PMID-30420685 Fast in-vivo voltage imaging using a red fluorescent indicator
- Kannan M, Vasan G, Huang C, Haziza S, Li JZ, Inan H, Schnitzer MJ, Pieribone VA.
- Other genetically encoded voltage indicators (GEVI):
- PMID-22958819 ArcLight (Peribone also last author) ; sign of negative, but large, 35%! Slow tho? improvement in speed
- ASAP3 large,
- PMID-26586188 Ace-mNeon FRET based, Acetabularia opsin, fast kinetics + brightness of mNeonGreen.
- Archon1 -- fast and sensitive, found (like VARNUM) using a robotic directed evolution or direct search strategy.
- VARNAM is based on Acetabularia (Ace) + mRuby3, also FRET based, found via high-throughput voltage screen.
- Archaerhodopsin require 1-12 W/mm^2 of illumination, vs. 50 mw/mm^2 for GFP based probes. Lots of light!
- Systematic optimization of voltage sensor function: both the linker region (288 mutants), which affects FRET efficiency, as well as the opsin fluorophore region (768 mutants), which affects the wavelength of absorption / emission.
- Some intracellular clumping (which will negatively affect sensitivity), but mostly localized to the membrane.
- Sensitivity is still imperfect -- 4% in-vivo cortical neurons, though it’s fast enough to resolve 100 Hz spiking.
- Can resolve post-synaptic EPSCs, but < 1 % .
- Tested all-optical ephys using VARNAM + blueshifted channelrhodopsin, CheRiff, both sparsely, and in PV targeted transgenetic model. Both work, but this is a technique paper; no real results.
-
- Tested TEMPO fiber-optic recording in freely behaving mice (ish) -- induced ketamine waves, 0.5-4Hz.
- And odor-induced activity in flies, using split-Gal4 expression tools. So many experiments.
|