A high-throughput all-optical platform for mapping gene perturbations to electrophysiology
Electrical phenotyping remains a bottleneck in linking genes to cellular function. We developed a high-throughput all-optical electrophysiology platform combining an engineered human induced pluripotent stem cell (hiPSC) line, wide-field voltage imaging, and automated analysis. The hiPSC line integrates inducible NGN2, CRISPR interference, and degron-gated expression of Voltron2 and CheRiff, enabling neuronal differentiation, targeted gene knockdown, and optical electrophysiology. The microscope records approximately 1,500 spiking glutamatergic neurons at 800 Hz across a 4x4 mm field, with larger fields supporting approximately 3,000 neurons at 660 Hz. Automated analysis extracts single-cell voltage traces and quantifies firing patterns, action-potential waveforms, and subthreshold dynamics. Arrayed knockdown of five sodium- and potassium-channel genes produced distinct electrophysiological phenotypes. The engineered hiPSC line also supported optical electrophysiology recordings from motor neurons, astrocytes, and cardiomyocytes. These results establish a scalable platform for high-throughput electrophysiological phenotyping and functional genomics in hiPSC-derived cells.
Authors
- Richard Theodore Lee (ORCID: https://orcid.org/0000-0003-4687-1381)
- Russell T. Walton (ORCID: https://orcid.org/0000-0002-0546-474X)
- Paul C. Blainey (ORCID: https://orcid.org/0000-0002-4889-8783)
- Ralda Nehme (ORCID: https://orcid.org/0000-0001-7215-3311)
- Daniel G. Itkis (ORCID: https://orcid.org/0009-0003-8948-2260)
- Luke D. Lavis (ORCID: https://orcid.org/0000-0002-0789-6343)
- Ernest Fraenkel (ORCID: https://orcid.org/0000-0001-9249-8181)
- Hannah K. Fandl (ORCID: https://orcid.org/0000-0002-4971-0738)
- Daozheng Gong (ORCID: https://orcid.org/0000-0002-3951-1890)
- Alec M. Barrios
- Adam E. Cohen (ORCID: https://orcid.org/0000-0002-8699-2404)
- Takeshi Uenaka (ORCID: https://orcid.org/0000-0001-5786-5038)
- Samouil L. Farhi (ORCID: https://orcid.org/0000-0003-1359-4568)
- Marius Wernig (ORCID: https://orcid.org/0000-0002-5309-515X)
- J Zhang
- Sarah E. Plutkis (ORCID: https://orcid.org/0009-0003-1822-0466)
- Keemaya Sahni
Institutions
- Broad Institute (US)
- Howard Hughes Medical Institute (US)
- Harvard University (US)
- Janelia Research Campus (US)
- Stanford Medicine (US)
- Koch Institute for Integrative Cancer Research At MIT (US)
- Stanley Center for Psychiatric Research (US)
- Institute for Stem Cell Biology and Regenerative Medicine (US)
- Massachusetts Institute of Technology (US)
- Stanford University (US)
Publication Details
- Journal
- bioRxiv (Cold Spring Harbor Laboratory)
- Published
- 2026-11-11
- DOI
- https://doi.org/10.64898/2026.11.09.765233
- Primary Topic
- Neuroscience and Neural Engineering
- Type
- preprint