In 1906, the Nobel Prize in Medicine was shared between Camilo Golgi and Santiago Ramón y Cajal. Golgi was largely awarded for creating a type of staining that marked neurons and their ramifications, but this one had the problem that because there were so many neurons it was very difficult to see where each neuron began and ended.

Cajal altered the staining to make many fewer neurons distinguishable, which made it possible to see each cell individually and discover that in the communication between neurons there are spaces of separation.

The Brainbow

Thus, from the beginning of histology it has been a constant problem to distinguish neurons and their extensions within a dye. And it is not for less, since our brain contains more than 100,000 kilometers of connections, but fortunately in 2007 was published the first version of a technique that allows us to see individually each one of these connections: the Brainbow.

The concepts of the Brainbow

This technique is based on two very simple concepts:

1. It is possible to generate transgenic animals that express fluorescent proteins of different colors. For example, there is Green Fluorescent Protein (GFP) or Red Fluorescent Protein (RFP).
2. If each cell of an animal tissue has different types of these fluorescent proteins and in different amounts, then each cell will have a different color.

From this base, animals were generated (initially worked with mice) that had three or four different fluorescent proteins that were mixed thanks to the activation of a gene (a recombinase also artificially introduced into the genome), which randomly mixes the amounts and types of fluorescent proteins in each cell.

The result is that approximately 100 different color combinations are obtained. Here the key is that this color change is permanent and marks the entire cell, including its ramifications. As this happens in the neurons of the brain, it allows a detailed study of what is known as the connectome, (the set of connections between neurons) and already makes it easier to analyze under the microscope where each neuron connects. However, an important amount of bioinformatics and electron microscopy work is required to handle such a large volume of connections. But the study of the connector deserves it since it is the connections that allow the passage of information between neuronal bodies and are the ones that really explain how the brain works each time it executes a task.

El Brainbow in other organisms

The importance of the Brainbow is such that it has been imported into other organisms where genetic manipulation is possible. An example is Drosophila melanogaster, the fruit fly, where there is intense neuroscientific research both at a basic level studying brain development, and at a biomedical level with works that use Drosophila for research on Parkinson or Alzheimer.

And another example is the Dario rerio or zebrafish, which thanks to the fact that it is transparent during the first phases of its development is used for the study of the initial formation of the nervous system of vertebrates.

Furthermore, it should not be forgotten that although Brainbow was originally a technique developed for neuroscientific studies, it is possible to adapt this tool to other types of tissues.

Brainbow and Cell Biology

With all this, the Brainbow has become one of the most popular tools in cell biology. It still remains an important tool in the study of the connectome, which is undoubtedly the great neuroscientific challenge of the 21st century.

But it can also be used in other tasks such as the study of cellular lineages: once the technique is activated, the change of color in neurons is permanent and also hereditary. In other words, if that cell divides, its daughter cells will maintain the color of its progenitor cell. This makes it possible to study precisely which cells give rise to which tissues, thus helping to understand the field of neuronal stem cells in greater depth.

The Brainbow is not only one of the most eye-catching techniques in molecular biology, it is also one of the most versatile.

References

• Benjamin Richier and Iris Salecker. Versatile genetic paintbrushes: Brainbow technologies. WIREs Developmental Biology (2015). Volume 4.
• Dawen Cai, Kimberly B. Cohen, Tuanlian Luo, Jeff W. Lichtman, and Joshua R. Sanes. New tools for the brainbow toolbox. Nat Methods (2013). 10(6): 540–547.
• Jean Livet, Tamily A. Weissman, Hyuno Kang, Ryan W. Draft, Ju Lu, Robyn A. Bennis, Joshua R. Sanes & Jeff W. Lichtman. Transgenic strategies for combinatorial expression of fluorescent proteins in thenervous system. Nature (2007). Vol 450.
• Michel A. Hofman. Evolution of the human brain: when bigger is better. Frontiers in Neuroanatomy (2014). Volume 8, Article 15.
• Zoe T. Cook, Nicole L. Brockway, Zachary J. C. Tobias, Joy Pajarla, Isaac S. Boardman, Helen Ippolito, Sylvia Nkombo Nkoula, and Tamily A. Weissman. Combining near-infrared fluorescence with Brainbow to visualize expression of specific genes within a multicolor context. Molecular Biology of the Cell (2019). Volume 30