Progress in Optogenetics Research

lightheredityOptogenetics is a rapidly developing field that integrates opticssoftwareInterdisciplinary bioengineering techniques such as control, gene manipulation, and electrophysiology. The main principle is to first use gene manipulation techniques to transfer photosensitive genes (such as ChR2, eBR, NaHR3.0, Arch or OptoXR, etc.) into specific types of cells in the nervous system for the expression of special ion channels or GPCRs. Light sensitive ion channels selectively stimulate the passage of cations or anions under different wavelengths of light, resulting in changes in membrane potential on both sides of the cell membrane, achieving the goal of selectively exciting or inhibiting cells.

Optogenetic technology has two major characteristics: high spatiotemporal resolution and cell type specificity, overcoming many shortcomings of traditional methods for controlling cell or organism activity. It can perform non-invasive localization and stimulation operations on neurons, which has changed the research status in the field of neuroscience and provided revolutionary research methods for neuroscience. lightheredityTechnology may also develop new therapies for a range of central nervous system diseases in the future.

lightheredityThe application of learning technology has developed rapidly since 2010, covering multiple classic experimental animal species such as fruit flies, nematodes, mice, rats, macaques, and crab eating monkeys, and involving multiple aspects of neuroscience research, including neural circuitsbasic researchLearning and memory research, addiction research, movement disorders, sleep disorders, Parkinson's disease modelsdepressionAnd applications such as animal models of anxiety disorders.

In the process of applying optogenetics technology, scientists first need to search for suitable photosensitive proteins; Secondly, corresponding genetic information transmission is carried out through transfection, viral transductionGenetically modified animalsThe establishment of systems and other methods will enhance the photosensitivity of photosensitive proteinsheredityInformation is transmitted to the target cell; Subsequently, scientists utilized controllability demonstrations to demonstrate cellular activity by controlling the specificity of the demonstration light in time and space; After reading the research results, researchers can use electrodes to measure the fluorescence effect changes of photosensitive proteins by detecting the voltage inside and outside the cell membrane, and can use fluorescent biosensors to detect the readout values of different cells, and then evaluate the impact of adjusting cell activity on the entire animal through behavioral testing.

Based on this, regarding lightheredityI will take stock of the recent progress made in learning technology for readers.

1. Cell: Significant progress! Expanding the optogenetics toolkit by flipping rhodopsin
doi:10.1016/j.cell.2018.09.026

In a new study, researchers from institutions such as the Howard Hughes Medical Institute in the United States have discovered a novel method for modifying a class of photosensitive proteins called rhodopsin. By flipping these proteins in the cell membrane, they can produce tools with different characteristics. The relevant research results were published online in the Cell journal on October 18, 2018, under the title "Expanding the Optogenetics Toolkit by Topological Inversion of Rhodopsin". The corresponding authors of the paper are Joshua Dudman and Alla Karpova from the Howard Hughes Medical Institute. This technology can be used for lightheredityThe number of proteins in optogenetics technology - a technique that uses light to manipulate neuronal activity - doubles. These newly developed hybrid rhodopsin proteins enable these researchers to conduct new experiments that help analyze brain circuits and study the neuroscience behind the treatment of Parkinson's disease.

Inspired by evolution, under the leadership of Jennifer Brown, Reza Behnam, Luke Coddington, and Gowan Tervo, these researchers have developed a complementary technique for modifying new rhodopsin. In addition to mutations, the existence of recombination - which combines protein domains with different functions through gene reshuffling - also leads to protein diversity in nature. Scientists believe that recombination is crucial for the emergence of a small subset of proteins whose orientation in the cell membrane has changed through evolution.

