Yue Chunmei, Mutsuga Noriko, Scordalakes Elka M, Gainer Harold
Molecular Neuroscience Section, Laboratory of Neurochemistry, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892, USA.
Am J Physiol Regul Integr Comp Physiol. 2006 May;290(5):R1233-41. doi: 10.1152/ajpregu.00709.2005. Epub 2005 Dec 15.
To develop a comprehensive approach for the study of oxytocin (OT) and vasopressin (VP) gene expression in the rat hypothalamus, we first developed an intronic riboprobe to measure OT heteronuclear RNA (hnRNA) levels by in situ hybridization histochemistry (ISHH). Using this 84-bp riboprobe, directed against intron 2 of the OT gene, we demonstrate strong and specific signals in neurons confined to the supraoptic (SON) and paraventricular (PVN) nuclei of the rat hypothalamus. We used this new intronic OT probe, together with other well-established intronic and exonic OT and VP probes, to reevaluate OT and VP gene expression in the hypothalamus under two classical physiological conditions, acute osmotic stimulation, and lactation. We found that magnocellular neurons in 7- to 8-day lactating female rats exhibit increased OT but not VP hnRNA. Since VP mRNA is increased during lactation, this suggests that decreased VP mRNA degradation during lactation may be responsible for this change. In contrast, whereas there was the expected large increase in VP hnRNA after acute salt loading, there was no change in OT hnRNA, suggesting that acute hyperosmotic stimuli produce increased VP but not OT gene transcription. Hence, the use of both exon- and intron-specific probes, which distinguish the changes in hnRNA and mRNA levels, respectively, can provide insight into the relative roles of transcription and mRNA degradation processes in changes in gene expression evoked by physiological stimuli.
为了开发一种全面的方法来研究大鼠下丘脑催产素（OT）和加压素（VP）基因的表达，我们首先开发了一种内含子核糖探针，通过原位杂交组织化学（ISHH）来测量OT异核RNA（hnRNA）水平。使用这种针对OT基因内含子2的84个碱基对的核糖探针，我们在局限于大鼠下丘脑视上核（SON）和室旁核（PVN）的神经元中证明了强烈而特异的信号。我们使用这种新的内含子OT探针，连同其他成熟的内含子和外显子OT及VP探针，在两种经典生理条件下，即急性渗透刺激和泌乳，重新评估下丘脑OT和VP基因的表达。我们发现，7至8日龄泌乳雌性大鼠的大细胞神经元中OT hnRNA增加，但VP hnRNA没有增加。由于泌乳期间VP mRNA增加，这表明泌乳期间VP mRNA降解减少可能是造成这种变化的原因。相比之下，急性盐负荷后VP hnRNA如预期大幅增加，而OT hnRNA没有变化，这表明急性高渗刺激会使VP基因转录增加，但不会使OT基因转录增加。因此，分别区分hnRNA和mRNA水平变化的外显子特异性和内含子特异性探针的使用，可以深入了解转录和mRNA降解过程在生理刺激引起的基因表达变化中的相对作用。
