SAGE in Cerebellum After Alcohol Exposure

描述(申请人提供)：众所周知，过量饮酒会对胎儿、成人和衰老的神经系统产生毒害作用。众所周知，在整个发育过程中，小脑神经元都会受到酒精摄入的损害。由于小脑的主要功能是通过整合感觉输入和运动输出来调节运动和平衡，因此组成神经元的任何损伤都可能导致运动协调的障碍。酒精引起的小脑损伤似乎是许多基因表达累积变化的结果。到目前为止，只有一小部分构成人类和小鼠基因组的基因被单独或通过DNA微阵列测试了酒精诱导的变化。由于DNA微阵列技术是一个封闭的系统，只能检测已知的基因，我们建议使用基因表达序列分析(SAGE)对酒精暴露小鼠小脑的基因表达进行全局分析。SAGE是一个“开放”的系统，能够检测和数字量化已知和未知的基因。拟议的实验包括构建、测序和生物信息学分析常规小脑SAGE文库，在出生后早期、成年期和衰老后不久，接触到大量的酒精。由此产生的关于酒精诱导的基因表达变化的全面和定量的SAGE数据库，包括新基因的变化，将通过NCBI网站传播。未来的长期实验将超越本R03的范围并作为计划中的R01提交的一部分，旨在：(1)构建酒精暴露后的几个时间间隔的“定时”小脑SAGE文库；(2)制备长期酒精暴露小鼠的小脑SAGE文库；(3)全面鉴定因饮酒而使表达发生显著变化的新的小脑基因；以及(4)研究旨在逆转酒精摄入对小脑神经元毒性影响的实验策略。

DESCRIPTION (provided by applicant): Excessive consumption of alcohol is known to have toxic effects on the fetal, adult and aging nervous system. It is well established that cerebellar neurons are subject to the damaging effects of alcohol administration throughout development. Because the major function of the cerebellum is to modulate movement and balance by integrating sensory input and motor output, any injury to component neurons is likely to lead to the impairment of motor co-ordination. It is plausible that alcohol-induced cerebellar damage is the result of cumulative changes in the expression of many genes. To date, only a fraction of the genes comprising the human and mouse genomes have been assayed for alcohol-induced changes, either individually or by DNA microarrays. Because DNA microarray technology is a 'closed' system, capable of detecting only known genes, we propose to conduct a global analysis of gene expression in the alcohol-exposed mouse cerebellum using Serial Analysis of Gene Expression (SAGE). SAGE is an 'open' system capable of detecting and digitally quantifying both known and, as yet, unknown genes. The proposed experiments involve the construction, sequencing and bioinformatic analysis of routine cerebellar SAGE libraries during the early postnatal period, adulthood and senescence, shortly after exposure to "binge" amounts of alcohol. The resulting comprehensive and quantitative SAGE database of alcohol-induced changes in gene expression, which includes that of novel genes, will be disseminated via the NCBI website. Future long-term experiments, beyond the scope of this R03 and proposed as part of a planned R01 submission, will be aimed at (1) construction of "timed" cerebellar SAGE libraries at several time intervals after alcohol exposure; (2) preparation of cerebellar SAGE libraries from mice chronically exposed to alcohol; (3) the full characterization of novel cerebellar genes whose expression is substantially altered as a result of alcohol administration, and (4) the examination of experimental strategies aimed at reversing the toxic effects of alcohol ingestion on cerebellar neurons.