m8ta
use https for features.
text: sort by
tags: modified
type: chronology
{1504}
hide / / print
ref: -0 tags: GEVI review voltage sensor date: 03-18-2020 17:43 gmt revision:22 [21] [20] [19] [18] [17] [16] [head]

Various GEVIs invented and evolved:

Ace-FRET sensors

  • PMID-26586188 Ace-mNeonGreen, an opsin-FRET sensor, might still be better in terms of SNR, but it's green.
    • Negative ΔF/F\Delta F / F with depolarization.
    • Fast enough to resolve spikes.
    • Rational design; little or no screening.
    • Ace is about six times as fast as Mac, and mNeonGreen has a ~50% higher extinction coefficient than mCitrine and nearly threefold better photostability (12)

  • PMID-31685893 A High-speed, red fluorescent voltage sensor to detect neural activity
    • Fusion of Ace2N + short linker + mScarlet, a bright (if not the brightest; highest QY) monomeric red fluorescent protein.
    • Almost as good SNR as Ace2N-mNeonGreen.
    • Also a FRET sensor; negative delta F with depolarization.
    • Ace2N-mNeon is not sensitive under two-photon illumination; presumably this is true of all eFRET sensors?
    • Ace2N drives almost no photocurrent.
    • Sought to maximize SNR: dF/F_0 X sqrt(F_0); screened 'only' 18 linkers to see what worked the best. Yet - it's better than VARNAM.
    • ~ 14% dF/F per 100mV depolarization.

Arch and Mac rhodopsin sensors

  • PMID-22120467 Optical recording of action potentials in mammalian neurons using a microbial rhodopsin Arch 2011
    • Endogenous fluorescence of the retinal (+ environment) of microbial rhodopsin protein Archaerhodopsin 3 (Arch) from Halorubrum sodomense.
    • Proton pump without proton pumping capabilities also showed voltage dependence, but slower kinetics.
      • This required one mutation, D95N.
    • Requires fairly intense illumination, as the QY of the fluorophore is low (9 x 10-4). Still, photobleaching rate was relatively low.
    • Arch is mainly used for neuronal inhibition.

  • PMID-25222271 Archaerhodopsin Variants with Enhanced Voltage Sensitive Fluorescence in Mammalian and Caenorhabditis elegans Neurons Archer1 2014
    • Capable of voltage sensing under red light, and inhibition (via proton pumping) under green light.
    • Note The high laser power used to excite Arch (above) fluorescence causes significant autofluorescence in intact tissue and limits its accessibility for widespread use.
    • Archers have 3-5x the fluorescence of WT Arch -- so, QY of ~3.6e-3. Still very dim.
    • Archer1 dF/F_0 85%; Archer2 dF/F_0 60% @ 100mV depolarization (positive sense).
    • Screened the proton pump of Gloeobacter violaceus rhodopsin; found mutations were then transferred to Arch.
      • Maybe they were planning on using the Geobacter rhodopsin, but it didn't work for some reason, so they transferred to Arch..
    • TS and ER export domains for localization.

  • PMID-24755708 Imaging neural spiking in brain tissue using FRET-opsin protein voltage sensors MacQ-mOrange and MacQ-mCitrine.
    • L. maculans (Mac) rhodopsin (faster than Arch) + FP mCitrine, FRET sensor + ER/TS.
    • Four-fold faster kinetics and 2-4x brighter than ArcLight.
      • No directed evolution to optimize sensitivity or brightness. Just kept the linker short & trimmed residues based on crystal structure.
    • ~5% delta F/F, can resolve spikes up to 10Hz.
    • Spectroscopic studies of the proton pumping photocycle in bacteriorhodopsin and Archaerhodopsin (Arch) have revealed that proton translocation through the retinal Schiff base changes chromophore absorption [24-26]
    • Used rational design to abolish the proton current (D139N and D139Q aka MacQ) ; screens to adjust the voltage sensing kinetics.
    • Still has photocurrents.
    • Seems that slice / in vivo is consistently worse than cultured neurons... in purkinje neurons, dF/F 1.2%, even though in vitro response was ~ 15% to a 100mV depolarization.
    • Imaging intensity 30mw/mm^2. (3W/cm^2)

  • PMID-24952910 All-optical electrophysiology in mammalian neurons using engineered microbial rhodopsins QuasAr1 and QuasAr1 2014
    • Directed evolution approach to improve the brightness and speed of Arch D95N.
      • Improved the fluorescence QY by 19 and 10x. (1 and 2, respectively -- Quasar2 has higher sensitivity).
    • also developed a low-intensity channelrhodopsin, Cheriff, which can be activated by blue light (lambda max = 460 nm)dim enough to not affect QuasAr.
    • They call the two of them 'Optopatch 2'.
    • Incident light intensity 1kW / cm^2 (!)

