基于上转换纳米粒子的低损伤神经界面技术

Optogenetics is a neuromodulation technology that combines light control technology with genetic technology, thus allowing the selective activation and inhibition of the electrical activity in specific types of neurons with millisecond time resolution. Over the past several years, optogenetics has become a powerful tool for understanding the organization and functions of neural circuits, and it holds great promise to treat neurological disorders. To date, the excitation wavelengths of commonly employed opsins in optogenetics are located in the visible spectrum. This poses a serious limitation for neural activity regulation because the intense absorption and scattering of visible light by tissues lead to the loss of excitation light energy and also cause tissue heating. To regulate the activity of neurons in deep brain regions, it is necessary to implant optical fibers or optoelectronic devices into target brain areas, which however can induce severe tissue damage. Non- or minimally-invasive remote control technologies that can manipulate neural activity have been highly desirable in neuroscience research. Upconversion nanoparticles (UCNPs) can emit light with a short wavelength and high frequency upon excitation by light with a long wavelength and low frequency. Therefore, UCNPs can convert low-frequency near-infrared (NIR) light into high-frequency visible light for the activation of light-sensitive proteins, thus indirectly realizing the NIR optogenetic system. Because NIR light has a large tissue penetration depth, UCNP-mediated optogenetics has attracted significant interest for deep-tissue neuromodulation. However, in UCNP-mediated in vivo optogenetic experiments, as the up-conversion efficiency of UCNPs is low, it is generally necessary to apply high-power NIR light to obtain up-converted fluorescence with energy high enough to activate a photosensitive protein. High-power NIR light can cause thermal damage to tissues, which seriously restricts the applications of UCNPs in optogenetic technology. Therefore, the exploration of strategies to increase the up-conversion efficiency, fluorescence intensity, and biocompatibility of UCNPs is of great significance to their wide applications in optogenetic systems. This review summarizes recent developments and challenges in UCNP-mediated optogenetics for deep-brain neuromodulation. We firstly discuss the correspondence between the parameters of UCNPs and employed opsins in optogenetic experiments, which mainly include excitation wavelengths, emission wavelengths, and luminescent lifetimes. Thereafter, we introduce the methods to enhance the conversion efficiency of UCNPs, including optimizing the structure of UCNPs and modifying the organic dyes in UCNPs. In addition, we also discuss the future opportunities in combining UCNP-mediated optogenetics with flexible microelectrode technology for the long-term detection and regulation of neural activity in the case of minimal injury.     

植入式光电极器件发展

Optogenetics transforms specific types of neurons through genetic engineering to achieve the cell membrane expression of photosensitive channel protein. When a specific wavelength of light irradiates the photosensitive channel protein, the cell is either excited or inhibited. Optogenetics provides a precise and fast method to control the activity of individual neurons for neuroscience research, which has gained increasing attention as a means of neural regulation. To realize the photogenetic regulation of neurons, light should be introduced into the brain safely and efficiently. Thus, specialized photoelectric devices are needed. Optrode plays a significant role in the application of optogenetics tools, which is the technical basis for the application of optogenetics. Optrode is a kind of implantable neural interface device. It can introduce light into the brain to regulate neural activity and record the changes of neural electrical signals under the control of lights. As the research of optogenetic technology continues, More and more optrodes are being developed and applied in the study of neuroscience and diseases, such as neural circuit, cognition and memory, epilepsy, and sensory function damage. The combination of optrode with optogenetic technologies provides various developmental modes in terms of material selection, device structure, light supply method, and integrated ways. The difficulty in fabricating optrodes lies in performing light stimulation and electrical signal recording without causing the immune rejection of the test animal and affecting its normal physiological activities simultaneously. In this study, based on structural characteristics and manufacturing process, optrodes are classified into two categories: waveguide-based and micro-light emitting diode-based. Subsequently, based on manufacturing process and light supply method, waveguide-based optrodes are further divided into optical fiber-optrode, optical waveguide-optrode based on MENS technology, and LD/LED waveguide-optrode. Similarly, micro-light emitting diode-based optrodes are divided into hard μLED optrode and soft μLED optrode. The advantages and disadvantages of different types of optrodes, as well as the evolution direction, are reviewed and summarized. Additionally, problems with existing optrodes, such as signal quality, biocompatibility, and device reliability, are discussed. Further, the ideal form of the device is presented as possessing the following characteristics: μLED and recording electrode integrated on flexible substrate, small size, high spatial resolution, high biocompatibility, wireless energy supply, wireless data transmission, etc. As optrode technologies are continuously updated, in the application of optogenetic technologies, research on brain neural circuit and functional structure will be better studied, and various nerve diseases will be gradually tamed.     

