Designer Retinal Circuits: Interfacing optical tweezers with an electronic device

描述(申请人提供)：我们的目标是研究视网膜回路的发展和重建：1)视网膜移植后必须形成的回路，如果要恢复一些视力，2)在视网膜重塑时在退化的视网膜中发生的回路，以及3)在正常视网膜中起作用的回路。我们建议通过将光学镊子技术与玻璃芯片上的微电极阵列相结合来实现这一点。芯片将被设计为具有非粘附区和粘附区，分别用于细胞运动和细胞培养。使用光学镊子，分离的成人视网膜细胞通过其独特的形态被显微操作到电极上，并允许形成细胞与细胞的接触。突触的发育将通过免疫细胞化学和突触小泡回收染料进行监测。突触活性将通过电极刺激和对双极细胞和一些无长突细胞的无毒电压敏感染料的成像，以及通过记录无长突细胞和神经节细胞的尖峰和动作电位的电极来测试。使用镊子可以精确控制所建造的电路。随着时间的推移，设计并在功能上测试特定神经细胞相互作用的能力将为系统提供前所未有的实验控制水平。

DESCRIPTION (provided by applicant): Our goals are to examine the development and reconstruction of retinal circuits 1) that must form after retinal transplantation if some vision is to be restored, 2) that occur in the degenerate retina as the retina is remodeled, and 3) that function in the normal retina. We propose to do this by combining the technologies of optical tweezers with microelectrode arrays on glass chips. Chips will be designed to have non-adherent and adherent zones for cell movement and cell culture respectively. Using the optical tweezers, isolated adult retinal cells that are identified by their distinctive morphologies will be micromanipulated onto electrodes and allowed to form cell-cell contacts. Synaptic development will be monitored by immunocytochemistry and synaptic vesicle recycling dyes. Synaptic activity will be tested by electrode stimulation and imaging of non- toxic voltage-sensitive dyes for bipolar and some amacrine cells, and by recording electrodes for the spiking and action potentials of amacrine and ganglion cells. The use of the tweezers allows precise control over the circuits that are built. The ability to design and then functionally test specific nerve cell interactions over time will provide a system with an unprecedented level of experimental control.