Published online 6 February 2011 | Nature | doi:10.1038/news.2011.73

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Fly brain structure illuminated

Multicoloured imaging techniques highlight neural networks in fruitflies.

BRAAIIIIIINS!The two techniques for imaging fruitfly neurons, dBrainbow (left) and Flybow (right).

Fly brains have never looked so good. Spectacular images of the insects' complex neural circuitry have now been produced using a pair of techniques that allow individual nerve-cell lineages to be visualized using a range of colours.

Both methods are adaptations of the 'Brainbow' techniques devised at Harvard University in Cambridge, Massachusetts, to visualize mouse neurons, and reported in Nature in 20071.

"We were inspired by the elegance of the Brainbow approach," says Iris Salecker, a neuroscientist at the National Institute for Medical Research in London, who worked on one of the fruitfly methods.

The new techniques, reported in two papers published online today in Nature Methods23, involve inserting strings of genes into the neurons of Drosophila melanogaster embryos. Each gene produces a different fluorescent colour, lighting up individual neurons, or even all of the cells descended from an embryonic neuron - because they will carry the same gene and therefore be the same colour. Both techniques result in colourful visualizations that allow all the nerve cells in any one lineage to be distinguished and their development traced, illuminating how neural circuits develop and interact.

The string includes a selection of colour-producing genes, but only one gene is active in each modified nerve cell — the one closest to a region of DNA called a promoter. As the strings are identical, all the modified neurons would be the same colour, and would be impossible to separate visually.

But the researchers altered the gene strings using enzymes, so that a different colour-producing gene sat next to the promoter in different cells, and individual neurons could then be identified.

Two routes, one destination

The main difference between the two studies lies in the methods used to alter the gene strings.

The first technique, called dBrainbow, was developed by Julie Simpson, a neuroscientist at the Howard Hughes Medical Institute's Janelia Farm Research Campus in Ashburn, Virginia, and her colleagues2. This method uses enzymes called recombinases to randomly delete some of the colour-producing genes from the string, leaving different genes next to the promoter regions in different cells. Individual cells are therefore uniquely coloured and so can be easily distinguished.

Although Simpson's method uses a string of just three colour-producing genes — red, green and blue — the team inserted two strings into the fly neurons, allowing six different hues to be produced.

"It's like a television displaying different colours by mixing red, green and blue," explains Simpson. "Each of the six colours is made by mixing two colours."

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The second technique, called Flybow, was developed by Salecker and her colleagues3. They used an enzyme that 'flips' pairs of colour-producing genes on the string, leaving different genes next to the promoter region. The 'flipping' enzyme is also a recombinase, and so after being inverted, some of the colour-producing genes are randomly deleted. This ensures that all the different genes on the string can potentially end up next to the promoter, and be displayed by individual modified neurons.. Flybow uses a single string of four colours — red, green, blue and yellow.

Cooperation

Simpson and Salecker realized that they were working towards similar goals when they submitted abstracts about their work to the same conference. "We decided to co-submit the work," says Simpson. "People should be more collegial about this kind of thing — it's benefited us both."

Salecker says these techniques could be useful in investigating how individual genes affect neuronal development, because fruitfly genetics are very well charatcterised. "You could knock-out or over-express genes and watch the effects in the nervous system," she says.

"It's gratifying to see these techniques used in animals other than mice," says Jeff Lichtman, a Harvard neuroscientist who helped to develop Brainbow. But he adds that we may never really know how nervous systems work. "These tools allow us to see just how complex they are," he says, "and the more we learn the harder they are to actually understand." 

  • References

    1. Livet, J. et al. Nature 450, 56-62 (2007).
    2. Hampel, S. et al. Nature Meth. doi:10.1038/nmeth.1566 (2011).
    3. Hadjieconomou, D. et al. Nature Meth. doi:10.1038/nmeth.1567 (2011).
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