Laboratory behavioral tests of mainland and island populations of Podarcis siculus

In accordance with the permit of the Ministry of Environment and Energy (Class: AP /L612-07 I 18-481 65; No: 517-07-l-l-l-18-4), and the permit of the Ethical committee of the Faculty of Science, University of Zagreb, Croatia (No: 251-58-10617-18-16), animals (10 males from each location) were captured by noosing, and transported to the laboratory in cloth bags. Since the population on islet Pijavica is strictly protected, this was the maximum number of animals we were permitted to collect. In the laboratory, animals were maintained in individual terraria (40x30cm) with peat as a substrate, fed every two days with crickets (Acheta domestica), that were supplemented with calcium and vitamins, with water available ad libitum. The light cycle was the same as the natural cycle (14:10 h, light: dark). Room temperature varied between 22°C during the night and 30° C during the day, to simulate natural day-night temperature variation. A plastic tube with one entrance was put in each terrarium to serve as a hiding place. These tubes were also used as containers for transferring the lizards from their terrarium to the experimental setup, to reduce handling stress. Lizards were allowed to habituate to laboratory conditions for one week before being subjected to behavioral testing. The experiments were conducted in accordance with the Directive of the European Parliament and of the Council (2010/63/EU) and the Croatian Animal Protection Law (“Narodne Novine”, 102/17 and 32/19). All efforts were made to reduce the number of animals used and to ensure animal welfare.

本研究依据克罗地亚环境与能源部许可（等级：AP /L612-07 I 18-481 65；编号：517-07-l-l-l-18-4）以及克罗地亚萨格勒布大学理学院伦理委员会许可（编号：251-58-10617-18-16）开展实验。实验动物为各采集地点的10只雄性个体，采用套索法诱捕后以布袋转运至实验室。由于皮亚维察岛（Pijavica）的种群受到严格保护，本次采集的动物数量已达许可上限。实验室中，所有动物均饲养于规格为40×30cm的独立饲养箱内，以泥炭作为饲养基质；每两日投喂一次家蟋蟀（Acheta domestica），并在饲料中补充钙制剂与维生素，同时保证饮水自由取用。光照周期设置为14小时光照/10小时黑暗，与自然光周期一致；室温模拟自然昼夜温差，夜间维持22℃，日间调整为30℃。每个饲养箱内放置单入口塑料管作为躲避场所，该塑料管同时可作为转运容器，用于将蜥蜴从饲养箱转移至实验装置，以降低捕捉应激。在行为实验开始前，所有蜥蜴均需在实验室环境中适应一周。本实验严格遵循欧盟议会与理事会第2010/63/EU号指令以及克罗地亚《动物保护法》（《国家公报》102/17与32/19）的相关要求，尽可能减少实验动物使用数量，并全程保障动物福利。