High-speed laser microsurgery of alert fruit flies for fluorescence imaging of neural activity.

Microscopy and neurophysiology experiments in live animals commonly involve complex surgical preparations, which are often time-consuming, demand considerable manual dexterity, and can sharply limit experimental throughput. Here we present a spatially precise laser microsurgical technique using a pulsed UV laser. Our approach reduces surgical time by up to two orders of magnitude while substantially improving reproducibility. Using the fruit fly as a model, we show that laser microsurgery leaves complex behaviors intact and allows us to visualize brain activity in live flies for up to 18 h, more than four times longer than reported previously using hand dissection. We also demonstrate laser microsurgery on nematodes, ants, and the mouse cranium, illustrating broad potential utility for both optical and electrophysiological studies.