation could be that the hearing system has evolved in order to hear sounds made by members of an animal’s own species (voices in the case of humans) in noisy surroundings. Vocalisations always consist of harmonic complexes of several simultaneous tones having an octave relationship with each other.’

Hearing aid
The researchers, who work at Utrecht University, the UMC Utrecht Brain Center and Radboud University Nijmegen, can easily think up applications for this fundamental research. If, for example, someone no longer hears high tones because of damage to the cochlear hair cells, the hearing aid can be adjusted in such a way that it converts those tones so they sound one or more octaves lower. Since the brain itself ‘fills in’ tones with an octave relationship, that person’s perception should then become more normal. It is also important for commercial sound producers to know how tones are perceived. That is why Philips Research is involved in this research in their department ‘Brain, Body and Behavior’.

Hearing loss from loud blasts may be treatable

Long-term hearing loss from loud explosions, such as blasts from roadside bombs, may not be as irreversible as previously thought, according to a new study by researchers at the Stanford University School of Medicine.

Using a mouse model, the study found that loud blasts actually cause hair-cell and nerve-cell damage, rather than structural damage, to the cochlea, which is the auditory portion of the inner ear. This could be good news for the millions of soldiers and civilians who, after surviving these often devastating bombs, suffer long-term hearing damage.

“It means we could potentially try to reduce this damage,” said John Oghalai, MD, associate professor of otolaryngology and senior author of the study, published July 1 in PLOS ONE. If the cochlea, an extremely delicate structure, had been shredded and ripped apart by a large blast, as earlier studies have asserted, the damage would be irreversible. (Researchers presume that the damage seen in these previous studies may have been due to the use of older, less sophisticated imaging techniques.)

“The most common issue we see veterans for is hearing loss,” said Oghalai, a scientist and clinician who treats patients at Stanford Hospital & Clinics and directs the hearing center at Lucile Packard Children’s Hospital.

The increasingly common use of improvised explosive devices, or IEDs, around the world provided the impetus for the new study, which was primarily funded by the U.S. Department of Defense. Among veterans with service-connected disabilities, tinnitus — a constant ringing in the ears — is the most prevalent condition. Hearing loss is the second-most-prevalent condition. But the results of the study would prove true for anyone who is exposed to loud blasts from other sources, such as jet engines, air bags or gunfire.

More than 60 percent of wounded-in-action service members have eardrum injuries, tinnitus or hearing loss, or some combination of these, the study says. Twenty-eight percent of all military personnel experience some degree of hearing loss post-deployment. The most devastating effect of blast injury to the ear is permanent hearing loss due to trauma to the cochlea. But exactly how this damage is caused has not been well understood.

The ears are extremely fragile instruments. Sound waves enter the ear, causing the eardrums to vibrate. These vibrations get sent to the cochlea in the inner ear, where fluid carries them to rows of hair cells, which in turn stimulate auditory nerve fibers. These impulses are then sent to the brain via the auditory nerve, where they get interpreted as sounds.

Permanent hearing loss from loud noise begins at about 85 decibels, typical of a hair dryer or a food blender. IEDs have noise levels approaching 170 decibels.

Damage to the eardrum is known to be common after large blasts, but this is easily detected during a clinical exam and usually can heal itself — or is surgically repairable — and is thus not typically the cause of long-term hearing loss.

In order to determine exactly what is causing the permanent hearing loss, Stanford researchers created a mouse model to study the effects of noise blasts on the ear.

After exposing anesthetized mice to loud blasts, researchers examined the inner workings of the mouse ear from the eardrum to the cochlea. The ears were examined from day one through three months. A micro-CT scanner was used to image the workings of the ear after dissection.

“When we looked inside the cochlea, we saw the hair-cell loss and auditory-nerve-cell loss,” Oghalai said.

“With one loud blast, you lose a huge number of these cells. What’s nice is that the hair cells and nerve cells are not immediately gone. The theory now is that if the ear could be treated with certain medications right after the blast, that might limit the damage.”

Previous studies on larger animals had found that the cochlea was torn apart and shredded after exposure to a loud blast. Stanford scientists did not find this in the mouse model and speculate that the use of older research techniques may have caused the damage.

“We found that the blast trauma is similar to what we see from more lower noise exposure over time,” said Oghalai. “We lose the sensory hair cells that convert sound vibrations into electrical signals, and also the auditory nerve cells.”

