Download Optogenetics is the combination of genetics and optics to control

BIOL463 Fall 2016
Three-minute technique
Optogenetics
1.
Names and contributions of group members:
Julius Booth: Research, Write-up, Write-up final copy
Celina Li: Research, Write-up, Powerpoint
Martin Thygesen: Research, Write-up, Powerpoint
2.
Technique chosen:
Optogenetics
3.
What general biological, chemical, and/or physical principles and concepts is this
technique based on?
Optogenetics is the combination of genetics and optics to control events within
specific cells of living tissue.
Some proteins are light sensitive, meaning that certain wavelengths of light can
change their properties/conformation. Light-sensitive ion channels can be introduced
to neurons. The neuron will then either inhibit or elicit (depending on the type of
channel) action potentials when illuminated by a laser. This is caused by the channel
opening and allowing ions to flow in to the cell, sodium for excitatory channels
(channelrhodopsins), chlorine for inhibitory (halorhodopsins).
The technique also relies on the associated technologies for delivering light deep into
organisms as complex as freely moving mammals, for targeting light-sensitivity to
cells of interest, and for assessing specific readouts, or effects, of this optical control.
4.
What does this technique ‘do’?
Biologists want to be able to control specific events in specific locations on cue in
vivo.
When working with neurons, electrodes aren’t specific enough and drugs act too
slowly, but optogenetics is both specific and practically instantaneous (millisecondscale timing precision).
The control optogenetics offers allows scientists to test complex systems that are
only meaningful when kept whole ie. neural networks that produce behavior; not
possible with one neuron on a petri dish. Biologists can trigger specific neurons and
see how the neuron interacts with the rest of the system and what behaviors it
produces in organism.
BIOL463 Fall 2016
Three-minute technique
Similarly, other pathways can be triggered by light, like using modified versions of
rhodopsin to initiate G-protein dependent signal pathways in specific cell lines at
will.
http://www.annualreviews.org/doi/full/10.1146/annurev-neuro-061010113817?url_ver=Z39.882003&rfr_id=ori%3Arid%3Acrossref.org&rfr_dat=cr_pub%3Dpubmed
Figure 1. Different microbial ion channels respond to different light wavelengths
and transport different ions. (a) For this particular protein, when light is
illuminating onto the protein, the channel responds to open for cation ion to enter
the intracellular space. (b) ChR2 responds to light of approximately 470 nm and
allows for sodium ions to enter depolarizing the cell. Action potentials are
generated. (c) NpHR responds to light of approximately 580 nm and allows for
chloride ions to enter hyperpolarizing the cell. The train of action potentials is
stopped. (d) VChR1 responds to light of approximately 589 nm and allows for
sodium ions to enter depolarizing the cell and firing action potentials.
5.
What applications is this technique employed for?
Can be used to examine neural circuitry.
Can be used to study brain disorders such as Parkinson’s disease and psychiatric
diseases such as schizophrenia. Some results showing optogenetics could be used to
treat these diseases (in mice only… so far).
New experiments have investigated the possibilities of using optogenetics in order to
achieve a light-dependent genome editing tool. This technique uses a split Cas9
BIOL463 Fall 2016
Three-minute technique
protein bound to light activated fusion proteins, allowing the assembly of functional
Cas9 only when the fusion proteins are activated by light.
http://www.nature.com/nbt/journal/v33/n7/full/nbt.3245.html
6.
What questions relating to gene regulation and/or development can be addressed
using this technique? Provide two examples (peer-reviewed papers) that use this
technique.
Examining sensorimotor development in chick embryos:
http://jn.physiology.org/content/jn/106/5/2776.full.pdf
Understanding the epigenetic regulatory mechanisms of telomere length
maintenance:
http://www.impactjournals.com/oncotarget/index.php?journal=oncotarget&page=arti
cle&op=view&path[]=10394&pubmed-linkout=1
7.
What critical reagents are required to use this technique?
Transformation mechanism (Viral vector or CRE system).
Gene expression of high-fidelity mutants of microbial channelrhodopsins,
halorhodopsins.
Light source able to reach neurons/cells and selectively turn on/off.
8.
What critical information is required to be able to employ this technique?
Need to know how to target specific cells and which cells you want to target.
Need to know how to selectively express light-sensitive channel proteins.
Microbial opsins require all-trans retinal (Vitamin A) to absorb photons – Vertebrate
tissues contain natural all-trans retinal, so optogenetic control is feasible in mammals.
9.
At least two resources/information sources that you recommend for learning more
about this technique (one or more may be developed by your group):
http://www.nature.com/nmeth/journal/v8/n1/full/nmeth.f.324.html
BIOL463 Fall 2016
Three-minute technique
http://www.scientificamerican.com/article/revolutionary-neuroscience-techniqueslated-for-human-clinical-trials/
http://www.scientificamerican.com/article/optogenetics-controlling/
10. List of references consulted:
http://www.annualreviews.org/doi/full/10.1146/annurev-neuro-061010113817?url_ver=Z39.882003&rfr_id=ori%3Arid%3Acrossref.org&rfr_dat=cr_pub%3Dpubmed
http://www.impactjournals.com/oncotarget/index.php?journal=oncotarget&page=article
&op=view&path[]=10394&pubmed-linkout=1
http://jn.physiology.org/content/jn/106/5/2776.full.pdf
http://www.nature.com/nmeth/journal/v8/n1/full/nmeth.f.324.html
http://www.nature.com/nrm/journal/v15/n8/full/nrm3837.html
http://www.nature.com/nbt/journal/v33/n2/full/nbt.3124.html
http://www.nature.com/nchembio/journal/v10/n3/full/nchembio.1430.html
http://www.nature.com/nbt/journal/v33/n7/full/nbt.3245.html
https://www.ncbi.nlm.nih.gov/pubmed/25896279
http://www.scientificamerican.com/article/revolutionary-neuroscience-technique-slatedfor-human-clinical-trials/
http://www.scientificamerican.com/article/optogenetics-controlling/