When these researchers simulated recombination by adding a new protein to one end of a rhodopsin, it flipped. This is really unexpected. If every existing modified or newly discovered rhodopsin can acquire new functions when flipped, it may lead to the use of lightheredityDoubling the protein tools in learning technology. They can not only change the orientation of proteins in the cell membrane, but also discover that these newly modified rhodopsin have * and useful new functions. One of them, called FLInChR (Full Length Inversion of ChR), initially activates neurons in rhodopsin. When flipping occurs, it becomes a potent and rapid inhibitor that can be used to conduct new experiments.

2. A major breakthrough in optogenetics! Upconversion nanoparticles assist in deep brain stimulation! Or it may overturn the treatment of neurological diseases!
doi:10.1126/science.aaq1144; doi:10.1126/science.aar7379

Thomas McHugh, the head of the Brain Science Research Institute at RIKEN in Japan, and his colleagues have now found a new method to non invasively introduce light into the depths of the brain. In their article published yesterday in Science, they used upconversionnanoUCNPs introduce laser into the deep part of the skull. this kind ofnanoParticles can withstand traditional lightheredityLearn to absorb near-infrared light at depths that cannot be achieved and convert them into visible light. This method is used to activate neurons in different regions of the brain, silence epilepsy, and activate memory cells. Nanoparticles can effectively extend the depth that our optical fibers can reach, enabling remote delivery of light and achieving non-invasive therapy, "McHugh said.

In addition to activating neurons, UCNPs can also be used to suppress the condition of epileptic mice. Researchers injected green emitting nanoparticles into the hippocampus of mice and then irradiated the skull surface with near-infrared light. As a result, the epileptic neurons of these mice were effectively silenced. In another region called the medial septal nucleus, the light emitted by nanoparticles promotes the synchronization of neuronal theta cycles (an important type of brainwave). In mice with fear memories, researchers successfully triggered fear memories in the hippocampus using luminescent UCNPs. These neuronal activation, inhibition, and memory activation effects were only observed in mice injected with nanoparticles.

3. Science: Boosting the development of optogenetics! Analyze the three-dimensional structure of rhodopsin channel protein 2
doi:10.1126/science.aan8862; Download:10.1126/science.aar2299

Channel rhodopsin 2 (ChR2) is a membrane protein widely used in optogenetics technology. lightheredityLearning technology is a relatively new technique that involves using light to manipulate neurons and muscle cells in living organisms. Similar methods have been used to partially reverse hearing/vision loss and control muscle contractions.

To reveal the structure of ChR2, researchers from Germany, France, Russia, and the Czech Republic used an analytical technique called X-ray diffraction. This technique is only used for analyzing protein samples that exist in crystalline form. They cultured ChR2 protein crystals in a so-called cubic lipid interphase that allows proteins to move freely without leaving the membrane. They used X-rays with a wavelength of approximately 1 angstrom to irradiate their cultured ChR2 protein crystals, and successfully resolved the structure of ChR2 protein by analyzing the diffraction pattern of X-rays in these protein crystals. The relevant research results were published in the Science journal on November 24, 2017, with the paper titled "Structural Insights into ion Conduct by channelrhodopsin 2".

4. Hippocampus: "Optogenetics" therapy may be able to restore memory in some Alzheimer's patients
doi:10.1002/hipo.22756

Recently, researchers from Columbia University published an article in the Hippocampus journal stating that through lightheredityLearning methods can restore the memory of mice with Alzheimer's disease. This discovery may change our understanding of this disease.

Firstly, the author genetically modified mice to emit light colored fluorescence when storing memories and red fluorescence when reacquiring memories. Afterwards, the author accepted itheredityThe modified wild-type mice and Alzheimer's disease mice were stimulated with lemon scent, followed by electrical stimulation, to establish a correlation between these two memories. A week later, the author once again stimulated these mice with the scent of lemon. The results showed that wild-type mice were able to exhibit both yellow and red fluorescence simultaneously, and showed signs of fear, indicating that they also underwent memory recall while forming memories. However, the glowing areas in the brains of Alzheimer's mice are significantly different, indicating that their brains are disrupted during the process of memory retrieval. Afterwards, the researchers stimulated the brains of mice with a beam of blue light, which reactivated their memory of lemon scent and electrical stimulation. As a result, the mice trembled when they smelled the scent again.

