Biosensors for determination of multiple neurotransmitters in vertebrate retina

项目概述：该项目的长期目标是开发一种改变范式的神经传感技术 用于直接、同时监测多种神经递质的活动，以了解大脑功能。 视网膜被选为我们的模型系统，因为它很容易接近和成熟的神经生理学。 而且迫切需要这种工具来了解神经递质在各种眼病中的作用，例如 糖尿病视网膜病变。视网膜感光细胞(视杆细胞和视锥细胞)将光转化为电信号。这个 电信号通过双极细胞传递到神经节细胞，即视网膜的输出神经元，然后 对大脑的影响。通过这一途径的信号传递由无长突细胞调制，无长突细胞是视网膜 中间神经元。无长突细胞有多种类型，但它们都能合成和释放神经调节剂，如 多巴胺(DA)、γ-氨基丁酸(GABA)和乙酰胆碱(ACh)。具体地说，多巴胺能 无长突细胞(DAC)共同释放GABA和DA，它们在调节视网膜光敏感性方面起着关键作用 和眼睛发育。星爆无长突细胞共同释放GABA和ACh，启动运动方向 视觉系统的一部分。从历史上看，视网膜神经元和无长突细胞释放的神经递质 通过电生理方法和/或使用电分析进行氧化还原检测来间接研究 使用碳纤维微电极的技术。然而，细胞中的电活动并不总是匹配的 细胞内神经递质的释放。氧化还原方法只适用于数量相对较少的分析物 比如地方检察官。我们构建了一种新型的生物传感器，它采用互补的电化学和 压电传感器，我们的初步结果表明，它可以区分氧化还原和非氧化还原 活跃的神经递质。R21项目的目标是开发一种小型化的多模式生物传感器，以 实时以高空间和时间分辨率同时测量多种神经递质，标签- 不需要试剂，有两个目的：1.小型化多式联运车的设计、制造和表征 电化学(E)和压电式传感器(薄膜体声谐振器(FBAR)(即E-FBAR)) 神经传感探针；和2：通过监测多巴胺、GABA和ACh来验证神经传感探针 在活着的正常和糖尿病视网膜神经元中。该项目的成功完成将证明一种不含试剂的， 无标记实时同时检测大鼠脑内氧化还原活性和非氧化还原活性神经递质 视网膜具有多方面的信息，具有高度的敏感性和选择性。这样的工具将具有无价的研究价值。 旨在了解视网膜神经退行性疾病的原因和机制 作为糖尿病视网膜病变，也测试用于治疗此类疾病的治疗剂。这部小说 技术也可以被用来监测大脑中的其他重要神经递质，增加我们的 对大脑功能的理解。我们成熟的、高技能的、多学科的团队拥有专业知识 在电化学和声学生物传感器、微器件和微传感器的设计和制造以及视觉方面 神经科学来开发和验证提出的神经传感技术。

Project Summary: The long-term goal of this project is to develop a paradigm-shifting neurosensing technology for direct, simultaneous monitoring of the activity of multiple neurotransmitters for understanding brain function. The retina is selected as our model system due to its easy accessibility and well-established neurophysiology and the urgent needs in such tool to understand the roles of neurotransmitters in various eye diseases such as diabetic retinopathy. Retinal photosensitive cells (rods and cones) convert light into an electrical signal. The electrical signal is transmitted through bipolar cells to ganglion cells, the output neurons of the retina, and then to the brain. Signal transmission through this pathway is modulated by amacrine cells, which are retinal interneurons. There are multiple types of amacrine cells, but all synthesize and release neuromodulators such as dopamine (DA), gamma-aminobutyric acid (GABA) and acetylcholine (ACh). Specifically, dopaminergic amacrine cells (DACs) co-release GABA and DA, which play a critical role in modulating retinal light sensitivity and eye development. Starburst amacrine cells co-release GABA and ACh, which initiates the motion direction of the visual system. Historically, the release of neurotransmitters from retinal neurons and amacrine cells has been studied indirectly, through electrophysiological methods and/or redox detection using electroanalytical techniques employing carbon fiber microelectrodes. However, electrical activity in a cell does not always match the release of neurotransmitter from the cell. Redox methods only work for a relatively small number of analytes such as DA. We have constructed a novel biosensor that employs complementary electrochemical and piezoelectric sensors, and our preliminary results show that it can differentiate between redox and non-redox active neurotransmitters. The objective of this R21 project is to develop a miniaturized multimodal biosensor to measure multiple neurotransmitters simultaneously with high spatial and temporal resolution in real time, label- and reagent-free with two Aims: 1. Design, fabrication, and characterization of a miniaturized multimodal electrochemical (E) and piezoelectric sensor (thin film bulk acoustic resonator (FBAR) (i.e. E-FBAR) neurosensing probe; and 2: Validation of the neurosensing probe through monitoring dopamine, GABA, and ACh in living normal and diabetic retinal neurons. Successful completion of this project will certify a reagent-free, label-free and real-time simultaneously detection of both redox active and non-redox active neurotransmitters in retina with multifaceted information in high sensitivity and selectivity. Such a tool will be invaluable to research aimed at understanding the causes and mechanisms responsible for retinal neurodegenerative diseases such as diabetic retinopathy, and also to test therapeutic agents for the treatment of such diseases. This novel technology could also be adapted to monitor other important neurotransmitters in the brain, increasing our understanding of brain functions. Our well- established, highly skilled, multidisciplinary team has the expertise in electrochemical and acoustic biosensors, microdevice and microsensor design and fabrication, and visual neuroscience to develop and validate the proposed neurosensing technology.