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(Fig source from: Mixing fiber optics with genetics has created a revolutionary tool for studying the brain)
To understand and manipulate components of the brain, “the central challenge in neuroscience is the difficulty of controlling just one type of neuron at a time!” observed Francis Crick prophetically in 1979. Karl Deisseroth, who devised ‘Optogenetics’, noted it helps scientists overcome ‘this’ challenge. Optogenetics is the combination of genetic and optical methods to achieve functional control of events in specific living cells.
It is hardly a couple of decades since the possibility of the technique, Optogenetics, developed with the discovery of ‘channelrhodopsin-1’, an ion channel protein that responds to light. The first gated channel light activated single component protein was discovered in 1971. In 2002, the light-sensitive protein channelrhodopsin-1 was discovered in single-celled green algae (where it aided the algae to swim towards light for photosynthesis); in just three years, the rhodopsin was genetically brought into hippocampal neurons, giving rise to Optogenetics (conferring millisecond-precision control of neuronal spiking).
Optogenetics is a technology that allows targeted, fast control of precisely defined events in biological systems as complex as freely moving mammals.
Karl Deisseroth, in Special Feature on Optogenetics, “Method of the Year” 2010, Nature Methods
The technique is based on the principle of ions as the common signal transmission mediator, between the light-sensitive proteins and the activity-behavior response of neurons. The protein channels are engineered into the neurons resulting in responses to light, subsequent generation of ion influx, and transmission of the signal across neurons. Using genetics, these proteins (optogenetic probes) are expressed in the desired and specific neurons. Light-sensitive ion channels bacteriorhodopsin, halorhodopsin, and channelrhodopsin, function as fast, single-component optogenetic tools in neurons. No avert immune responses are observed on introducing a primitively lower organism’s protein into a highly complex mammalian system; the two separated by a large evolutionary gap, yet, surprisingly, the technique is functioning and successful with a few tweaks of general strategies to enable mammalian expression. These ions can also be used in other excitable cells such as cardiac myocytes and skeletal muscle cells.
Development of this technique involves tunability of different variables. The variability includes many light-sensitive ion channels, dynamics of inactivation and recovery, differing wavelengths of light these ion channels respond to, type of ions allowed to influx and transmit signal, channel conductance, and membrane trafficking. This facilitates high temporal precision.
Applications of Optogenetics
Optogenetics has opened up new possibilities in the study of biology, for health, disease, and therapeutics. Optogenetics illuminates brain function and neurophysiology. Optogenetics led to understanding neuropsychiatric disorders enormously in a short span of time. Examples of applications are to study basic neural circuits, including the innate escape response, to understand more complex circuits that are involved in anxiety, depression, and drug addiction and also to study the proboscis extension reflex in butterflies. Optogenetics is also applied to restore vision. After the loss of photoreceptor cells in retinal degenerative diseases, optogenetic tools impart light sensitivity to the retina, restoring responses in retinal neurons; this has led to the demonstration of visually guided behaviors in animal models.
The single-component control of freely moving mammals through optogenetics came in 2007, with the advent of fiber optics interface. The readouts were obtained using “a fiber optic cable integrated with a tungsten electrode called an optrode.” Many strategies have been devised by synergistically integrating optogenetics with complementary technologies.
“The microbial opsin approach is heir to a long tradition of using light as an intervention in biology”.
Fundamentally, optogenetics relies on light technology. The laser development and its application in optogenetics are well-suited with the availability of synchronized dual-wavelength pulses. These lasers make a perfect source for techniques like coherent anti-Stokes Raman scattering and stimulated Raman scattering. A large band spanning ~900 nm to 1350 nm allows for CARS spectra from 500 cm-1 to 3500 cm-1 covering almost entire biological Raman window. Using two-photon imaging in optogenetics, simultaneous stimuli and signal capture are done. Multi-Photon Microscopy is often used in real-time imaging of neural activity in living organisms. With optogenetics, multi-photon microscopy allows to manipulate and observe the dynamic responses in the neurons. It also allows ‘the ability of spectral unmixing techniques to reveal an intimate relationship of spatial resolution with biomolecular specificity’. Integrating optogenetics with quantitative volumetric Raman Imaging (qVRI) allows three-dimensional visualization of cells with fine details, identification of specific endogenous biomolecules, and spatial observation of complex biological processes such as differentiation. This technique was demonstrated in a Nature paper published in 2017. The tools together will empower the imaging and analysis techniques, paving an understanding of the complex biological processes.
References
- Optogenetics. Deisseroth, K. (2010), Nature Methods
- Mixing fiber optics with genetics has created a revolutionary tool for studying the brain, Bruce Goldman
- Integration of optogenetics with complementary methodologies in systems neuroscience
- The Development and Application of Optogenetics, Lief Fenno, Ofer Yizhar, and Karl Deisseroth, Annual Review of Neuroscience, 34(1), 389–412
- Optogenetic Approaches to Restoring Vision, Annual Review of Vision Science, Vol. 1:185-210
- Neurophotonics and Biomedical Spectroscopy
- Quantitative volumetric Raman imaging of three dimensional cell cultures, Nature Communications, 2017
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