5. Cell: Optogenetics turns mice into killers
doi:10.1016/j.cell.2016.12.027

Every mouse hides a killer inside its body. Researchers have identified the region in the brain that controls hunting behavior and have successfully found a way to control its on/off state. Dr. Ivan de Araujo and colleagues from Yale University have discovered two sets of nerves in the brain that control hunting behavior in mice. One group coordinates the pursuit of prey, while the other controls muscles in the neck and jaw. Both sets of nerves are located in the amygdala, which is the area of the human body involved in movement, emotion, and fear.

By modifying these nerves, they can activate them through laser, a technique called lightheredityThe research team is therefore able to control the switching of these pathways at any time. When the laser is turned off, the mice walk around the cage normally. Once the laser is turned on, the mice will crazily attack anything on their way: live crickets, fake insects, even tree branches or bottle caps. They will pounce on their prey, grab them with their claws and repeatedly bite them.

Afterwards, the researchers attempted to suppress the function of each group of nerves separately. When they suppressed the nerves responsible for chasing prey, the mice chased slower but still bit; On the other hand, if the nerve responsible for tearing is suppressed, mice will chase prey but not bite.

6. Oncotarget: Using optogenetics to control tumor occurrence
doi:10.18632/oncotarget.8036

In a new study, researchers from Tufts University in the United States demonstrated the use of light to control electrical signals between cells, prevent tumor formation, and normalize them after tumor formation based on a frog model. This study reports the use of lightheredityLearn to specifically manipulate bioelectric signals to prevent tumor formation induced by oncogenes and lead to tumor formation induced by oncogenestumorFade. The relevant research results were published online on March 16, 2016 in the Oncotarget journal, with the title "Use of Genetically Encoded, Light Coated Translators to Control Tumorigenesis".

Frogs are a good model organism for cancer basic scientific research because their tumors share many common characteristics with mammals, including rapid cell division, tissue destruction, increased vascular growth, invasiveness, and cells containing abnormal internal positive voltages. Compared to the outside of the cell, almost all healthy cells maintain a larger negative voltage inside the cell; Opening and closing ion channels in the cell membrane can cause the voltage to become more positive (depolarizing the cell) or more negative (polarizing the cell). Under normal conditions in other aspects, utilizetumorThe abnormal bioelectric signal characteristics can detect them.

7. Neuron: Deleting memories? The future may really be possible
doi:10.1016/j.neuron.2014.09.037

Recently, in a research paper published in the journal Neuron, researchers from the Neuroscience Research Center at the University of California, Davis successfully removed special memories from the mouse brain using light. This study may provide some insights into how different parts of the brain work together to restore episodic memory.

lightheredityOptogenetics is a novel technology that uses light to study nerve cells, and in recent years, it has been rapidly adopted by scientists as a standard method for studying brain function. In the article, researcher Kazumasa Tanaka applied this technology to research on memory recovery, among other things. For 40 years, scientists have hypothesized that restoring episodic memory (even in special events that occur in specific locations) involves coordinated activity between the cerebral cortex and the hippocampus. This theory aims to study the patterns of brain activity that reactivate the cerebral cortex and hippocampus during episodic memory recovery, allowing individuals to experience those events again. If the hippocampus is damaged, patients will lose decades of memory.

In the article, researchers utilizeheredityModified mice were used for research, and when mouse nerve cells were activated, they could all emit green fluorescence and express special proteins to promote the closure of nerve cells by light. The researchers trained the mice in cages, where they experienced electroconvulsive therapy. Normally, mice in a new environment would use their sense of smell to adapt to the environment, but when they were electroconvulsive and placed in a new environment, they would experience a fear response.

