Data for: INFLUENCE OF MATERNAL AGE ON THE EFFECTS OF SELENO-L-METHIONINE IN THE MODEL ORGANISM DAPHNIA PULEX UNDER STANDARD AND HEAT STRESS CONDITIONS

We evaluated the influence of maternal age on the effects of seleno-methionine (SeMet) for lifespan, reproduction, and heat-stress resistance in D. pulex. Populations of animals were maintained under previously established laboratory culture protocol conditions. All D. pulex cultures were maintained in COMBO media at a density of 20 animals per 1 L media. COMBO media and Animate stock solutions were prepared as described by Kilham, Kreeger, Lynn, Goulden, & Herrera, 1998. For dose-finding study a total of five SeMet treatment concentrations were analyzed: 0-g SeMet/L, 1-g SeMet/L, 2-g SeMet/L, 4-g SeMet/L, and 8-g SeMet/L. The highest treatment (8-g SeMet/L) is the concentration of SeMet at which 50% of the population were dead (LD 50) in adult Daphnia magna after a 48-hour exposure48. A 50% dilution was applied to obtain the remaining treatments. In the control group (0-g SeMet/L) no additional Se was added. Each SeMet treatment was assessed using a population of n=20 individuals, that were followed as populations throughout their lifespan. Mortality and presence of offspring were recorded throughout the lifespan of each population. Daphnia pulex maternal populations were established from the third brood of our laboratory maintenance D. pulex culture (maternal age ~ 12 days). Neonates ( 24 hours of age; n=80) were obtained from the maintenance population and randomized into four maternal populations. These maternal populations each consisted of n=20 individuals that were housed in 1 L beakers containing 1,000 ml COMBO media and allowed to age under standard laboratory maintenance conditions. All experimental animals exposed to SeMet were obtained from these maternal populations when the mothers were 8-days of age (early life (2nd-3rd) brood) and when the mothers were 32-days of age (late life (12th-14th) brood).
For the lifespan analysis, offspring from maternal populations were combined into a single pool from which offspring were randomized into three SeMet treatment groups [(1 maternal population x 3 SeMet treatments) x 25 replicates per SeMet treatment]. This was conducted once when the maternal populations produced an early life brood and repeated when the maternal populations produced a late life brood. For the heat induced stress analysis, offspring were tracked by maternal population. Within the maternal population tracks, offspring were randomized into the three SeMet treatments and housed as populations, creating one experimental group for each maternal population. This was repeated for replicate groups to ensure that each SeMet treatment is assessed in 8 populations for each maternal age group; giving a total of n=24 exposure groups per maternal age [(4 maternal populations x 3 SeMet treatments) x 2 replicates].
Daphnia were submitted to the Virginia-Maryland College of Veterinary Medicine Toxicology Lab for quantification of Se content by atomic absorption spectrometry.

本研究评估了母龄对硒代蛋氨酸（seleno-methionine, SeMet）作用于蚤状溞（Daphnia pulex，下称D. pulex）的寿命、繁殖性能及热应激抗性的影响。实验所用蚤状溞种群均按照已建立的实验室培养规程进行维持培养。所有D. pulex种群均饲养于COMBO培养基（COMBO media）中，养殖密度为每1升培养基放养20只个体。COMBO培养基与Animate储备液（Animate stock solutions）均参照Kilham、Kreeger、Lynn、Goulden与Herrera于1998年发表的方法进行配制。

在剂量探索实验中，共设置5个硒代蛋氨酸处理浓度：0 μg SeMet/L、1 μg SeMet/L、2 μg SeMet/L、4 μg SeMet/L及8 μg SeMet/L。最高处理浓度（8 μg SeMet/L）为前期研究中大型溞（Daphnia magna）成体经48小时暴露后的半数致死浓度（LD₅₀）。其余处理浓度均通过对该最高浓度进行50%梯度稀释得到。对照组（0 μg SeMet/L）不添加额外的硒元素。每个硒代蛋氨酸处理组均采用n=20只个体组成的种群进行评估，并对该种群的整个生命周期进行追踪观测。在每个种群的整个生命周期中，均记录其死亡率与子代产生情况。

本研究的蚤状溞母代种群均来自实验室维持培养的蚤状溞种群的第三胎幼体（母龄约12天）。从维持培养种群中获取日龄≤24小时的新生幼体（n=80），并将其随机分为4个母代种群。每个母代种群包含n=20只个体，饲养于盛有1000 mL COMBO培养基的1 L烧杯中，并在标准实验室维持培养条件下进行培育。所有接受硒代蛋氨酸处理的实验动物均取自上述母代种群，分别在母代8日龄（早期生殖阶段，即第2-3胎）与32日龄（晚期生殖阶段，即第12-14胎）时获取。

在寿命分析实验中，将各母代种群的子代合并为一个混合种群，再从中随机分配至3个硒代蛋氨酸处理组[（1个母代种群×3个硒代蛋氨酸处理组）×每个处理组25个重复]。该实验分别在母代种群产生早期胎与晚期胎时各开展一次。

在热应激抗性分析实验中，子代按母代种群进行分组追踪。在每个母代种群的子代中，将幼体随机分配至3个硒代蛋氨酸处理组并以种群形式饲养，每个母代种群对应一个实验组。通过设置重复组开展该实验，以确保每个母龄组的每个硒代蛋氨酸处理组均有8个种群接受评估；最终每个母龄组共设置n=24个暴露组[（4个母代种群×3个硒代蛋氨酸处理组）×2个重复]。

所有蚤状溞样本均送至弗吉尼亚-马里兰兽医学院毒理学实验室，采用原子吸收光谱法对其硒含量进行定量检测。