A near-infrared genetically encoded calcium indicator for in vivo imaging

Nat Biotechnol. 2021 Mar;39(3):368-377. doi: 10.1038/s41587-020-0710-1. Epub 2020 Oct 26.

Abstract

While calcium imaging has become a mainstay of modern neuroscience, the spectral properties of current fluorescent calcium indicators limit deep-tissue imaging as well as simultaneous use with other probes. Using two monomeric near-infrared (NIR) fluorescent proteins (FPs), we engineered an NIR Förster resonance energy transfer (FRET)-based genetically encoded calcium indicator (iGECI). iGECI exhibits high levels of brightness and photostability and an increase up to 600% in the fluorescence response to calcium. In dissociated neurons, iGECI detects spontaneous neuronal activity and electrically and optogenetically induced firing. We validated the performance of iGECI up to a depth of almost 400 µm in acute brain slices using one-photon light-sheet imaging. Applying hybrid photoacoustic and fluorescence microscopy, we simultaneously monitored neuronal and hemodynamic activities in the mouse brain through an intact skull, with resolutions of ~3 μm (lateral) and ~25-50 μm (axial). Using two-photon imaging, we detected evoked and spontaneous neuronal activity in the mouse visual cortex, with fluorescence changes of up to 25%. iGECI allows biosensors and optogenetic actuators to be multiplexed without spectral crosstalk.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Calcium / chemistry*
  • Fluorescence Resonance Energy Transfer
  • HeLa Cells
  • Humans
  • Mice
  • Neurons / physiology
  • Spectroscopy, Near-Infrared / methods*
  • Visual Cortex / diagnostic imaging
  • Visual Cortex / physiology

Substances

  • Calcium