  • PMID-29483642 A robotic multidimensional directed evolution approach applied to fluorescent voltage reporters. Archon1 2018
    • Started with QuasAr2 (above), which was evolved from Arch. Intrinsic fluorescence of retinal in rhodopsin.
    • Expressed in HEK293T cells; then FACS, robotic cell picking, whole genome amplification, PCR, cloning.
    • Also evolved miRFP, deep red fluorescent protein based on bacteriophytochrome.
    • delta F/F of 80 and 20% with a 100mV depolarization.
    • We investigated the contribution of specific point mutations to changes in localization, brightness, voltage sensitivity and kinetics and found the patterns that emerged to be complex (Supplementary Table 6), with a given mutation often improving one parameter but worsening another.
    • If the original QY of Arch was 9e-4, and Quasar2 improved this by 10, and Archon1 improved this by 2.3x, then the QY of Archon1 is 0.02. Given the molar extinction coefficient is ~ 50000 for retinal, this means the brightness of the fluorescent probe is low, 1. (good fluorescent proteins and synthetic dyes have a brightness of ~90).
    • Big paper, moderate improvement.
    • SomArchon1 and SomCheriff serve as the basis of Optopatch4, e.g. All-optical electrophysiology reveals excitation, inhibition, and neuromodulation in cortical layer 1
    • Slow photobleaching, consistent with other Arch based GEVIs.

VSD - FP sensors

  • PMID-28811673 Improving a genetically encoded voltage indicator by modifying the cytoplasmic charge composition Bongwoori 2017
    • ArcLight derivative.
    • Arginine (positive charge) scanning mutagenesis of the linker region improved the signal size of the GEVI, Bongwoori, yielding fluorescent signals as high as 20% ΔF/F during the firing of action potentials.
    • Used the mutagenesis to shift the threshold for fluorescence change more negative, ~ -30mV.
    • Like ArcLight, it's slow.
    • Strong baseline shift due to the acidification of the neuron during AP firing (!)

  • Attenuation of synaptic potentials in dentritic spines
    • Found that SNR / dF / F_0 is limited by intracellular localization of the sensor.
      • This is true even though ArcLight is supposed to be in a dark state in the lower pH of intracellular organelles.. a problem worth considering.
      • Makes negative-going GEVI's more practical, as those not in the membrane are dark.

  • Fast two-photon volumetric imaging of an improved voltage indicator reveals electrical activity in deeply located neurons in the awake brain ASAP3 2018
    • Opsin-based GEVIs have been used in vivo with 1p excitation to report electrical activity of superficial neurons, but their responsivity is attenuated for 2p excitation. (!)
    • Site-directed evolution in HEK cells.
    • Expressed linear PCR products directly in the HEK cells, with no assembly / ligation required! (Saves lots of time: normally need to amplify, assemble into a plasmid, transfect, culture, measure, purify the plasimd, digest, EP PCR, etc).
    • Screened in a motorized 384-well conductive plate, electroporation electrode sequentially activates each on an upright microscope.
    • 46% improvement over ASAP2 R414Q
    • Ace2N-4aa-mNeon is not responsive under 2p illum; nor is Archon1 or Quasar2/3
    • ULOVE = AOD based fast local scanning 2-p random access scope.

  • Bright and tunable far-red chemigenetic indicators
    • GgVSD (same as ASAP above) + cp HaloTag + Si-Rhodamine JF635
    • ~ 4% dF/F_0 during APs.
    • Found one mutation, R476G in the linker between cp Halotag and S4 of the VSD, which doubled the sensitivity of HASAP.
    • Also tested a ArcLight type structure, CiVSD fused to Halotag.
      • HarcLght had negative dF/F_0 and ~ 3% change in response to APs.
    • No voltage sensitivity when the synthetic dye was largely in the zwitterionic form, eg. tetramethylrodamine.

