Researchers use ultrasound to precisely and safely activate brain cells in mice

SAN DIEGO — What if you could turn any neuron in the brain on or off whenever you wanted, and for however long you wanted?

Researchers say that precise, targeted control of the brain’s circuits could be the key to treating everything from epilepsy to Parkinson’s disease to depression. And they’re already using a variety of tools to work toward that goal, from light to electricity to magnets. Some of these approaches are already being used in patients.

A new study from scientists at the Salk Institute suggests there’s another way to stimulate neurons that would be less invasive than current methods while reaching regions deep within the brain — ultrasound.

Researchers discovered a protein that made lab-grown cells that wouldn’t react to ultrasound suddenly responsive. When they engineered mice to make that same protein in a population of neurons that control movement, scientists found that ultrasound triggered spikes of electrical activity in the animals’ limbs — and a few small kicks, too.

The study, published in the journal Nature Communications, is the latest development in a small but growing field known as sonogenetics. For now, researchers still have much to do before these tools can be used in people — from figuring out how to deliver genes encoding ultrasound-sensitive proteins to understanding how this all works. But they’re learning quickly.

“It’s a very exciting contribution and an important step,” said Mikhail Shapiro of Caltech, a sonogenetics expert who was not involved in the study. “This is one of the papers that’s come out over the last several years that shows that it’s a real possibility that you can use ultrasound to directly modulate the activity of specific neurons.”

Sonogenetics has a lot of catching up to do compared to other, more established fields. Researchers have long known, for instance, that cells can be activated with flashes of light if they carry certain light-sensitive proteins, a technique known as optogenetics. They have used this approach to make mice stop and swerve or even hallucinate. And in May 2021, researchers reported that they’d managed to use optogenetics to help a blind person see.

Scientists are also using controlled electrical zaps to stimulate neurons, in some cases by implanting electrodes in the brain or with magnetic fields, the latter strategy known as transcranial magnetic stimulation. Patients with depression, Parkinson’s, and epilepsy, among other disorders, are already being treated with these approaches.

But they have their limitations, says neurobiologist Sreekanth Chalasani, senior author of the recent study. Light can’t travel very far through tissues, which makes it hard to use optogenetics unless you’re targeting nerve cells in an organ that is clear (like the eye).

Electrical stimulation, while effective in stimulating deeper parts of the brain, requires surgical implants. And while transcranial magnetic stimulation isn’t invasive, magnetic fields quickly weaken as they move through the brain.

Enter ultrasound. These high-frequency sound waves are widely used in medicine to peer deep inside the body and do everything from breaking up kidney stones to scorching tumors to glimpsing developing fetuses.

“Everybody gets ultrasounds; things turn out to be OK,” Chalasani said. “Can we find a protein that senses this?”

The answer, his team reported in a 2015 study, was “yes,” based on experiments in Caenorhabditis elegans — a teeny, transparent nematode about the length of a pencil point. Researchers found pulsing these worms with ultrasound caused them to wriggle away. But worms with a mutation in a particular protein didn’t react, a telltale sign the molecule was indispensable for the response.

In their latest study, Chalasani’s team hunted for a protein that would have similar effects in mammalian cells. Researchers spent about a year and a half testing nearly 200 genes encoding proteins known to respond to pressure for signs they also responded to a 7-megahertz burst of ultrasound. Any cells that did would light up green under the microscope.

About five months into this effort, researchers still hadn’t found any promising candidates. That’s when Chalasani, who was in his office, heard two members of his team screaming one night. He went over to see what all the fuss was about, and that’s when he saw it.

“It wasn’t an occasional thing,” Chalasani said of the microscope images. “This was just like a blazing field of light.”

That field of light was the result of cells producing TRPA1, a protein researchers believe is usually activated by chemical irritants and toxins. When the authors coaxed neurons isolated from mice into producing the protein, these cells started firing off electrical signals in response to ultrasound.

To see whether those findings held up in living animals, scientists used a mouse strain that only produced TRPA1 in a certain group of cortical motor neurons, cells that control voluntary muscle movements. Pulsing these mice with ultrasound led to spikes of electrical activity in their limbs and small but visible movements.

Chalasani’s team is now tweaking the protein to get stronger responses. And while turning neurons on is useful, he says, it’s equally important to be able to turn cells off with ultrasound. He thinks his lab may have found a protein that does just that, a molecule made by the carnivorous Venus flytrap plant, but it’ll take careful follow-up work to show whether that’s true.

Then there’s the question of how you’d deliver genes for ultrasound-sensitive proteins to specific neurons. In this study, researchers injected mice directly into the brain with a virus that delivered the gene. But that negates one of ultrasound’s main benefits — noninvasive use. Caltech’s Shapiro thinks a possible workaround could be to use ultrasound to temporarily weaken the blood brain barrier, which could let viral vectors through that would otherwise be unable to reach the brain.

Even if researchers don’t figure out a way to deliver ultrasound-sensitive proteins to cells in the brain, they might learn how to stimulate proteins that neurons are already equipped with, said Colleen Hanlon, a biologist at Wake Forest School of Medicine. She’s one of the leaders in transcranial magnetic stimulation, but she welcomes other approaches aimed at regulating how neurons communicate with one another.

“We’ve spent so much time over the last few decades focusing on pharmacologic therapies,” she said. “This paper is another really important piece to this puzzle of developing neural circuit-based therapeutics for disease.”