Optoelectronics and Biomedical Photonics Group, AIPT, Aston University, Birmingham, United Kingdom.
Optoelectronics and Measurement Techniques, University of Oulu, Oulu, Finland; Cell Physiology and Pathology Laboratory, Orel State University, Orel, Russia.
Biophys J. 2021 Mar 2;120(5):964-974. doi: 10.1016/j.bpj.2021.01.028. Epub 2021 Feb 3.
In nature, sensory photoreceptors underlie diverse spatiotemporally precise and generally reversible biological responses to light. Photoreceptors also serve as genetically encoded agents in optogenetics to control by light organismal state and behavior. Phytochromes represent a superfamily of photoreceptors that transition between states absorbing red light (Pr) and far-red light (Pfr), thus expanding the spectral range of optogenetics to the near-infrared range. Although light of these colors exhibits superior penetration of soft tissue, the transmission through bone and skull is poor. To overcome this fundamental challenge, we explore the activation of a bacterial phytochrome by a femtosecond laser emitting in the 1 μm wavelength range. Quantum chemical calculations predict that bacterial phytochromes possess substantial two-photon absorption cross sections. In line with this notion, we demonstrate that the photoreversible Pr ↔ Pfr conversion is driven by two-photon absorption at wavelengths between 1170 and 1450 nm. The Pfr yield was highest for wavelengths between 1170 and 1280 nm and rapidly plummeted beyond 1300 nm. By combining two-photon activation with bacterial phytochromes, we lay the foundation for enhanced spatial resolution in optogenetics and unprecedented penetration through bone, skull, and soft tissue.
在自然界中，感光受体是对光产生各种时空精确且通常可逆的生物学反应的基础。感光受体也是光遗传学中用于通过光控制生物体状态和行为的基因编码剂。光敏色素是一类光受体，它们在吸收红光（Pr）和远红光（Pfr）之间转换状态，从而将光遗传学的光谱范围扩展到近红外范围。尽管这些颜色的光在软组织中的穿透性更强，但在骨骼和头骨中的传输效果较差。为了克服这一基本挑战，我们探索了利用发射 1 微米波长的飞秒激光来激活细菌光敏色素。量子化学计算预测，细菌光敏色素具有相当大的双光子吸收截面。这一概念与我们的实验结果一致，我们证明了在 1170 到 1450 纳米波长范围内的双光子吸收可以驱动光可逆的 Pr ↔ Pfr 转换。在 1170 到 1280 纳米波长之间，Pfr 的产量最高，而超过 1300 纳米时则迅速下降。通过将双光子激活与细菌光敏色素相结合，我们为光遗传学中增强空间分辨率和前所未有的穿透骨骼、头骨和软组织奠定了基础。
