Independent two-color optogenetic excitation of neural populations

The optical modulation of neurons with channelrhodopsins, a class of genetically encoded light-gated ion channels, has enabled the spatiotemporally precise interrogation of the roles individual cell types play in neural circuit dynamics. A topic of great interest to the neuroscience community is the independent optical excitation of two distinct neuron populations with different wavelengths, which would enable the interrogation of emergent phenomena such as circuit dynamics, plasticity, and neuromodulation. Previous implementations have focused on maximizing spectral separation by driving one channelrhodopsin in the violet (405 nm) and the other in the yellow (590 nm), yet it has not been possible to achieve independent violet excitation without eliciting spikes from both populations, due to the intrinsic UV-blue light sensitivity of the retinal chromophore. This thesis designs and implements an improved two-color excitation scheme where effective light sensitivity is utilized to achieve independent optical excitation in blue (470 nm) and red (625 nm) channels. Zero post-synaptic crosstalk is demonstrated in acute murine slice, using two novel channeirhodopsins identified from a systematic screen of 80 naturally occurring, previously uncharacterized opsins in primary neuron culture. Gene88 is the first known yellow-peaked channelrhodopsin, with a peak 45 nm more red-shifted than any previous channelrhodopsin, while Gene90 has the fastest channel turn on, turn off, and recovery kinetics of any known channelrhodopsin. These opsins' novel properties enable the first known demonstration of post-synaptic crosstalk-free two-color excitation with temporally precise modulation of spatially inseparable neuron populations.

Department dc:contributor.department

Massachusetts Institute of Technology. Department of Brain and Cognitive Sciences.

Grantor dc:publisher

Massachusetts Institute of Technology

Year dc:date.issued

2013

Author dc:creator

• Klapoetke, Nathan Cao

Advisor dc:contributor.advisor

• Edward S. Boyden, III.

• Brain and Cognitive Sciences.

Statement dc:rights

• M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission.

Licence dc:rights.uri

• http://dspace.mit.edu/handle/1721.1/7582

Language dc:language.iso

eng

Handle dc:identifier.uri

http://hdl.handle.net/1721.1/86285

OAI identifier oai:identifier

oai:dspace.mit.edu:1721.1/86285