Wireless Optogenetics by relay nano-illuminators

描述(由申请人提供)：由于光遗传工具的突破性发展，最近朝着更好地理解神经电路功能迈出了重要的一步。在这种方法中，微生物视蛋白基因[最值得注意的是通道视紫红质(chr-2)和卤视紫质(NpHR)]通过病毒转导或转基因在神经元中表达。然后，表达视蛋白的神经元可以被特定波长的光激活或抑制。由于其巨大的时空分辨率，光遗传学能够从功能上剖析大脑回路，并为大脑之间的因果关系提供新的见解。 活动和行为，并可能导致神经精神疾病的治疗。然而，由于激活光遗传结构所需波长的光在组织中的穿透性有限，对行为动物的刺激必须依赖长期植入的光纤或安装的LED来将光传输到深部脑组织。虽然这种方法非常有用，并产生了大量关于大脑回路的信息，但通过光纤进行刺激也有重要的局限性，特别是在清醒动物的慢性刺激方面。为了解决这一具有挑战性的问题，我们建议开发一种无线光遗传策略，使用继电纳米照明器远程激活活体中的opsins。这一方法是建立在我们实验室最近在掺镧上转换纳米颗粒(UCNPs)方面取得的关键技术进步的基础上的，UCNPs是具有意想不到的性质的新一代纳米颗粒。UCNP最显著的优点是其不自然的反向激发和发射轮廓；即，它们使用生物兼容的、低功率、深组织穿透、近红外辐射来激发，这些辐射在不同的频率下有效地转换为更高的能量输出发射 波长更短，包括激活视蛋白的可见光。我们提出了两个具体目标。对于目标1，我们将描述UNCPs在体外和体内作为“接力照明器”的能力。我们首先将重点放在第一代UCNP纳米粒子(CaF2包裹的核/壳UCNP)，我们的初步实验已经证明，显示出强大的发射从深部脑组织。在AIM2中，我们将开发新的组合合成来提高稀土掺杂UCNP的光遗传性能。这一新战略将克服当前基于光纤的方法的许多局限性，并将在基础科学和人类健康方面实现新的应用。

DESCRIPTION (provided by applicant): An important step toward better understanding neural circuit function was recently made possible thanks to the breakthrough development of optogenetic tools. In this approach, the microbial opsin genes [most notable, Channelrhodopsin (ChR-2) and halorhodopsin (NpHR)] are expressed in neurons either by viral transduction or transgenesis. Neurons expressing opsin can then be activated or inhibited by light at specific wavelengths. Due to its great spatiotemporal resolution, optogenetics is able to functionally dissect brain circuits, as well as to offer new insights into the causal relationship between brain activity and behavior and, possibly, lead to therapies for neuropsychiatric diseases. However, due to the limited tissue penetration of light at the wavelengths necessary to activate optogenetic constructs, the stimulation of behaving animals has to rely on chronically implanted, fiber-optics or mounted LEDs to deliver light into deep brain tissues. Although this method is very useful and has yielded a wealth of information about brain circuits, stimulation via fiber-optics also has important limitations, particularly in regard to chronic stimulation in awake animals. To address this challenging issue, we propose to develop a wireless optogenetic strategy to remotely activate opsins in vivo using a relay nano- illuminator. This approach is buil upon key technologic advances recently made in our laboratory in lanthanide-doped upconversion nanoparticles (UCNPs), a new generation of nanoparticles with unexpected properties. The most significant advantage of UCNPs is their unnatural inverse excitation and emission profiles; i.e., they are excited using biocompatible, low power, deep tissue-penetrant, near infrared radiation that is effectively converted to a higher energy output emission at various shorter wavelengths, including visible light for activation of opsins. We propose two specific aims. For Aim 1, we will characterize the ability of UNCPs to act as "relay illuminators" in vitro and in vivo. We will initially focus on the first generation of UCNP nanoparticles (CaF2 coated core/shell UCNP) that our preliminary experiments have demonstrated to exhibit robust emission from deep brain tissue. In Aim2, we will develop novel combinatorial synthesis to enhance optogenetic performance of lanthanide-doped UCNPs. This new strategy will overcome many of the limitations of current fiber-optic based approaches, and will enable new applications in both fundamental science and human health.