Measuring neuronal input-output operations in vivo with random access projection microscopy
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.
Authors
- Giulio Tononi (ORCID: https://orcid.org/0000-0002-3892-4087)
- Michael E. Xie (ORCID: https://orcid.org/0000-0002-4026-9181)
- Matthew D. Golub (ORCID: https://orcid.org/0000-0003-4508-0537)
- Chiara Cirelli (ORCID: https://orcid.org/0000-0003-2563-677X)
- Yukun Alex Hao (ORCID: https://orcid.org/0000-0002-3454-5733)
- Daniel Flickinger (ORCID: https://orcid.org/0000-0002-8361-3122)
- Maedeh Seyedolmohadesin (ORCID: https://orcid.org/0000-0003-3742-394X)
- Jackie Swapp (ORCID: https://orcid.org/0009-0008-4965-6242)
- Kaspar Podgorski (ORCID: https://orcid.org/0000-0002-0374-2005)
- Jonathan King
- Bruce E. Kimmel
- Michael Z. Lin (ORCID: https://orcid.org/0000-0002-0492-1961)
- Adam S. Charles (ORCID: https://orcid.org/0000-0002-9045-3489)
- Jacob R. Glaser (ORCID: https://orcid.org/0000-0003-4463-207X)
- Kort Driessen (ORCID: https://orcid.org/0009-0003-5051-5213)
- Karel Svoboda (ORCID: https://orcid.org/0000-0002-6670-7362)
- Manni He (ORCID: https://orcid.org/0009-0001-4237-900X)
- Henry Loeffler (ORCID: https://orcid.org/0009-0007-3935-6863)
- Georg Jaindl (ORCID: https://orcid.org/0000-0001-6134-6767)
- Yanru Li (ORCID: https://orcid.org/0000-0002-8048-7220)
- Ali Williford (ORCID: https://orcid.org/0009-0000-7282-7515)
- Carter R. Peene (ORCID: https://orcid.org/0009-0000-6660-2264)
- Katrina Nguyen (ORCID: https://orcid.org/0009-0009-8547-9703)
- Madeline Tom
- Jimin Park
- Ryan Gibiser
Institutions
- Howard Hughes Medical Institute (US)
- University of Wisconsin–Madison (US)
- Johns Hopkins University (US)
- Janelia Research Campus (US)
- University of Washington (US)
- MBF Bioscience (United States) (US)
- Allen Institute (US)
- Allen Institute for Neural Dynamics (US)
- Stanford University (US)
Publication Details
- Journal
- bioRxiv (Cold Spring Harbor Laboratory)
- Published
- 2026-11-06
- DOI
- https://doi.org/10.64898/2026.11.05.764682
- Primary Topic
- Advanced Fluorescence Microscopy Techniques
- Type
- preprint
Funders
- Paul and Daisy Soros Fellowships for New Americans
- Chan Zuckerberg Initiative