Gassó S, Cristòfol R M, Selema G, Rosa R, Rodríguez-Farré E, Sanfeliu C
Department of Pharmacology and Toxicology, Institut d'Investigacions Biomèdiques de Barcelona, CSIC, IDIBAPS, Barcelona, Spain.
J Neurosci Res. 2001 Oct 1;66(1):135-45. doi: 10.1002/jnr.1205.
The effects of the environmental contaminants methylmercury (MeHg) and inorganic mercury (HgCl(2)) on cell viability, intracellular calcium concentration (Ca(2+)), and reactive oxygen species (ROS) generation were studied in rat cerebellar granule neuron cultures using fluorescent methods. MeHg exhibited an LC(50) (2.47 microM) tenfold lower than that of HgCl(2) (26.40 microM). To study the involvement of oxidative stress and Ca(2+) homeostasis disruption in mercury-induced cytotoxicity, we tested the neuroprotective effects of several agents that selectively interfere with these mechanisms. After a 24 hr exposure, the cytotoxic effect of both mercury compounds was reduced by thapsigargin, an inhibitor of endoplasmic reticulum Ca(2+)-ATPase; the Ca(2+) channel blocker flunarizine; and the Na(+)/Ca(2+) exchanger blocker benzamil. All these compounds decreased the mercury-mediated Ca(2+) rise. These results indicate that Ca(2+) influx through Ca(2+) channels and the Na(+)/Ca(2+) exchanger and Ca(2+) mobilization from the endoplasmic reticulum are involved in mercury-mediated cytotoxicity. The antioxidants probucol and propyl gallate reduced the HgCl(2) toxicity. Probucol and vitamin E partially inhibited the MeHg toxicity after a 24 hr period, whereas propyl gallate completely prevented this effect. Probucol slightly reduced ROS generation in methylmercury-exposed cultures and decreased mercury-mediated rise of Ca(2+). Propyl gallate abolished ROS generation and partially inhibited the increase of Ca(2+) induced by both mercury compounds. Propyl gallate also protected human cerebral cortical neuron cultures from the MeHg effect even after 72 hr of MeHg exposure, thus showing a long-lasting effect. Our data suggest that disruption of redox equilibrium and Ca(2+) homeostasis contribute equally to HgCl(2)-mediated toxicity, whereas oxidative stress is the main cause of MeHg neurotoxicity.
采用荧光法在大鼠小脑颗粒神经元培养物中研究了环境污染物甲基汞（MeHg）和无机汞（HgCl₂）对细胞活力、细胞内钙浓度（[Ca²⁺]i）以及活性氧（ROS）生成的影响。MeHg的半数致死浓度（LC₅₀）（2.47 μM）比HgCl₂（26.40 μM）低十倍。为了研究氧化应激和Ca²⁺稳态破坏在汞诱导的细胞毒性中的作用，我们测试了几种选择性干扰这些机制的药物的神经保护作用。暴露24小时后，内质网Ca²⁺-ATP酶抑制剂毒胡萝卜素、Ca²⁺通道阻滞剂氟桂利嗪和Na⁺/Ca²⁺交换体阻滞剂苄胺咪都降低了两种汞化合物的细胞毒性作用。所有这些化合物都减少了汞介导的[Ca²⁺]i升高。这些结果表明，通过Ca²⁺通道和Na⁺/Ca²⁺交换体的Ca²⁺内流以及内质网的Ca²⁺动员参与了汞介导的细胞毒性。抗氧化剂普罗布考和没食子酸丙酯降低了HgCl₂的毒性。普罗布考和维生素E在24小时后部分抑制了MeHg的毒性，而没食子酸丙酯完全阻止了这种作用。普罗布考略微降低了甲基汞暴露培养物中的ROS生成，并降低了汞介导的[Ca²⁺]i升高。没食子酸丙酯消除了ROS生成，并部分抑制了两种汞化合物诱导的[Ca²⁺]i增加。即使在暴露于MeHg 72小时后，没食子酸丙酯也保护人脑皮质神经元培养物免受MeHg的影响，从而显示出持久的作用。我们的数据表明，氧化还原平衡和Ca²⁺稳态的破坏对HgCl₂介导的毒性贡献相同，而氧化应激是MeHg神经毒性的主要原因。
