Uhl G R, Zingg H H, Habener J F
Proc Natl Acad Sci U S A. 1985 Aug;82(16):5555-9. doi: 10.1073/pnas.82.16.5555.
Hybridizable vasopressin mRNA may be quantitatively localized in situ in sections from rat hypothalamus. Radiolabeled oligonucleotide cDNA probes, synthesized by chemical and enzymatic means, provide strong hybridization in zones known to contain vasopressin cell bodies. Multiple single-stranded 32P-, 35S-, or 3H-labeled oligonucleotides demonstrate localized hybridization that increases as probes are lengthened from 8 to 75 bases. Competition studies, RNase experiments, anatomic localization, and use of multiple probes all support hybridization specificity. An approximate doubling of hybridizable mRNA in both supraoptic and paraventricular nuclei can be detected with dehydration of the animals. Hybridizable mRNA densities are virtually normal in hypothalamic nuclei of Brattleboro rats given free access to water. These methods can provide insight into regional mRNA dynamics and may reflect functional activity of peptidergic neurons.
可杂交的血管加压素信使核糖核酸(mRNA)可在大鼠下丘脑切片中进行原位定量定位。通过化学和酶促方法合成的放射性标记寡核苷酸互补脱氧核糖核酸(cDNA)探针,在已知含有血管加压素细胞体的区域能产生强烈杂交信号。多个单链的32P、35S或3H标记的寡核苷酸显示出局部杂交,随着探针从8个碱基延长至75个碱基,杂交信号增强。竞争研究、核糖核酸酶实验、解剖定位以及使用多个探针均支持杂交特异性。动物脱水后,视上核和室旁核中可杂交的mRNA含量大约会翻倍。能自由饮水的布拉特洛维大鼠下丘脑核中,可杂交的mRNA密度实际上是正常的。这些方法能深入了解区域mRNA动态变化,可能反映肽能神经元的功能活动。