For a long time, scientists have been mystified by how horses are able to produce their unique whinny. The vocalization includes both high-pitched and low-pitched noises, a phenomenon known as biphonation. No other animal can do this simultaneously.

Now, researchers think they have finally figured out how a horse's distinctive whinny can include high- and low-pitched components. The research team performed a series of tests, scans, and experiments on horse larynxes.

They discovered that the low-pitched part of a horse's whinny is made by vibrating the vocal cords, which is similar to human speech and singing. The high-pitched part is produced by whistling with its voice box, also known as a larynx.

A horse has to contract the muscles around its larynx and force air through a small opening to make the whistling sound. Mice and rats can also do this, but their vocalizations are too high-pitched for human ears to detect.

Around 4,200 years ago, humans tamed the ancestors of modern domestic horses, but we still don't fully understand their communication.

It wasn't until 2015 that scientists realized a horse's whinny contains a high-pitched sound and a low-pitched sound, both produced at the same time.

They were surprised to learn that horses could make high-frequency sounds because the general rule of thumb is that large animals make lower-pitched sounds while small animals make higher-pitched sounds.

For the new study, the research team used an endoscopic camera to get a closer look at live horses' larynxes as they made sounds. Their vocal cords vibrated when making lower-pitched sounds but stayed still when producing higher-pitched ones.

However, the muscles above the vocal cords strongly contracted during higher-pitched sounds.

Red horse standing in meadowkwadrat70 - stock.adobe.com - illustrative purposes only

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Then, the researchers blew compressed air and helium through larynxes from a horse meat supplier. The larynxes had been removed from dead horses.

The team compared the frequencies of the sounds produced by the two gases, so they could test where the high-frequency sound was coming from.

"The low-frequency component is produced by vocal fold vibration, just like human singing or a cow mooing, and it didn't shift when we changed between gases," said Tecumseh Fitch from the University of Vienna, Austria.

"But the high-frequency component does significantly shift to be higher in helium, just as predicted for whistles."

The experiment confirmed that the high-frequency sound was a whistle. It's still unclear exactly how horses control the whistle sound, though, or why they evolved to have this vocalization in the first place.

According to Elodie Mandel-Briefer, a co-author of the study and a biologist at the University of Copenhagen in Denmark, the high pitch is possibly for long-distance communication between horses, while the low pitch is for short distances.

The findings were published in Current Biology.