Optogenetic Population Clamp to Study Long-term Plasticity in Vitro

描述(由申请人提供)： 这是一项针对PA-10-009的多学科生物工程研究补助金(BRG)提案，具有设计驱动和发现驱动的元素。它基于一个越来越受欢迎的假设，即许多神经疾病的进展植根于已变得不适应的体内平衡可塑性。这些疾病可被归类为去传入障碍，在中枢神经系统组织中，由于白质损伤、中风或感觉感受器或周围神经的损伤，这些组织中的破坏性同步群体爆发活动会在几天或几周内发展起来，其输入已大大减少或消除。人群爆发时产生的低频、高幅度电放电可表现为癫痫发作、慢性疼痛、肌张力障碍、耳鸣或其他致残症状，这取决于神经系统的哪个部分在去传入后变得高度兴奋。药物治疗通常是完全无效的。这导致许多人提出了包括植入电极的直接、局部脑刺激或经颅磁刺激的治疗方法。光遗传学提供了一种更局部化和更具体的方式来刺激脑组织，因为它可以使特定类型的神经细胞对特定颜色的光敏感。也就是说，光可以激活或沉默目标神经元，以努力使异常神经正常化。 活动。 在利用多电极阵列刺激抑制培养的大脑皮层网络中癫痫样群体爆发的成功的闭环方法的基础上，本项目旨在开发和优化一种闭环光遗传学工具，以获得对稳态可塑性机制的控制，并逆转去传入组织表达同步爆发的趋势。这一群体钳制将使用来自多电极阵列底物的细胞外记录作为反馈信号，快速而连续地调整彩色光的脉冲，选择性地激活和抑制不同类型的神经元，以维持所需的活动水平。由于体外制剂典型的去传入，表达群体放电的皮质网络将被夹在不同的活动设定值-几天内。动态平衡反应，如突触强度的变化，将通过细胞内记录和细胞外群体活动测量来监测。使用腺相关病毒载体针对兴奋性锥体神经元或抑制性中间神经元的光遗传结构的组合，将根据它们作为操纵柄的能力进行比较，通过这些手柄可以操纵内稳态可塑性。将开发反馈控制算法，以实现最有效和最持久的人口激增缓解，同时加强网络功能的措施，如复杂光输入和尖峰输出之间的互信息。通过为研究不同的结构和参数提供一个可访问和可操作的试验台，光遗传种群钳将为通过植入光纤使用闭环光刺激的各种神经疾病的基因治疗铺平道路。

DESCRIPTION (provided by applicant): This is a multi-disciplinary _Bioengineering Research Grants_ (BRG) proposal in response to PA-10-009, with design-driven and discovery-driven elements. It is based on a hypothesis that is gaining in popularity, that the progression of a number of neurological disorders is rooted in homeostatic plasticity that has become maladaptive. These can be classified as de-afferentation disorders, where disruptive synchronized population bursting activity develops across days or weeks, in CNS tissue whose inputs have been greatly reduced or eliminated by white matter damage, stroke, or damage to sensory receptors or peripheral nerves. Low-frequency, high-amplitude electrical discharges from population bursting can manifest as seizures, chronic pain, dystonia, tinnitus, or other disabling symptoms, depending on which part of the nervous system has become hyper-excitable after deafferentation. Pharmacological treatments are often completely ineffective. This has lead many to propose therapies that involve direct, localized brain stimulation with implanted electrodes or transcranial magnetic stimulation. Optogenetics provides a much more localized and specific way to stimulate brain tissue, because it can render defined neural cell types sensitive to light of specific colors. Wit it, light can either activate or silence targeted neurons in an effort to normalize aberrant neural activity. Based on a successful closed-loop approach to quieting seizure-like population bursting in cultured cortical networks with multi-electrode array stimulation, this project is to develop and optimize a closed-loop optogenetic tool to gain control over homeostatic plasticity mechanisms, and to reverse the tendency of deafferented tissue to express synchronized bursting. This _Population Clamp_ will employ extracellular recording from multi-electrode array substrates as the feedback signal, to rapidly and continuously adjust pulses of colored light, selectively activating and inhibiting different neuron types, to maintain a desired activity level. Cortical networks expressing population discharges due to the deafferentation typical of in vitro preparations will be clamped to different activity set----points for days. Homeostatic responses, such as changes in synaptic strength, will be monitored with intracellular recording and extracellular measures of population activity. Combinations of optogenetic constructs, directed at excitatory pyramidal neurons or inhibitory interneurons using adeno-associated viral vectors, will be compared in terms of their ability to serve as handles by which homeostatic plasticity can be manipulated. Feedback control algorithms will be developed that enable the most effective and enduring remission of population bursting, while enhancing measures of network function, such as the mutual information between complex light input and spiking output. By providing an accessible and manipulable test bed for studying different constructs and parameters, the Optogenetic Population Clamp will pave the way for gene-therapeutic treatments of a variety of neurological disorders that employ closed-loop light stimulation via implanted fiber optics.