Basic NeuroscienceMonitoring synaptic and neuronal activity in 3D with synthetic and genetic indicators using a compact acousto-optic lens two-photon microscope

- We expand the utility of acousto-optic lens (AOL) 3D 2P microscopy.
- We show rapid, simultaneous monitoring of synaptic inputs distributed in 3D.
- First use of genetically encoded indicators with AOL 3D functional imaging.
- Measurement of sensory-evoked neuronal population activity in 3D in vivo.
- Strategies for improving the measurement of the timing of neuronal signals.

BackgroundTwo-photon microscopy is widely used to study brain function, but conventional microscopes are too slow to capture the timing of neuronal signalling and imaging is restricted to one plane. Recent development of acousto-optic-deflector-based random access functional imaging has improved the temporal resolution, but the utility of these technologies for mapping 3D synaptic activity patterns and their performance at the excitation wavelengths required to image genetically encoded indicators have not been investigated.New methodHere, we have used a compact acousto-optic lens (AOL) two-photon microscope to make high speed [Ca2+] measurements from spines and dendrites distributed in 3D with different excitation wavelengths (800-920 nm).ResultsWe show simultaneous monitoring of activity from many synaptic inputs distributed over the 3D arborisation of a neuronal dendrite using both synthetic as well as genetically encoded indicators. We confirm the utility of AOL-based imaging for fast in vivo recordings by measuring, simultaneously, visually evoked responses in 100 neurons distributed over a 150 μm focal depth range. Moreover, we explore ways to improve the measurement of timing of neuronal activation by choosing specific regions within the cell soma.Comparison with existing methodsThese results establish that AOL-based 3D random access two-photon microscopy has a wider range of neuroscience applications than previously shown.ConclusionsOur findings show that the compact AOL microscope design has the speed, spatial resolution, sensitivity and wavelength flexibility to measure 3D patterns of synaptic and neuronal activity on individual trials.