In Vivo Activation Of Neurons Based On X-ray Excitated Downconversion Nanomaterial

Temporally precise,noninvasive and remote control of activity in specific types of neuronal populations is a long-term goal in neuroscience.Optogenetic techniques can accurately control genetically targeted neuronal activity over millisecond-timescale through light-gated ion channel.However,there are still a few defects in the conventional optogenetic techniques.For example,inserting of the invasive optical fiber may damage the organization and behavior of the experimental object.Besides,optical fiber is difficult to be used for the optical stimulation of peripheral neurons limited by fixed conditions.Another optogenetic technique is based on red light excitation,which is limited by the ability to penetration and the excitation depth is shallow.Other optogenetic technique is near infrared optogenetics,which is based on upconversion nanomaterials and near-infrared light excitation（upconversion optogenetics）,but it is limited by the large absorption of water in tissues and results in low luminescence efficiency and overheating phenomenon.Moreover,it is a great challenge to have simultaneous neuron stimulations in different brain regions with the use of the fixed fiber or near-infrared laser.Inspired by the recent research in upconversion optogenetics,this thesis proposes a novel strategy for neuron activation based on X-ray excited downconversion nanomaterials（DCNP）,in which X-ray is converted into visible light by DCNP to activate channelrhodopsin and neurons.Based on emission spectrum well-matched with the activation spectra of the optogenetic tools,terbium（III）(Tb³⁺)doped DCNP were adopted to activate channelrhodopsin.The effects of nanomaterial performance parameters(such as the shape and Tb³⁺doping concentration)and excitation parameters of X-ray（such as the tube current and voltage）on the luminescence intensity of nanomaterials were analyzed and tested.All of these provides screening basis for optimal luminescence performance of Tb³⁺doped DCNP and its optimal X-ray excitation conditions.Based on these results,an experimental method for X-ray activated neurons based on DCNP has been designed.Finally,this thesis combines the recombinant AAV virus and transgenic mice to mediate dopaminergic neurons in VTA and GABAergic neurons in the cerebral cortex to express channelrhodopsin,and uses the expression of c-Fos protein as a marker of neuronal activation after X-ray excites DCNP.The experimental results revealed that the percentage of c-Fos-positive neurons has risen significantly in both the VTA brain region of TH-Cre mice and the cerebral cortex of Vgat-Cre mice after the function of X-ray excites DCNP system.This provides experimental support for the method that X-ray activates neurons based on DCNP.