Chemical Physics in Living Cells-using Light to Visualize and Control Intracellular Signal Transduction?

Cells are crowded microenvironments filled with macromolecules undergoing constant physical and chemical interactions. The physicochemical makeup of the cells affects various cellular responses, determines cell-cell interactions and influences cell decisions. Chemical and physical properties differ between cells and within cells. Moreover, these properties are subject to dynamic changes in response to environmental signals, which often demand adjustments in the chemical or physical states of intracellular molecules. Indeed, cellular responses such as gene expression rely on the faithful relay of information from the outside to the inside of the cell, a process termed signal transduction. The signal often traverses a complex path across subcellular spaces with variable physical chemistry, sometimes even influencing it. Understanding the molecular states of such signaling molecules and their intracellular environments is vital to our understanding of the cell. Exploring such intricate spaces is possible today largely because of experimental and theoretical tools. Here, we focus on one tool that is commonly used in chemical physics studies-light. We summarize recent work which uses light to both visualize the cellular environment and also control intracellular processes along the axis of signal transduction. We highlight recent accomplishments in optical microscopy and optogenetics, an emerging experimental strategy which utilizes light to control the molecular processes in live cells. We believe that optogenetics lends unprecedented spatiotemporal precision to the manipulation of physicochemical properties in biological contexts. We hope to use this work to demonstrate new opportunities for chemical physicists who are interested in pursuing biological and biomedical questions.     

活细胞中的化学物理-光学观测和控制细胞内信号转导

Cells are crowded microenvironments filled with macromolecules undergoing constant physical and chemical interactions. The physicochemical makeup of the cells affects various cellular responses, determines cell-cell interactions and influences cell decisions. Chemical and physical properties differ between cells and within cells. Moreover, these properties are subject to dynamic changes in response to environmental signals, which often demand adjustments in the chemical or physical states of intracellular molecules. Indeed, cellular responses such as gene expression rely on the faithful relay of information from the outside to the inside of the cell, a process termed signal transduction. The signal often traverses a complex path across subcellular spaces with variable physical chemistry, sometimes even influencing it. Understanding the molecular states of such signaling molecules and their intracellular environments is vital to our understanding of the cell. Exploring such intricate spaces is possible today largely because of experimental and theoretical tools. Here, we focus on one tool that is commonly used in chemical physics studies-light. We summarize recent work which uses light to both visualize the cellular environment and also control intracellular processes along the axis of signal transduction. We highlight recent accomplishments in optical microscopy and optogenetics, an emerging experimental strategy which utilizes light to control the molecular processes in live cells. We believe that optogenetics lends unprecedented spatiotemporal precision to the manipulation of physicochemical properties in biological contexts. We hope to use this work to demonstrate new opportunities for chemical physicists who are interested in pursuing biological and biomedical questions.     

