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Optogenetics, Opsins and nVoke

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Posted by Swagata Basu - 06.13.2017

 

Opsins are transmembrane proteins and a fundamental component of the optogenetic toolkit. As optogenetic actuators, opsins modify the activity of cells in which they are expressed when exposed to light and can induce single to multiple action potentials or inhibit neural activity with millisecond precision. Opsins can be categorized into two main classes: microbial opsins and vertebrate opsins. Microbial opsins like rhodopsins are found in prokaryotic and eukaryotic microbial organisms like algae and bacteria with a single membrane-bound protein component that functions as a pump or channel. Vertebrate opsins are found in animal cells and are mainly used for vision.

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Image: Rhodopsins are used as actuators in optogenetics. Channelrhodopsin (ChR) and halorhodopsin (HR) mediate light-dependent transport of ions across the cell to modulate neuronal activity.                           Credit: Jacqueline DeRose, Inscopix.

 


Our Newest Innovation


At Inscopix, we developed nVoke, an integrated optogenetics and calcium imaging platform that relies on red-shifted microbial opsins and enables temporally precise control of genetically defined neurons in the mammalian brain in vivo using light. The system integrates two LED light sources to allow for simultaneous or sequential cellular-resolution imaging and optogenetic manipulation within the same field of view in freely behaving mice. Our scientists have validated genetically-encoded tools like excitatory and inhibitory red-shifted opsins paired with GCaMP calcium indicators that show negligible biological crosstalk (at biologically relevant specs).


Choosing the Right Opsin with nVoke

Excitatory protocols with opsins

nVoke is well suited for pairing GCaMP6 with Chrimson for excitatory protocols in terminals of freely behaving mice. By modulating presynaptic terminals and imaging postsynaptic activity, nVoke can be used to study how two brain regions communicate effectively. You can read more about how nVoke enables this kind of inter-regional functional connectivity study here.


Chrimson Key Features:

Chrimson is a red light-drivable channelrhodopsin derived from Chlamydomonas noctigama (freshwater algae).

  • Spectral peak at 590 nm (45 nm more red shifted than previously known channelrhodopsins and has the highest wavelength absorbance maximum)1,2
  • Ideal for optogenetic experiments requiring deep tissue penetration.
  • Has been previously used to control behavioral responses in Drosophila1 and C. elegans3


Inhibitory protocols with opsins


nVoke can be used for pairing one-photon Ca2+ imaging of GCaMP with full-field somatic and terminal inhibition using NpHR3.0 or Jaws.


NpHR3.0 Key Features:

NpHR3.0 is a light-gated chloride pump halorhodopsin derived from Natromonas pharaonis4.


  • Enables optical control of membrane potential and reversible silencing of targeted neurons with far-red light and has a spectral peak at 590 nm.
  • Shows improved localization to the plasma membrane and has a significantly enhanced inhibitory capacity without requiring increased light power.
  • Penetrates deeper into biological tissue, thereby increasing the ease with which the effector is activated.

Jaws Key Features:

Jaws is a derived from the most red-shifted cruxhalorhodopsin Halo57 from Haloarcula salinarium (strain Shark)5.

  • Mediates strong red light-driven neural inhibition and dramatically increased photocurrent without altering the red-shifted spectrum of Halo57 with a spectral peak at 600 nm.
  • Has an enhanced spike frequency distribution.

 

With nVoke, researchers now have a unique platform that enables simultaneous and sequential cellular-resolution imaging and optical manipulation within the same field of view in freely behaving mice and allows you to dissect the neural basis of behavior at the circuit level. Choosing the right opsin for your particular scientific application is critical; some example research applications are highlighted in these case studies that test the relationship between specific neural circuits in freely moving behaviors.

Do you use these opsins for your research? Share your experience with us!

 nVoke Application Highlight


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