Much of the resulting hearing loss after such blast damage to the ear is actually caused by the body’s immune response to the injured cells, Oghalai said. The creation of scar tissue to help heal the injury is a particular problem in the ear because the organ needs to vibrate to allow the hearing mechanism to work. Scar tissue damages that ability.

“There is going to be a window where we could stop whatever the body’s inflammatory response would be right after the blast,” Oghalai said. “We might be able to stop the damage. This will determine future research.”

New understanding of hearing loss

A major breakthrough in the understanding of hearing and noise-induced hearing loss has been made by hearing scientists from three Pacific Rim universities.

Scientists from The University of Auckland, the University of New South Wales in Sydney, and the University of California in San Diego have collaborated for nearly 20 years on this research.

“This work represents a paradigm shift in understanding how our ears respond to noise exposure,” says Professor Peter Thorne from The University of Auckland, who is one of the co-authors of two papers published recently in the prestigious journal, the Proceedings of the National Academy of Sciences (PNAS) [1, 2].

“We demonstrate that what we traditionally regard as a temporary hearing loss from noise exposure is in fact the cochlea of the inner ear adapting to the noisy environment, turning itself down in order to be able to detect new signals that appear in the noise,” he says.

After the noise is turned off, hearing remains temporarily dull for some time while it readjusts to the lack of noise.

“Clinically, this is what we measure as a temporary hearing loss,” says Professor Thorne. “This has always been regarded as an indication of noise damage rather than, in our new view, a normal physiological process.”

The researchers show that this is due to a molecular signalling pathway in the cochlea, mediated by a chemical compound called ATP, released by the cochlear tissue with noise and activating specific ATP receptors in the cochlear cells.

“Interestingly, if the pathway is removed, such as by genetic manipulations, this adaptive mechanism doesn’t occur and the ear becomes very vulnerable to longer term noise exposure and the effects of age, eventually resulting in permanent hearing loss.”

“In other words the adaptive mechanism also protects the ear,” says Professor Thorne.

The second paper, done in collaboration with United States colleagues, reveals a new genetic cause of deafness in humans which involves exactly the same mechanism.

People (two families in China) who had a mutation in the ATP receptor showed a rapidly progressing hearing loss which was accelerated if they worked in noisy environments.

“This work is important because it shows that our ears naturally adapt to their environment, a bit like pupils of the eye which dilate or constrict with light, but over a longer time course,” Professor Thorne says.

This inherent adaptive process also provides protection to the ear from noise and age-related wear and tear. If people don’t have the genes that produce this protection, then they are more likely susceptible to developing hearing loss.

“This may go some way to explaining why some people are very vulnerable to noise or develop hearing loss with age and others don’t,” he says.

“Our research demonstrates that what we have always thought was temporary noise damage (i.e. the temporary hearing loss experienced in night clubs or a day’s work in factories), may not be this, but instead, is the ear regulating its sensitivity in background noise”.

“Although our research suggests that our hearing adapts in some noise environments, this has limits,” says Professor Thorne. “If we exceed the safe dose of noise, our ears can still be damaged permanently despite this apparent protective mechanism.”

“People need to protect their ears from constant noise exposure to prevent hearing loss and this is particularly important in the workplace and with personal music devices which can deliver high sound levels for long periods of time,” he says.

Deep inside a mouse’s ear, a swirling galaxy of cells

Is this a churning galaxy in some faraway corner of the universe? A neon rose plucked by a 1990s raver? Or just a dollop of fluorescent paint swirling down the drain? Nope - it’s the cochlea of a mouse that has been stained with antibodies to reveal cells with different functions.

The image, created by Karen Avraham and Shaked Shivatzki of Tel Aviv University in Israel, was the winning entry in the GenArt 2012 human genetics image competition.

Overlaid on the twisting cochlea is a cascade of green letters that make up the DNA sequence of connexin 26. Mutations in this gene are the most common cause for deafness, says Avraham. The image is an artistic representation of deep sequencing, a technique for detecting variances in DNA.

Avraham says deep sequencing is revolutionising the hunt for genetic mutations because of its speed and low cost. Where sequencing a genome once cost millions of dollars and took years, it now takes weeks and costs about $1000.

"By finding the mutations responsible for human disease, scientists can diagnose disorders in a way that was impossible before," she says.