A group of Stanford University researchers have paved the way for increased transparency -- literally and scientifically -- of one of the least understood organs, the brain.
The researchers developed a process that creates a completely intact, transparent mouse brain with three-dimensional views of the brain's fine wiring and molecular structures.
In the world of whole-organ imaging, the new process is a giant step forward.
"CLARITY is a way of making the biological tissue transparent to light, but also at the same time it can be assessed with molecular labels," said Stanford bioengineer and psychiatrist Karl Deisseroth, M.D., Ph.D., who led the team of neuroscientists and chemical engineers that created CLARITY. "What this means is we can look at fine details of what makes up a tissue or an organ without losing sight of the big picture -- the whole organ, the whole biological system itself."
Deisseroth authored a paper published online April 10 in science journal Nature detailing the CLARITY process, which stands for "Clear, Lipid-exchanged, Anatomically, Rigid, Imaging/Immunostaining-compatible, Tissue, hYdrogel," postdoctoral student and first author of the paper Kwanghun Chung, wrote in an email.
Deisseroth is also one of 15 experts on President Barack Obama's $100 million brain research initiative "dream team," which was announced on April 2.
Other team members and paper authors include undergraduate student Jenelle Wallace; graduate students Sung-Yon Kim, Kelly Zalocusky, Joanna Mattis, Aleksandra Denisin and Logan Grosenick; research assistants Sandhiya Kalyanasundaram, Julie Mirzabekov, Sally Pak and Charu Ramakrishnan; postdoctoral scholars Aaron Andalman, and Tom Davidson; former undergraduate student Hannah Bernstein and former staff scientist Viviana Gradinaru.
The discovery is the product of a multidisciplinary effort, with the above team members bringing a range of expertise to the lab: bioengineering, chemical engineering, optics, genetics and neuroscience.
"This couldn't have happened without bringing all these very different fields together," Deisseroth said. "For me, that's really exciting. Not just the medical applications, which for me are very important, but advancing science by bringing different fields together."
Deisseroth's team was working on how to extract lipids, whose composition renders them opaque, while keeping other important features intact and viewable. Lipids are fatty molecules that help the body store energy and serve as the brain's structural component, so removing them causes the rest of the brain tissue to fall apart.
Chung explained in a press release how the team solved the opaque problem.
"We drew upon chemical engineering to transform biological tissue into a new state that is intact but optically transparent and permeable to macromolecules."
This transformation begins by replacing the brain's lipids with a hydrogel that is created from within the brain itself. The researchers then immersed a postmortem mouse brain in the hydrogel and heated it to about body temperature. The heat causes hydrogel monomers to congeal into polymers, becoming a mesh throughout the brain that holds everything together without binding to the lipids.
The team was then able to extract lipids through electrophoresis, a process that separates molecules for analysis.
The end product? A 3-D, completely see-through and intact mouse brain ready to be probed, analyzed and more fully understood.
CLARITY also uses fluorescent antibodies that attach themselves to specific proteins to target certain structures within the brain. Their fluorescence illuminates these structures, creating an image of the brain with certain sections and their compositions clarified with neon light. This allows researchers to gather information about molecules, cell function, relationships between cells and more in a way that is not possible with any other method.
Though Deisseroth's team primarily worked on mouse brains, they have also applied the hydrogel to zebrafish and preserved human brain samples with similar results, holding much promise for future human research.
"CLARITY is applicable to any types of tissue, organs, organisms. For example, we can use this technique to study how blood vessels or cancer cells organized in tumors in 3-D," Chung said.
Deisseroth also said that his team has been approached by people who study Alzheimers and cancer commissions who hope that the process can help further their understanding of how those diseases affect the brain. As a psychiatrist who treats patients with depression, Deisseroth said the process can also be applied to psychiatric research.
"Although it is not compatible with living brains, we have very rare and precious samples that have been donated from human patients and we can study those to come to a deeper understanding of the important structures that are involved."
Though a breakthrough in the world of science and medicine, the process does not come without challenges.
"CLARITY will produce a huge amount of data," Chung said. "How to store, how to process, and how to extract useful information out of the huge dataset will pose a significant challenge."
Deisseroth echoed Chung's sentiments, explaining that the data produced is so vast it cannot be transferred from one computer to another, and it overloads typical fiber networks.
"We have new kinds of problems that the new opportunity has created," he said. "It's exciting, but we have to solve them."