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Brief review: Simultaneous two-color optogenetics using novel probes (Klapoetke et al., 2014)

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Cartoon structure of rhodopsin – Wikipedia Commons

To catch up on the field of optogenetics, here is a primer and here is an update on some new stuff.  Also refer to the literature (Mattis et al., 2011; Fenno et al., 2011).  This post assumes a working understanding of Optogenetics, Electrophysiology and some genetics.

A recent paper out of MIT (Ed Boyden lab) identifies two new probes in the ever growing quest to find improve optogenetic tools that will allow for greater spectral separation of activation/excitation wavelengths.  Some of the major challenges for optogenetics research are:

  • genetically expressing opsins, or other light sensitive molecules in model organisms/specific cells
  • finding the right probe that addresses a specific need (i.e. high frequency stimulation, or activate a specific Gi/o pathway)
  • delivering light of a specific wavelength to a particular target in tissue and/or specific cell types
  • spectrally separating a single probe (i.e. ChR2 activated at 470nm) from an imaging probe (i.e. GCamp3, or even the new RCamp which can be activated by blue light)
  • finding two or more probes to express that will not “cross-talk”- such that excitation wavelength of probe 1 will not activate probe 2

Optogenetics allows for very precise control of cell excitability and/or signaling pathways and researchers continue to push the limit of the existing tools and pharmacological agents.  For instance, if studying the interaction of interneurons in the hippocampus area CA1 it would be beneficial to be able to simultaneously activate CCK interneurons while optically inhibiting PV interneurons, or vice versa.  There are currently methods of blocking one versus the other (machR agonist carbachol for instance activates CCK only) but to rapidly be able to control the activity of these neurons would be of great interest in studying the details of synaptic transmission of individual or small groups of neurons (worth noting is these types of on/off, two wavelength probes exist for other applications such as PIF-2.

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Written by Michael Mohammadi

March 31, 2014 at 05:17

Targeting Light: Two recent papers that use active illumination!

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Screen Shot 2014-03-20 at 5.02.10 PMIn my day job I get to travel quite a bit and visit labs around the world who use methods in microscopy in imaging.  These labs range from neuroscience to developmental biology and everything in between, and many of these labs have very specific needs when it comes to light delivery.  The applications I most often work with can involve from high power lasers for ablation and thrombosis, or FRAP or photoconversion.  I’m very focused on optogenetics and uncaging, as well as any type of imaging, especially the use of genetically encoded indicators (for voltage, ions, temperature, pH, etc).  

While many people tend to use a simple configuration for single spot/point illumination, or full field illumination (and we provide these tools as well) where we are unique is in systems that allow for “targeted” or “active illumination.”  There are many methods for targeting light (possibly a good topic for an upcoming review) but the two of the main methods used are either using a galvo-mirror system (very fast mirrors that move a single spot of light in X-Y to create regions with the ability to target to a diffraction limited spot) or a digital micromirror device, or DMD.  

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Written by Michael Mohammadi

March 20, 2014 at 16:04