{1487}
hide / / print
ref: -0 tags: adaptive optics sensorless retina fluorescence imaging optimization zernicke polynomials date: 11-15-2019 02:51 gmt revision:0 [head]

PMID-26819812 Wavefront sensorless adaptive optics fluorescence biomicroscope for in vivo retinal imaging in mice

  • Idea: use backscattered and fluorescence light to optimize the confocal image through imperfect optics ... and the lens of the mouse eye.
    • Optimization was based on hill-climbing / line search of each Zernicke polynomial term for the deformable mirror. (The mirror had to be characterized beforehand, naturally).
    • No guidestar was needed!
  • Were able to resolve the dendritic processes of EGFP labeled Thy1 ganglion cells and Cx3 glia.

{87}
hide / / print
ref: Nicolelis-1993 tags: neurons somatosensory nicolelis rats thalamus date: 01-03-2012 23:30 gmt revision:2 [1] [0] [head]

from the Scientific American:

  • blocking (single?) neuron activity in S1 cortex affects the responses of VPM neurons in the thalamus - indicating that descending feedback signals in the cortex to the VPM could have a major role in modulating the ascending information.
  • if you implant a cuff electrode aroung the trigeminal nerve, the evoked responses in S1 and VPM are dependent on the behavioral state of the animal (of course!). this effect is so pronounced that, when the rats were not 'paying attention', only the first stimulus of a series evoked a response; when the rat was whisking, stimulation was faithfully reported.

{926}
hide / / print
ref: Nicolelis-1998.11 tags: spatiotemporal spiking nicolelis somatosensory tactile S1 3b microwire array rate temporal coding code date: 12-28-2011 20:42 gmt revision:3 [2] [1] [0] [head]

PMID-10196571[0] Simultaneous encoding of tactile information by three primate cortical areas

  • owl monkeys.
  • used microwires arrays to decode the location of tactile stimuli; location was encoded through te population, not within single units.
  • areas 3b, S1 & S2.
  • used LVQ (learning vector quantization) backprop, LDA to predict/ classify touch trials; all yielded about the same ~60% accuracy. Chance level 33%.
  • Interesting: "the spatiotemporal character of neuronal responses in the SII cortex was shown to contain the requisite information for the encoding of stimulus location using temporally patterned spike sequences, whereas the simultaneously recorded neuronal responses in areas 3b and 2 contained the requisite information for rate coding."
    • They support this result by varying bin widths and looking at the % of correctly classivied trials. in SII, increasing bin width decreases (slightly but significantly) the prediction accuracy.

____References____

[0] Nicolelis MA, Ghazanfar AA, Stambaugh CR, Oliveira LM, Laubach M, Chapin JK, Nelson RJ, Kaas JH, Simultaneous encoding of tactile information by three primate cortical areas.Nat Neurosci 1:7, 621-30 (1998 Nov)

{896}
hide / / print
ref: Friston-2002.1 tags: neuroscience philosophy feedback top-down sensory integration inference date: 10-25-2011 23:24 gmt revision:0 [head]

PMID-12450490 Functional integration and inference in the brain

  • Extra-classical tuning: tuning is dependent on behavioral context (motor) or stimulus context (sensory). Author proposes that neuroimaging can be used to investigate it in humans.
  • "Information theory can, in principle, proceed using only forward connections. However, it turns out that this is only possible when processes generating sensory inputs are invertible and independent. Invertibility is precluded when the cause of a percept and the context in which it is engendered interact." -- proof? citations? Makes sense though.
  • Argues for the rather simplistic proof of backward connections via neuroimaging..

{619}
hide / / print
ref: -0 tags: thalamus sensory Ochoa date: 10-05-2008 16:41 gmt revision:2 [1] [0] [head]

http://jp.physoc.org/cgi/reprint/342/1/633

{476}
hide / / print
ref: bookmark-0 tags: wideband oxygen sensor diffusion nernst lambda date: 10-22-2007 03:41 gmt revision:0 [head]

http://www.wbo2.com/lsu/lsuworks.htm