背景:迷走神经传入神经元代表肠-脑轴的关键神经感觉分支,描述了胃肠系统和大脑之间的双向通信。这些神经元对于检测和将感觉信息从外围传递到中枢神经系统以调节摄食行为非常重要。新陈代谢,和炎症。混杂变量使分离迷走神经传入在介导这些生理过程中的作用的过程复杂化。因此,我们建立了肠-脑轴感觉分支的微流控模型.我们证明了这种微流体模型成功地分隔了神经元的细胞体和神经突末端,从而模拟这些神经元的解剖布局,以更准确地研究生理相关过程。
方法:我们将原代大鼠迷走神经传入神经元培养物实施到由两个与径向微通道互连的同心腔室组成的微流体平台中。微流体平台将细胞体与迷走神经传入神经元的神经突末端分离。然后,我们在神经末梢引入了生理相关的胃肠效应分子,并使用活细胞钙成像评估了它们沿神经突的逆行转运或引起电生理反应的能力。
结果:微通道出口的角度决定了神经突生长到腔室中与沿着腔室壁追踪的可能性。当神经突末端暴露于荧光标记的霍乱毒素亚基B时,蛋白质被吸收并在24小时内沿着神经突逆行运输。此外,机械扰动(例如,冲洗)的神经突末端显着增加了远端体细胞中的细胞内钙浓度。最后,辣椒素的膜展示受体被表达并沿着新投射的神经突运输,通过共聚焦显微镜显示。
结论:在这项工作中,我们开发了一种微流控装置,可以概括体外迷走神经传入神经元的解剖布局。我们展示了该平台的两种生理相关应用:逆行运输和电生理反应。我们希望该工具能够对迷走神经传入神经元在肠-脑轴中的作用进行控制研究。
BACKGROUND: Vagal afferent neurons represent the key neurosensory branch of the gut-brain axis, which describes the bidirectional communication between the gastrointestinal system and the brain. These neurons are important for detecting and relaying sensory information from the periphery to the central nervous system to modulate feeding behavior, metabolism, and inflammation. Confounding variables complicate the process of isolating the role of the vagal afferents in mediating these physiological processes. Therefore, we developed a microfluidic model of the sensory branch of the gut-brain axis. We show that this microfluidic model successfully compartmentalizes the cell body and neurite terminals of the neurons, thereby simulates the anatomical layout of these neurons to more accurately study physiologically-relevant processes.
METHODS: We implemented a primary rat vagal afferent neuron culture into a microfluidic platform consisting of two concentric chambers interconnected with radial microchannels. The microfluidic platform separated cell bodies from neurite terminals of vagal afferent neurons. We then introduced physiologically-relevant gastrointestinal effector molecules at the nerve terminals and assessed their retrograde transport along the neurite or capacity to elicit an electrophysiological response using live cell calcium imaging.
RESULTS: The angle of microchannel outlets dictated the probability of neurites growing into a chamber versus tracking along chamber walls. When the neurite terminals were exposed to fluorescently-labeled cholera toxin subunit B, the proteins were taken up and retrogradely transported along the neurites over the course of 24 h. Additionally, mechanical perturbation (e.g., rinsing) of the neurite terminals significantly increased intracellular calcium concentration in the distal soma. Finally, membrane-displayed receptor for capsaicin was expressed and trafficked along newly projected neurites, as revealed by confocal microscopy.
CONCLUSIONS: In this work, we developed a microfluidic device that can recapitulate the anatomical layout of vagal afferent neurons in vitro. We demonstrated two physiologically-relevant applications of the platforms: retrograde transport and electrophysiological response. We expect this tool to enable controlled studies on the role of vagal afferent neurons in the gut-brain axis.