SUPRACHIASMATIC NUCLEI-NEUROBIOLOGY OF A CIRCADIAN CLOCK

昼夜节律的视交叉上核神经生物学

• 批准号: 3409246
• 负责人: William J Schwartz
• 金额: $ 25.34万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-08-01 至 1995-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3409246
• 关键词: DNA binding protein; antisense nucleic acid; circadian rhythms; dizocilpine; environmental adaptation; experimental brain lesion; gel mobility shift assay; gene expression; immunocytochemistry; in situ hybridization; laboratory rat; melatonin; neuropeptides; neuropharmacology; northern blottings; photoactivation; photobiology; suprachiasmatic nucleus; transcription factor; visual pathways; western blottings

Circadian rhythms are the overt manifestation of an innate timekeeping system, i.e., a "circadian clock," that is synchronized (entrained) to the 24-hr day-night cycle by environmental light. The clinical and practical significance of the human clock is now well recognized. The overall objective of this research program is to understand the endogenous oscillatory mechanism of the circadian pacemaker in the suprachiasmatic nuclei (SCN) of mammals. During the current grant period, we began to study whether an alteration of gene expression might be involved in the entraining effect of light. We found that light regulates the expression of c-Fos and Fos-related transcriptional regulatory proteins in the ventrolateral SCN and in the intergeniculate leaflet (IGL) of rats. In this competing continuation application, experimental results are presented that lead to the hypothesis that events at the transcriptional level are part of the mechanism for photic entrainment of the circadian pacemaker to the light-dark cycle, and, more specifically, that Fos expression in the SCN may be a link in the entrainment mechanism. Protocols are proposed to study four questions directed to this hypothesis. (1) What are the effects of light and circadian phase on the expression of Fos, on its interaction with Jun, and on their binding to DNA? Patterns of Fos/Jun expression will be evaluated by immunohistochemistry and in situ hybridization, identification of the immunoreactive and mRNA species by Western and Northern blot analysis, and detection of sequence-specific DNA binding by mobility shift (gel retardation) assay. (2) What are the implications of the localization of Fos to the retino-recipient ventrolateral SCN and IGL? The possible colocalization of Fos to neurons that synthesize known neuropeptides and the dependence of light-induced Fos activation in the SCN on visual input from the IGL will be assessed. (3) Do stimuli that affect pacemaker entrainment modify Fos expression in the SCN? Changes of Fos expression will be investigated after injection of the N-methyl-D-aspartate antagonist MK-801, after daily injections of melatonin, and in the fetal SCN. (4) Will experimental reduction of intracellular Fos levels in the SCN inhibit the entraining action of light? An antisense oligonucleotide complementary to the 5' end of rat c-fos mRNA will be introduced by stereotaxic infusion in to the SCN via mini-osmotic pump, and its effects on light-induced phase shifts of overt behavioral rhythmicity will be measured.

昼夜节律是与生俱来的计时能力的明显表现 系统，即“生物钟”，与 环境光的 24 小时昼夜循环。 临床和 人类时钟的实际意义现已得到充分认识。 这 该研究计划的总体目标是了解 昼夜节律起搏器的内源性振荡机制 哺乳动物的视交叉上核（SCN）。 在本次赠款期间 在此期间，我们开始研究基因表达的改变是否可能 参与光的夹带效应。 我们发现那光 调节 c-Fos 和 Fos 相关转录的表达 腹外侧 SCN 和膝状体中的调节蛋白 大鼠传单（IGL）。 在这个竞争性的延续申请中， 实验结果得出以下假设： 转录水平的事件是光传递机制的一部分 昼夜节律起搏器与明暗周期的夹带，以及更多 具体来说，SCN 中的 Fos 表达可能是 夹带机制。 提议研究四个问题的方案 针对这个假设。 (1) 光线和昼夜节律相位对表情有何影响 Fos 与 Jun 的相互作用以及它们与 DNA 的结合？ Fos/Jun 表达模式将通过免疫组织化学进行评估 以及原位杂交、免疫反应性和mRNA的鉴定 通过 Western 和 Northern 印迹分析来检测物种 通过迁移率变化（凝胶阻滞）测定进行序列特异性 DNA 结合。 （2）Fos国产化对我国有何影响？ 视网膜受体腹外侧 SCN 和 IGL？可能的共定位 Fos 对合成已知神经肽的神经元的影响及其依赖性 SCN 中光诱导 Fos 激活对 IGL 视觉输入的影响 将被评估。 (3) 影响起搏器夹带的刺激会改变 Fos 的表达吗？ SCN？注射后研究Fos表达的变化 每天注射 N-甲基-D-天冬氨酸拮抗剂 MK-801 后 褪黑激素，以及胎儿 SCN。 (4) 实验性降低 SCN 细胞内 Fos 水平 抑制光的夹带作用？反义寡核苷酸 与大鼠 c-fos mRNA 5' 端互补的部分将通过以下方式引入 微型渗透泵立体定向输注至SCN及其效果 关于光诱导的明显行为节律的相移将是 测量。