Laboratory of Neurobiology, Structural Cell Biology Section, NINDS, NIH, Bethesda, MD 20892, USA.
Biochem J. 2010 Apr 14;427(3):535-40. doi: 10.1042/BJ20091344.
Biochemical studies have suggested that certain synaptic proteins associate with lipid rafts to perform key functions within the synapse. However, variability in biochemical preparations raises questions as to which synaptic proteins actually associate with lipid rafts. In the present study, we use both electron microscopy and biochemistry to investigate AMPA (alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid) receptor localization in synaptic membrane subfractions prepared in two different ways, by Triton X-100 detergent treatment or without detergent by sonication at high pH. Immunogold electron microscopy shows that a detergent-resistant synaptosomal membrane subfraction consists of empty vesicles 0.1-1.0 microm in diameter. A subpopulation of these vesicles labelled for glycosphingolipid GM1 ganglioside, a marker of lipid rafts, and 46% of the labelled vesicles also labelled for the AMPA receptor subunit GluR2. This co-segregation into specific vesicles does not depend on effects of detergent because a similar distribution of label was found in vesicles isolated without the use of detergent. Our results suggest that AMPA receptors localize within specific regions of synaptic membranes rich in GM1 ganglioside.
生化研究表明，某些突触蛋白与脂筏结合，在突触内发挥关键功能。然而，生化制剂的可变性引发了一个问题，即哪些突触蛋白实际上与脂筏结合。在本研究中，我们使用电子显微镜和生物化学方法来研究两种不同方法制备的突触膜亚部分中 AMPA（α-氨基-3-羟基-5-甲基异恶唑-4-丙酸）受体的定位，一种方法是用 Triton X-100 去污剂处理，另一种方法是在高 pH 值下用超声波处理而不使用去污剂。免疫金电子显微镜显示，一种去污剂抗性的突触体膜亚部分由直径 0.1-1.0 微米的空囊泡组成。这些囊泡中的一部分标记了神经节苷脂 GM1，这是脂筏的标志物，并且 46%的标记囊泡也标记了 AMPA 受体亚基 GluR2。这种共分离到特定囊泡中并不依赖于去污剂的作用，因为在不使用去污剂的情况下分离的囊泡中也发现了类似的标记分布。我们的结果表明，AMPA 受体定位于富含 GM1 神经节苷脂的突触膜特定区域内。
