Stanford Engineering researchers have developed a reversible and noninvasive method to make the scalp of juvenile mice transparent, allowing for repeated imaging of brain development. The new technique involves applying a solution containing ampyrone to the scalp, which temporarily changes the skin’s optical properties so that visible light can pass through. This enables scientists to observe neuron formation and activity in living mice over time.
The study, published August 26 in PNAS, aims to address the difficulty of studying brain changes during childhood and adolescence. Traditional methods do not allow for repeated imaging of neural pathways in the same animal as it grows. The Stanford team’s approach allows researchers to return to the same mouse over days and weeks without surgery.
“This opens a literal window to peek into the brain’s development,” said Guosong Hong, assistant professor of materials science and engineering and senior author on the paper. “Not only can we image the structures of these neurons, but we can also image the neural activity over time in an animal model. In the future, this approach could enable us to look at how these circuits form during the development of an animal.”
Mark Brongersma, Stephen Harris Professor and professor of materials science and engineering and co-author on the paper, explained that differences in optical properties between water and biomaterials cause light scattering in tissue: “From a physics perspective, we’re basically a bag of water with biomaterials,” he said. “And the mismatch in their optical properties is why we can’t see through the skin or scalp.”
By raising the refractive index of water in skin using ampyrone, which absorbs ultraviolet light but lets visible light pass through, researchers made mouse skin transparent for about 20 minutes per application. This allowed them to view fluorescent markers commonly used to track neural activity until mice reached about four weeks old.
“The fact that such fundamental optics laws can be applied and work in a biological system is just amazing to me,” Brongersma said. “It wasn’t clear whether the physics and the chemistry and the biology would all line up to make this happen.”
Jun Ding, associate professor of neurosurgery and co-author on the paper, highlighted challenges with previous methods: “It has been extremely challenging to do this day-by-day imaging in mice that are younger than two weeks because at this age, a mouse’s brain and skull are rapidly growing,” he said. “This is when synapses are formed and pruned during early development…and we’ve had no way to look at how these exciting things change at synapse resolution day by day over longitudinal time.”
The effect wears off unless ampyrone is reapplied, but tests showed no harm from temporary transparency; saline controls caused more irritation than ampyrone itself.
Hong noted ongoing efforts: “We may be able to design and synthesize much more efficient molecules based on ampyrone’s structure, such that we can reduce the concentration needed by another tenfold,” he said.
Brongersma added: “In some sense, the race is on to combine physics, chemistry, and neuroscience to start designing better molecules that can enable entirely new imaging methodology.”
Funding for this research came from several organizations including federal agencies like the National Institutes of Health (NIH) and National Science Foundation (NSF), as well as private foundations such as Rita Allen Foundation and Howard Hughes Medical Institute.
Stanford faculty involved include Guosong Hong (also a Wu Tsai Neurosciences Institute faculty scholar), Mark Brongersma (member of Stanford Bio-X), Jun Ding (member of Stanford Bio-X), Tony Wyss-Coray (D.H. Chen distinguished Professor), Carolyn Bertozzi (Robert M. Bass Professor), Richard Roth (Professor Emeritus), Kerriann Casey and Gordon Wang (clinical associate professors), Xiaoke Chen and Todd Coleman (associate professors), along with several research staff members, postdoctoral scholars, graduate students, undergraduate students from Stanford University as well as collaborators from Sonologi.
For media inquiries contact Jill Wu at Stanford School of Engineering.