Strategies for development of optogenetic systems and their applications

It has become clear that biological processes are highly dynamic and heterogeneous within and among cells. Conventional analytical tools and chemical or genetic manipulations are unsuitable for dissecting the role of their spatiotemporally dynamic nature. Recently, optical control of biomolecular signaling, a technology called “optogenetics,” has gained much attention. The technique has enabled spatial and temporal regulation of specific signaling pathways both in vitro and in vivo. This review presents strategies for optogenetic systems development and application for biological research. Combinations with other technologies and future perspectives are also discussed herein. Although many optogenetic approaches are designed to modulate ion channel conductivity, we mainly examine systems that target other biomolecular reactions such as gene expression, protein translocations, and kinase or receptor signaling pathways.     

Quantification of riboflavin,flavin mononucleotide,and flavin adenine dinucleotide in mammalian model cells by CE with LED‐induced fluorescence detection

Cultured mammalian cells essential are model systems in basic biology research, production platforms of proteins for medical use, and testbeds in synthetic biology. Flavin cofactors, in particular flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD), are critical for cellular redox reactions and sense light in naturally occurring photoreceptors and optogenetic tools. Here, we quantified flavin contents of commonly used mammalian cell lines. We first compared three procedures for extraction of free and noncovalently protein‐bound flavins and verified extraction using fluorescence spectroscopy. For separation, two CE methods with different BGEs were established, and detection was performed by LED‐induced fluorescence with limit of detections (LODs 0.5–3.8 nM). We found that riboflavin (RF), FMN, and FAD contents varied significantly between cell lines. RF (3.1–14 amol/cell) and FAD (2.2–17.0 amol/cell) were the predominant flavins, while FMN (0.46–3.4 amol/cell) was found at markedly lower levels. Observed flavin contents agree with those previously extracted from mammalian tissues, yet reduced forms of RF were detected that were not described previously. Quantification of flavins in mammalian cell lines will allow a better understanding of cellular redox reactions and optogenetic tools.     

轻掺杂硅基神经电极的光噪声消减

Silicon-based neural probes are practical tools for recording neural cell firing. A single silicon-based needle with a width of only 70 μm, prepared using the standard complementary metal-oxide-semiconductor (CMOS) process technology, can contain thousands of electrode-recording sites. Optogenetics has made control over neuronal activity more precise. By recording the electrical activity of neurons stimulated by light, more information about brain activity can be recorded and analyzed. When yellow light or blue light is used to stimulate neurons, the photon energy is greater than the forbidden bandwidth of the silicon substrate, and the valence-band electrons are excited to the conduction band, generating electron-hole pairs. The photoinduced carrier in the silicon substrate severely disrupts the probe's signal-to-noise ratio. Decreasing the disturbance caused by light is a pragmatic way to execute recording and stimulating simultaneously. The traditional noise reduction method involves using heavily doped silicon as the substrate, reducing the carrier life by increasing the impurity concentration, and then reducing the noise of the silicon electrode under illumination. However, the heavily doped silicon substrate has more lattice defects than its lightly doped counterparts, which makes the silicon electrode fragile, and this method is not compatible with the standard CMOS process technology. On analyzing the photoinduced noise mechanism of manufacturing electrodes on lightly doped silicon substrates, we found that the inhomogeneous distribution of carriers generated by light excitation polarizes lightly doped silicon substrates. The potential caused by photoinduced polarization will affect the electrodes fabricated on it. Metalizing and grounding the lightly doped silicon substrate will effectively decrease the polarization potential. On using this method, the noise amplitude caused by the illumination can drop to 0.87% of the original value. To ensure an appropriate firing rate of neurons, the photo-stimulation frequency was chosen to be 20 Hz. Under the illumination of 1 mW·mm^-2, the background noise of the electrode could be controlled below 45 μV, which meets the needs for general optogenetics applications. Modification of the lightly doped silicon substrate will meet the requirements of the neural electrode for optogenetics applications. Unlike the traditional method of reducing light-induced noise by heavily doping the entire substrate, the noise reduction method of lightly doped silicon substrate is compatible with the standard CMOS process technology. It provides a noise cancellation method for the preparation of silicon-based neural microelectrodes with dense recording sites and high channel count using standard CMOS processes.