Meacham C A, Freudenrich T M, Anderson W L, Sui L, Lyons-Darden T, Barone S, Gilbert M E, Mundy W R, Shafer T J
Neurotoxicology Division, National Health and Environmental Effects Research Laboratory/ORD, U.S. Environmental Protection Agency, Research Triangle Park, NC 27711, USA.
Toxicol Appl Pharmacol. 2005 Jun 1;205(2):177-87. doi: 10.1016/j.taap.2004.08.024. Epub 2004 Nov 19.
In vivo exposure levels for neurotoxicants are often reported in parts per million (ppm) concentration in tissue, whereas exposure levels in experiments utilizing in vitro models are most commonly reported in micromolar (muM) concentration in the exposure solution. The present experiments sought to determine whether or not in vitro solution concentration was an appropriate dose-metric for comparison to in vivo tissue levels for lipophilic compounds. To do so, the accumulation of the polychlorinated biphenyl (PCB) mixture Aroclor 1254 (A1254) or methylmercury (MeHg) was examined in three commonly utilized in vitro neuronal tissue models: nerve growth factor differentiated pheochromocytoma (PC12) cells, primary cultures of rat neocortical cells, and adult rat hippocampal slices. Tissues were exposed to A1254 (0.65 ppm) or to MeHg (0.0033-0.33 ppm) in serum-free media for 1 or 24 h. Total PCB or mercury accumulation was measured by dual column gas chromatography with electron capture detection or by cold vapor atomic absorption, respectively. PC12 cells accumulated 66.7 and 103.8 ppm PCBs after 1 and 24 h exposure to A1254. Neocortical neurons also accumulated significant concentrations of PCBs, but less so than PC12 cells. After 1 h exposure to 0.65 ppm A1254, slices contained 3.46 and 0.81 ppm PCBs when exposed in a static and perfused system, respectively. After 1 h exposure to 0.0033, 0.033, and 0.33 ppm MeHg, PC12 cells contained 0.3, 2.2, and 17.7 ppm mercury, respectively; after 24 h, PC12 cells contained 0.4, 2.8, and 21.9 ppm. Hippocampal slices accumulated 1.7 and 4.8 ppm mercury after 1 and 3 h exposure to 0.33 ppm MeHg. For comparison, mercury accumulation in rat fetal and pup brain tissue after maternal exposure [0, 0.1, 1.0, or 2.0 mg/kg/day MeHg from gestational day (GD) 6-15] ranged from 0.05 to 7.89 ppm in 0.1 mg/kg dose animals on postnatal day 10 and 2.0 mg/kg dose animals on GD16, respectively. These results demonstrate that accumulation of PCBs and MeHg in vitro is tissue-, time-, and concentration-dependent and indicates that tissue levels rather than exposure concentrations are a more appropriate metric for comparison of in vitro to in vivo effects.
神经毒物的体内暴露水平通常以组织中百万分之一（ppm）的浓度报告，而在利用体外模型的实验中，暴露水平最常以暴露溶液中微摩尔（μM）的浓度报告。本实验旨在确定体外溶液浓度是否是与亲脂性化合物的体内组织水平进行比较的合适剂量指标。为此，在三种常用的体外神经元组织模型中检测了多氯联苯（PCB）混合物Aroclor 1254（A1254）或甲基汞（MeHg）的积累：神经生长因子分化的嗜铬细胞瘤（PC12）细胞、大鼠新皮质细胞原代培养物和成年大鼠海马切片。将组织在无血清培养基中暴露于A1254（0.65 ppm）或MeHg（0.0033 - 0.33 ppm）1或24小时。分别通过带电子捕获检测的双柱气相色谱法或冷蒸气原子吸收法测量总PCB或汞的积累。PC12细胞在暴露于A1254 1和24小时后积累了66.7和103.8 ppm的PCB。新皮质神经元也积累了显著浓度的PCB，但比PC12细胞少。在暴露于0.65 ppm A1254 1小时后，在静态和灌注系统中暴露的切片分别含有3.46和0.81 ppm的PCB。在暴露于0.0033、0.033和0.33 ppm MeHg 1小时后，PC12细胞分别含有0.3、2.2和17.7 ppm的汞；24小时后，PC12细胞含有0.4、2.8和21.9 ppm。海马切片在暴露于0.33 ppm MeHg 1和3小时后积累了1.7和4.8 ppm的汞。作为比较，母体暴露[妊娠第（GD）6 - 15天给予0、0.1、1.0或2.0 mg/kg/天MeHg]后，大鼠胎儿和幼崽脑组织中的汞积累在出生后第10天的0.1 mg/kg剂量动物和GD16的2.0 mg/kg剂量动物中分别为0.05至7.89 ppm。这些结果表明，PCB和MeHg在体外的积累是组织、时间和浓度依赖性的，并表明组织水平而非暴露浓度是比较体外和体内效应的更合适指标。
