Optical Stimulation of the Auditory Pathway by Blue-Shifted Photoswitchable Glutamate Receptor Agonists.

光刺激耳中的螺旋神经节神经元(SGN)提供了一种未来的替代电刺激用于人工耳蜗术的方法。神经活动的光遗传操作是基于光敏感蛋白的表达，这需要基因治疗。光遗传学提供了一种替代光遗传学的方法，它对内源性受体起作用，不需要基因操作。在光开关中，由Pau Gorostiza开发的靶向共价光开关(Tcp)主要与离子型海人藻酸受体GluK1反应。在海马神经元上，最好的第一代化合物TCP9在紫外光(380 Nm)下激活天然的GluK1受体，在可见光(500 Nm)下使受体失活。我们通过圆窗将该化合物应用于沙土鼠耳蜗处，对新一代BluTCP1和BluTCP2进行了体内初步研究。经圆窗壁龛电极进行的耳蜗术显示，耳蜗声诱发微音器和复合动作电位(CAP)波幅保持正常，提示在TCPs存在的情况下，耳蜗生理正常，无急性毒性。在用一根光纤(λ=473 nm)进行光刺激时，我们观察到光诱发的帽(OCAP)。光脉冲辐射通量小于3 mW时可诱发OCAP，80µS光脉冲的OCAP幅度最大，重复频率可达1 kHz。这一性能使这些化合物成为刺激光学SGN的有趣工具。该项目的目的是在开发光学耳蜗植入物的背景下，使用BlueTCP1和BlueTCP2建立和表征对SGN的体内光药理刺激。主要涉及两个方面：i)SGN对BluTCP/光刺激的反应的生理学表征，以确定这两种化合物中最合适的。；ii)在耳聋模型中测试BluTcp用于光学听力恢复的翻译潜力，该模型配备了耦合到光学人工耳蜗的慢性药物输送系统。

Light stimulation of spiral ganglion neurons (SGNs) in the ear provides a future alternative to electrical stimulation used in cochlear implants. Optogenetic manipulation of neuronal activity is based on the expression of light-sensitive proteins, which requires gene therapy. An alternative to optogenetics is offered by photopharmacology which operates on endogenous receptors and does not require genetic manipulation. Among the “photoswitches”, the Targeted Covalent Photoswitches (TCP) developed by Pau Gorostiza, mainly reacts with the ionotropic kainate receptor GluK1. It was previously shown in vitro on hippocampal neurons that TCP9, the best first-generation compound, activates native GluK1 receptor upon ultraviolet light (380 nm) and a deactivates the receptor upon visible light (500 nm). We performed a preliminary study of a new generation of blueTCP1 and blueTCP2 in vivo by applying the compound to the gerbil cochlea via the round window. Electrocochleography via a round window niche electrode showed a preservation of acoustically-evoked cochlear microphonic and a compound action potential (CAP) amplitude, suggesting a normal cochlear physiology in the presence of TCPs and an absence of acute toxicity. Upon light stimulation using an optical fiber (λ = 473 nm), we observed optically evoked CAPs (oCAPs). oCAPs could be evoked by light pulse radiant flux as low than 3 mW, oCAP amplitudes were maximum in response to 80 µs light pulse and were sizable up to a repetition rate of 1 kHz. This performance makes these compounds interesting tools for optical SGN stimulation. The project aims to establish and characterize in vivo photopharmacological stimulation of SGNs using the blueTCP1 and blueTCP2 in the context of developing the optical cochlear implant. Two main aspects will be addressed: i) physiological characterization of SGN responses to blueTCP/light stimulation to identify the most suitable of the two compounds.; ii) testing the translational potential of blueTCP for optical hearing restoration in a deafness model fitted with a chronic drug delivery system coupled to an optical cochlear implant.