Brain, due to its exceeding complexity, is the most important but least understood organ. Existing methods of mammalian brain imaging either involve sectioning followed by reconstruction or are not compatible with molecular phenotyping. However, without understanding it as a whole, partial functions of brain remain open-ended quest. And only obtaining high-resolution structural information of brain together with access of functional study would bring a global view of how brain works. In the spring of 2013, Karl Deisseroth and colleagues addressed this challenge with a technique which they called CLARITY (Clear, Lipid-exchanged, Anatomically Rigid, Imaging/immunostaining compatible, TissuehYdrogel).
Fig. 1: Stanford researchers developed a method called CLARITY, making brain tissue transparent and permeable for macromolecules. It allows the 3D-imaging and 3D molecular analysis of brain.
The main idea of CLARITY is to remove the lipidwhich scatters the light and make the brain completely transparent for the detailed microscopic investigation. To preserve the structure of brain tissue while removing the lipid components, the first step is to inject formaldehyde and hydrogel. The formaldehyde cross-link all the molecules (proteins and amino acids) except lipid to hydrogel monomers, then polymerize into a hydrogel mesh when heated to body temperature. The lipid, then, can be cleaned by electrophoresis, leaving the fine-structure and cross-linked biological molecules in place.
After CLARITY, the brain tissue becomes not only transparent but also theoretically permeable for macromolecules. Karl Deisseroth and colleagues further proved this by using fluorescent antibodies which only attach to specific proteins and showed that only specific structures in the brain could be lighted up under illumination. This opens a new door for brain study, for instance, researchers can trace neural circuits through the entire brain, and go deeply into the brain to investigate the local wiring. Moreover, intracellularly, researchers can study the subcellular structures and interplay between molecules.
Fig. 2: A three-dimensionalimage of a CLARITY-processed adult mouse hippocampus (1mm). Neurons are labeled by different antibodies, excitatory neurons in green, inhibitory neurons in red, and astrocytes in blue.
We may be surprised how this simple method will improve the brain study in the future. Sometimes, a good idea could be the biggest step in the research. And the great ideas always come from the flexible application of multidisciplinary knowledge in the study.
Chung, Kwanghun, Jenelle Wallace, Sung-Yon Kim, SandhiyaKalyanasundaram, Aaron S. Andalman, Thomas J. Davidson, Julie J. Mirzabekov et al. "Structural and molecular interrogation of intact biological systems." Nature 497, no. 7449 (2013): 332-337.