Department of Biology, York University, Toronto, ON, Canada.
Lab Chip. 2017 Nov 21;17(23):4048-4058. doi: 10.1039/c7lc00786h.
The zebrafish larva is an important vertebrate model for sensory-motor integration studies, genetic screening, and drug discovery because of its excellent characteristics such as optical transparency, genetic manipulability, and genetic similarity to humans. Operations such as precise manipulation of zebrafish larvae, controlled exposure to chemicals, and behavioural monitoring are of utmost importance to the abovementioned studies. In this work, a novel microfluidic device is presented to easily stabilize an individual larva's head using a microfluidic trap while leaving the majority of the body and the tail unhindered to move freely in a downstream chamber. The device is equipped with a microvalve to prevent the larva's escape from the trap and a microchannel beside the larva's head to expose it to chemicals at desired concentrations and times, while investigating multiple behaviours such as the tail, eye, and mouth movement frequencies. An in situ air bubble removal module was also incorporated to increase the yield of experiments. The functionality of our device in comparison to a conventional droplet-based technique was tested using l-arginine exposure and viability assays. We found that the larvae in the device and the droplet exhibit similar tail and eye response trends to nM-mM concentrations of l-arginine, and that the survival of the larvae is not affected by the device. However, the tail responses in the device were numerically higher than the droplet-tested larvae at nM-mM l-arginine concentrations. In the future, our device has the potential to be used for conducting simultaneous whole-brain functional imaging, upon optimized immobilization of the brain, and behavioural analysis to uncover differences between diseased and healthy states in zebrafish.
斑马鱼幼虫是用于感觉运动整合研究、遗传筛选和药物发现的重要脊椎动物模型，因为它具有出色的特性，如光学透明度、遗传可操作性和与人类的遗传相似性。精确操纵斑马鱼幼虫、控制化学物质暴露和行为监测等操作对上述研究至关重要。在这项工作中，提出了一种新颖的微流控装置，该装置使用微流阱轻松稳定单个幼虫的头部，同时使大部分身体和尾巴不受阻碍地在下游腔室中自由移动。该装置配备了一个微阀，以防止幼虫从陷阱中逃脱，并在幼虫头部旁边设有一个微通道，以在所需浓度和时间下暴露于化学物质，同时研究尾巴、眼睛和嘴巴运动频率等多种行为。还集成了原位气泡去除模块，以提高实验的产量。通过使用 l-精氨酸暴露和生存能力测定，测试了我们的装置与传统基于液滴的技术相比的功能。我们发现，装置中的幼虫和液滴对 nM-mM 浓度的 l-精氨酸表现出相似的尾巴和眼睛反应趋势，并且装置不会影响幼虫的存活。然而，在 nM-mM l-精氨酸浓度下，装置中的尾巴反应数值高于液滴测试的幼虫。将来，我们的装置有可能用于进行全脑功能成像，在优化大脑固定后，进行行为分析，以揭示斑马鱼中疾病和健康状态之间的差异。