8. Nature: New hope for optogenetic tools, light driven sodium ion channel KR2 structure resolved
doi:10.1038/nature14322

Japanese scientists have published an academic article in the journal Nature, stating that they have deciphered the structure of the light driven sodium ion channel protein KR2, providing a new generation of light for the futureheredityLearning tools create possibilities.

Many organisms can absorb the energy of light or perceive information from light, relying on a type of rhodopsin molecule. This molecule has a seven alpha helix transmembrane protein (opsin) covalently linked to a retinal molecule. According to the types of visual proteins, they can be divided into animal and microbial visual proteins. The rhodopsin function of microorganisms is different from that of animals, mainly serving as ion channels, ion transporters, photosensitive molecules, or kinases. The rhodopsin of this microorganism is receiving increasing attention. This is because ion channel and ion pump type rhodopsin can be used for neural cell activity in many living organisms, and has become a very powerful light in the field of neuroscienceheredityLearning tools.

It is generally believed that the positively charged hydrogen ions of Schiff bases will be localized in all ion channels of photoion pumps and are believed to prevent anions and neutral molecules from passing through. The analysis of KR2 structure has raised a new question, which is how this ion pump transports sodium ions.

9. Neon: Chicago scientists achieve new breakthrough in optogenetics technology
doi:10.1016/j.neuron.2015.02.033

In recent years, scientists have beenepigeneticsWith the deepening of field research, people have begun to hope to control the state of cells, especially neurons, in the body through in vitro stimulation. This field has broad application prospects, such as treating genetic diseases such as macular degeneration. Based on this, lightheredityDisciplines such as learning have been established one after another. However, so far, in order to achieve this goal, researchers have had to genetically modify neurons. This also greatly hinders the popularization of this technology.

Researchers from the University of Chicago and the University of Illinois at Chicago have recently achieved breakthroughs in this field. Researchers use the heat generated by far-infrared light to control the normal life activities of normal neuronal cells. Unlike the previous practice of expressing photosensitive proteins in normal neuronal cells, scientists from Chicago have chosen to use gold nanoparticles to locate specific neurons. To address the issue of specificity of gold nanoparticles, researchers attached a scorpion neurotoxin Ts1 to the gold nanoparticles. Ts1 can target the recognition of neuronal cells by identifying sodium ion channels on their surface. This is also why humans are not modifying neuronsheredityBased on the characteristics, achieve the goal of light controlled neuronal activity. However, this study is still in its early stages, and researchers also suggest that Ts1 may be toxic to neuronal cells. This research work was published in the journal Neuron.

10. Nat Methods: Cell targeting controlled by photosensitizers
doi:10.1038/nmeth.3735

In a research report published in the journal Nature Methods, researchers from Carnegie Mellon University in the United States provided insights into lightheredityThe photosensitizer in the school has redesigned and developed a fluorescent probe for controlling cells. This technology may help understand the role of specific cells and proteins in disease development, and also provide hope for the development of targeted therapies for cancer or other diseases in the later stage.

Optogenetics is the use of light to control biological processes in the body. Researchers typically achieve light controlled cell processes by reprogramming light activated components into the genetic code of the organism. When these components are exposed to light, they promote different functions of certain body tissues. Researchers have spent nearly 10 years developing fluorescent probes called fluorescein activating proteins (FAPs), which are commonly used to monitor protein activity in live cells in real-time. FAPs are a type of fluorescent probe that typically activates proteins in cellshereditySexually expressed, when a fluorescent dye with the same name as fluorescein comes into contact, the complex will emit light, allowing scientists to observe and track it.

Researcher Bruchez said that in order to turn on the lightheredityIn the article, we engineered fluorescent dyes to not only emit light, but also produce singlet oxygen, which is a toxic form of oxygen. When it binds to fluorophore activating proteins and is exposed to light, the targeted fluorophore activating protein and photosensitizer activation method (FAP-TAPS) allows scientists to clearly see the labeled proteins and selectively inhibit their activity. (Biological Valley)