Centre for Environmental and Marine studies (CESAM), Department of Biology, University of Aveiro, Portugal; Department of Environmental Chemistry, Institute of Environmental Assessment and Water Research (IDAEA), Spanish Research Council (IDAEA, CSIC), Jordi Girona 18, 08034 Barcelona, Spain.
Instituto Politecnico Nacional, Escuela Nacional de Ciencias Biológicas,-Lab. de Hidrobiología Experimental, Carpio y Plan de Ayala S/N, Mexico City 11340, Mexico.
Sci Total Environ. 2019 Apr 20;662:160-167. doi: 10.1016/j.scitotenv.2019.01.187. Epub 2019 Jan 16.
Recent advances in imaging allow to monitor in real time the behaviour of individuals under a given stress. Light is a common stressor that alters the behaviour of fish larvae and many aquatic invertebrate species. The water flea Daphnia magna exhibits a vertical negative phototaxis, swimming against light trying to avoid fish predation. The aim of this study was to develop a high-throughput image analysis system to study changes in the vertical negative phototaxis of D. magna first reproductive adult females exposed to 0.1 and 1 μg/L of four neuro-active drugs: diazepam, fluoxetine, propranolol and carbamazepine. Experiments were conducted using a custom designed experimental chamber containing four independent arenas and infrared illumination. The apical-located visible light and the GigE camera located in front of the arenas were controlled by the Ethovision XT 11.5 sofware (Noldus Information Technology, Leesburg, VA). Total distance moved, time spent per zone (bottom vs upper zones) and distance among individuals were analyzed in dark and light conditions, and the effect of different intensities of the apical-located visible light was also investigated. Results indicated that light intensity increased the locomotor activity and low light intensities allowed to better discriminate individual responses to the studied drugs. The four tested drugs decreased the response of exposed organisms to light: individuals moved less, were closer to the bottom and at low light intensities were closer each other. At high light intensities, however, exposed individuals were less aggregated. Propranolol, carbamazepine and fluoxetine induced the most severe behavioural effects. The tested drugs at environmental relevant concentrations altered locomotor activity, geotaxis, phototaxis and aggregation in D. magna individuals in the lab. Therefore the new image analysis system presented here was proven to be sensitive and versatile enough to detect changes in diel vertical migration across light intensities and low concentration levels of neuro-active drugs.
近年来，成像技术的进步使得人们能够实时监测个体在特定压力下的行为。光作为一种常见的应激源，会改变鱼类幼虫和许多水生无脊椎动物的行为。大型溞（Daphnia magna）表现出垂直负趋光性，即逆着光游动，试图躲避鱼类捕食。本研究旨在开发一种高通量的图像分析系统，以研究暴露于 0.1 和 1μg/L 四种神经活性药物（地西泮、氟西汀、普萘洛尔和卡马西平）的大型溞第一代生殖雌体的垂直负趋光性变化。实验使用一个带有四个独立竞技场和红外照明的定制实验室进行。位于顶部的可见光和位于竞技场前面的 GigE 摄像机由 Ethovision XT 11.5 软件（Noldus Information Technology，Leesburg，VA）控制。在黑暗和光照条件下分析总移动距离、每个区域（底部和上部区域）的停留时间以及个体之间的距离，并研究不同强度的顶部可见光的影响。结果表明，光强度增加了运动活性，低光强度更有利于区分个体对研究药物的反应。四种测试药物降低了暴露生物对光的反应：个体移动较少，靠近底部，在低光强度下彼此靠近。然而，在高光强度下，暴露的个体聚集程度较低。普萘洛尔、卡马西平和氟西汀引起的行为影响最严重。在环境相关浓度下，测试药物改变了大型溞个体在实验室中的运动活性、地理趋性、趋光性和聚集性。因此，这里提出的新图像分析系统被证明足够敏感和通用，可以检测到不同光强度和低浓度水平的神经活性药物对昼夜垂直迁移的影响。
