When expressed in NDNF⁺ neurons and VIP⁺ neurons respectively, the negative-and positive-polarities of the spikes emitted by Ace-mNeon2 and pAce can be used to infer the neuronal subtype. By freeing up the spectral bandwidth, single-channel, dual-population imaging opens up wholly new avenues for scalability. In principle, one can record from up to 4 neuron types concurrently using two color channels. That’s a 2-fold improvement over the current state-of-the-art in calcium imaging…all that with added sub-second temporal resolution.

Imaging conditions: High-speed (0.4-1 kHz) single-channel (green) epifluorescence microscopy. The speed of the recording is reduced by a factor of 10 for visualization.