Geriatric Research, Education and Clinical Center (GRECC), Veterans Affairs Medical Center, Seattle, Washington 98108, USA; Department of Pharmacology, University of Washington, Seattle, Washington 98195, USA.
Mol Cell Neurosci. 1992 Jun;3(3):224-36. doi: 10.1016/1044-7431(92)90042-z.
In situ hybridization was performed in Long-Evans (LE) and heterozygous (HET) and homozygous (HOM) Brattleboro rats using a 48-base oligonucleotide complementary to the last 16 amino acids of the vasopressin messenger RNA (VP mRNA). The number of cells expressing the VP mRNA in the supraoptic (SON) and paraventicular (PVN) nuclei was not significantly different between LE, HET, and HOM rats; however, the relative amount of mutant VP mRNA expressed in neurons of the PVN and SON of the HOM rat was significantly lower than that in the other two genotypes. In contrast, the suprachiasmatic nucleus (SCN), the bed nucleus of the stria terminalis (BNST), and the medial amygdala (MA) of the HOM rat showed a significant reduction in both the number of neurons and the level of mutant VP mRNA expression per cell compared to those in either the LE or the HET rat. To determine whether the reduced level of mutant VP mRNA in the HOM rat was due to decreased transcription, in situ hybridization to detect the VP primary transcript was performed. The number of neurons and the amount of nuclear VP RNA expressed per cell for the PVN, SCN, BNST, and MA in the HET and HOM rats were not significantly different from those in the LE rat. However, in the SON, the HOM rat exhibited a significant increase (P < 0.10) in the amount of nuclear VP RNA expressed per cell compared to the LE and HET rats, but the number of positively labeled cells was not significantly different. Therefore, these data suggest that the mutant gene in the HOM rat is transcribed at rates comparable to those in the LE and HET rats, while cytoplasmic mutant VP mRNA is significantly reduced in all nuclei of the HOM rat. The reduced cytoplasmic VP mRNA in the HOM rat may be attributed to instability (degradation) of the mutant message.
原位杂交在长耳 (LE) 和杂合子 (HET) 和纯合子 (HOM) 布拉特雷伯大鼠中使用与最后 16 个氨基酸的血管加压素信使 RNA (VP mRNA) 互补的 48 个碱基的寡核苷酸进行。在 LE、HET 和 HOM 大鼠中，表达 VP mRNA 的细胞数量在视上核 (SON) 和室旁核 (PVN) 中没有显著差异;然而，在 HOM 大鼠的 PVN 和 SON 神经元中表达的突变 VP mRNA 的相对量明显低于其他两种基因型。相比之下，HOM 大鼠的视交叉上核 (SCN)、终纹床核 (BNST) 和内侧杏仁核 (MA) 中神经元的数量和每个细胞表达的突变 VP mRNA 水平均显著降低与 LE 或 HET 大鼠相比。为了确定 HOM 大鼠中突变 VP mRNA 水平降低是否是由于转录减少，进行了原位杂交以检测 VP 初级转录物。PVN、SCN、BNST 和 MA 中 HET 和 HOM 大鼠的神经元数量和每个细胞表达的核 VP RNA 量与 LE 大鼠无显著差异。然而，在 SON 中，与 LE 和 HET 大鼠相比，HOM 大鼠的每个细胞表达的核 VP RNA 量显著增加 (P < 0.10)，但阳性标记细胞的数量没有显著差异。因此，这些数据表明，HOM 大鼠中的突变基因以与 LE 和 HET 大鼠相当的速率转录，而所有 HOM 大鼠核中的细胞质突变 VP mRNA 显著减少。HOM 大鼠细胞质中 VP mRNA 的减少可能归因于突变信息的不稳定性 (降解)。
