Transcriptomic studies revealed that hundreds of mRNAs show differential expression in the brains of sleeping versus awake rats, mice, flies, and sparrows. Although these results have offered clues regarding the molecular consequences of sleep and sleep loss, their functional significance thus far has been limited. This is because the previous studies pooled transcripts from all brain cells, including neurons and glia. In the following experiment, we studied the specific effects of sleep and wake conditions on glia cells of mouse cerebral cortex using the genetically targeted translating ribosome affinity purification (TRAP) methodology. We used bacterial artificial chromosome (BAC) transgenic mice expressing EGFP tagged ribosomal protein L10a in oligodendrocytes which constitute a defined cellular population of the mouse brain. Using this approach, we could extract only the oligodendrocitic mRNAs, and only those already committed to be translated into proteins (L10a is part of the translational machinery). Six mice for each vigilant state group (sleep (S), waking (W), and sleep deprivation (SD)) were considered. For each animal, one forebrain sample was immediately processed. Samples were immunoprecipitated to isolate oligodendrocytes. The precipitated portion formed the bound sample (IP) containing olidodendrocytes and the remaining part formed the unbound sample (UB) containing all the remaining cell types (neurons and other glia cells). Then, both IP and UB samples were processed and RNA was extracted.
山东省济南市章丘区文博路2号
齐鲁师范学院 genelibs生信实验室
山东省济南市高新区舜华路750号
大学科技园北区F座4单元2楼
电话: 0531-88819269