Glutamate indicators with increased sensitivity and tailored deactivation rates
Understanding how neurons integrate signals from thousands of input synapses requires methods to monitor neurotransmission across many sites simultaneously. The fluorescent protein glutamate indicator iGluSnFR enables visualization of synaptic signaling, but the sensitivity, scale and speed of such measurements are limited by existing variants. Here we developed two highly sensitive fourth-generation iGluSnFR variants with fast activation and tailored deactivation rates: iGluSnFR4f for tracking rapid dynamics, and iGluSnFR4s for recording from large populations of synapses. These indicators detect glutamate with high spatial specificity and single-vesicle sensitivity in vivo. We used them to record natural patterns of synaptic transmission across multiple experimental contexts in mice, including two-photon imaging in cortical layers 1-4 and hippocampal CA1, and photometry in the midbrain. The iGluSnFR4 variants extend the speed, sensitivity and scalability of glutamate imaging, enabling direct observation of information flow through neural networks in the intact brain.
Authors
- Arthur Konnerth (ORCID: https://orcid.org/0000-0002-9548-2676)
- Getahun Tsegaye
- Michael E. Xie (ORCID: https://orcid.org/0000-0002-4026-9181)
- Jihong Zheng (ORCID: https://orcid.org/0000-0003-3024-0892)
- David Kleinfeld (ORCID: https://orcid.org/0000-0001-9797-4722)
- Mirosław Tarnawski (ORCID: https://orcid.org/0000-0001-5297-5668)
- Philipp Leippe (ORCID: https://orcid.org/0000-0002-1109-5610)
- Adrian Negrean (ORCID: https://orcid.org/0000-0002-9256-7412)
- Anyi Liu
- Kaspar Podgorski (ORCID: https://orcid.org/0000-0002-0374-2005)
- Ronak Patel (ORCID: https://orcid.org/0000-0002-6945-6466)
- Jonathan S. Marvin (ORCID: https://orcid.org/0000-0003-2294-4515)
- Bryan MacLennan (ORCID: https://orcid.org/0009-0000-2619-1608)
- Jason Vevea (ORCID: https://orcid.org/0000-0002-3068-973X)
- Srinivas C. Turaga (ORCID: https://orcid.org/0000-0003-3247-6487)
- Abhi Aggarwal (ORCID: https://orcid.org/0000-0002-6336-566X)
- Glenn Turner (ORCID: https://orcid.org/0000-0002-5341-2784)
- Julien Hiblot (ORCID: https://orcid.org/0000-0002-7883-8079)
- Alison G. Tebo (ORCID: https://orcid.org/0000-0003-0788-5617)
- Julianna L. Sun (ORCID: https://orcid.org/0000-0002-5172-4709)
- Kenta M. Hagihara (ORCID: https://orcid.org/0000-0003-0064-0852)
- Daniel Reep
- Rishyashring R. Iyer (ORCID: https://orcid.org/0000-0001-9126-9491)
- Karel Svoboda (ORCID: https://orcid.org/0000-0002-6670-7362)
- Anirudh Palutla
- Arthur Tsang (ORCID: https://orcid.org/0000-0002-2935-8933)
- Lucas W. Kinsey
- Matteo Carandini
- Yang Chen
- Benjamin J. Arthur
- Yonghai Zhang
- Pantong Yao
- L. Federico Rossi
- Jeremy P. Hasseman
Institutions
- St. Jude Children's Research Hospital (US)
- Howard Hughes Medical Institute (US)
- Austrian Academy of Sciences (AT)
- Johns Hopkins University (US)
- University of Calgary (CA)
- Janelia Research Campus (US)
- Italian Institute of Technology (IT)
- University of California San Diego (US)
- Max Planck Institute for Medical Research (DE)
- Munich Cluster for Systems Neurology (DE)
- CeMM Research Center for Molecular Medicine (AT)
- Center for Neuroscience and Cognitive Systems (IT)
- Allen Institute (US)
- Allen Institute for Neural Dynamics (US)
- University College London (GB)
- Libin Cardiovascular Institute of Alberta (CA)
Publication Details
- Journal
- Nature Methods
- Published
- 2025-12-23
- DOI
- https://doi.org/10.1038/s41592-025-02965-z
- Citations
- 4
- Primary Topic
- Neuroscience and Neuropharmacology Research
- Type
- article
- Field-Weighted Citation Impact
- 3.64