That's the title of my 'Thought Experiment' column in the next issue of 'The Scientist', due to appear on February 1. Sarah Greene from The Scientist approached me in my role as F1000 faculty member at this year's SfN annual meeting in San Diego and asked me if I didn't want to write something for The Scientist.

The short article is about visualizing neuronal activity in small brains. I've recently applied for a starting grant at the European Research Council to develop a microscope which can record the activity in most of the Drosophila brain in 4D (space + time). This application is a revised version after the first one got good reviews but wasn't funded due to the technique being too risky. We have since tested the originally proposed technique and found that indeed it didn't work, as the reviewers assumed. Therefore, we are now proposing to develop a new kind of microscope, rather than just using an existing one.

Interestingly, just around the time I was writing this article, a paper was published by colleagues from UCLA that used a related, but slightly different technique to record in 4D from mouse brains: "Simultaneous two-photon calcium imaging at different depths with spatiotemporal multiplexing" Their technique is not identical to what we have tried in flies, but similar. The authors are using multiple laser beams to excite molecules in the mouse brain that act as reporters of neural activity. If the neuron they are incorporated in is active, they fluoresce in one wavelength when hit by the laser, and in a different one when the neuron is inactive. Previous studies had just picked one plane in the brain and then recorded the activity of neurons in this plane. One of several reasons why it wasn't possible, until now, to record from more than one plane, was that the speed with which one could move the laser around was too slow. The authors of this study overcome this problem by using multiple beams at the same time. This allowed them to monitor 100-200 neurons in a ~400 × 400 μm cube of mouse cortex.

While I'm not surprised that this technique works great in mice, there is one main reason why this method does not work in flies. In order to capture sufficient light in the much denser fly brain, we need dwelling times of around one microsecond per voxel (a 3D pixel). If we do this with more than one laser, the heat being transfered into the fly brain is just too great: the brain will literally boil (ask us how we know: this was exactly the method we suggested in the previous grant and tried out before applying again in this round). While this is obviously a lesser problem for the much larger mouse brain, where the heat can dissipate much more easily, I'd still like to know the temperature of the cube Cheng at al. were recording from. After all, we wouldn't want to interfere more than absolutely necessary with the object we're attempting to measure.

Overall, this is the way microscopy is heading these days: after having increased the spatial resolution to below the optical diffraction limit, now researchers all over the world are turning towards increasing the temporal resolution. This will allow us to record 3D videos of brain activity, very similar to the way currently being done with fMRI in humans, but with ten times the temporal and many times the spatial resolution. This latest study (and hopefully our project, if the ERC funds it) is a great step in this direction. Hopefully, many more will follow.

Attached at the end of my article in The Scientist is a 5min video where I'm giving a brief summary of our latest research on the Drosophila orthologue of the FOXP2 gene. This work was presented as a poster at the abovementioned SfN meeting and is also uploaded to the F1000 website.

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Cheng, A., Gonçalves, J., Golshani, P., Arisaka, K., & Portera-Cailliau, C. (2011). Simultaneous two-photon calcium imaging at different depths with spatiotemporal multiplexing Nature Methods, 8 (2), 139-142 DOI: 10.1038/nmeth.1552