Centro de Investigación, Hospital Universitario La Fe, Valencia, Spain.
Toxicol Sci. 2010 May;115(1):202-13. doi: 10.1093/toxsci/kfq040. Epub 2010 Feb 4.
Endocytosis is required for many cellular pivotal processes, including membrane recycling, nutrient uptake, and signal transduction. This complex process is particularly relevant in polarized cells, such as neurons. Previous studies have demonstrated that alcohol alters intracellular traffic, including endocytosis, in several cell types. However, information on the effect of chronic alcohol exposure on this process in neurons is scarce. As an approach, we investigated the effect of alcohol exposure on the internalization of two widely used endocytic markers, albumin and transferrin, in developing hippocampal neurons in primary culture. The effect of this treatment on the levels of several representative proteins involved in the endocytic process was also analyzed. Some of these proteins are also involved in the organization of the actin cytoskeleton. Pretreatment of cells with inhibitors chlorpromazine or nystatin indicates that albumin is internalized mainly by caveolin-dependent endocytosis. On the other hand, alcohol decreases the endocytosis of both markers, although no qualitative changes in the distribution of either of these molecules were observed. Finally, the effect of ethanol on the proteins analyzed was heterogeneous. Alcohol decreases the levels of clathrin, AP-2, SNX9, Rab5, Rab11, EEA1, Cdc42, or RhoA but increases the amount of Arf6. Moreover, alcohol does not affect the levels of caveolin1, dynamin1, Rab7, and LAMP2. This toxic effect of alcohol on endocytosis could affect some of the important neuronal activities, which depend on this process, including cell signaling. Our results in neurons also stress the notion that one of the main targets of ethanol is intracellular transport.
内吞作用是许多细胞关键过程所必需的，包括膜循环、营养物质摄取和信号转导。这个复杂的过程在极化细胞中尤为重要，如神经元。先前的研究表明，酒精改变了几种细胞类型的细胞内运输，包括内吞作用。然而，关于慢性酒精暴露对神经元中这一过程的影响的信息却很少。作为一种方法，我们研究了酒精暴露对原代培养海马神经元中两种广泛使用的内吞标记物白蛋白和转铁蛋白内化的影响。还分析了这种处理对参与内吞过程的几种代表性蛋白水平的影响。其中一些蛋白也参与肌动蛋白细胞骨架的组织。用氯丙嗪或制霉菌素预处理细胞表明，白蛋白主要通过网格蛋白依赖性内吞作用内化。另一方面，酒精降低了这两种标记物的内吞作用，尽管没有观察到这两种分子的分布有任何定性变化。最后，乙醇对所分析的蛋白质的影响是不均匀的。酒精降低网格蛋白、AP-2、SNX9、Rab5、Rab11、EEA1、Cdc42 或 RhoA 的水平，但增加 Arf6 的量。此外，酒精不影响 caveolin1、dynamin1、Rab7 和 LAMP2 的水平。酒精对内吞作用的这种毒性作用可能会影响一些依赖于该过程的重要神经元活动，包括细胞信号转导。我们在神经元中的结果也强调了这样一种观点，即乙醇的主要靶点之一是细胞内运输。
