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Abstract
Neurons transform thousands of synaptic inputs into spiking output, and understanding this transformation requires measuring many submicron synapses at millisecond timescales in vivo. Here we present SLAP2, a two-photon random access projection microscope with no per-target access time cost, yielding over 5-fold faster imaging than existing methods, and performant real-time hardware motion correction. Flexible scanning modes enable measurement of hundreds of synapses at 200 Hz, >800 synapses volumetrically at 50-100 Hz, and compartment-resolved dendritic voltage at 2-10 kHz in behaving mice. Simultaneous dendritic glutamate and somatic calcium imaging in visual cortex pyramidal neurons revealed nonlinear input-output amplification of stimulus onset and preferred stimulus orientation responses. Amplified features are associated with spatially organized inputs and stronger trial-to-trial coupling between synaptic activity and somatic output. These measurements establish SLAP2 as a powerful tool for recording intricate patterns of subcellular fluorescence dynamics in behaving animals.
