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Mitsuo Yamashita, Eric Glasgow, Bing-Jun Zhang, Kiyoshi Kusano, Harold Gainer, Identification of Cell-Specific Messenger Ribonucleic Acids in Oxytocinergic and Vasopressinergic Magnocellular Neurons in Rat Supraoptic Nucleus by Single-Cell Differential Hybridization, Endocrinology, Volume 143, Issue 11, 1 November 2002, Pages 4464–4476, https://doi.org/10.1210/en.2002-220516
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Abstract
Magnocellular neurons (MCNs) in the hypothalamo-neurohypophysial system synthesize high levels of the peptides oxytocin (OT) and vasopressin (VP) in separate cells. We used RT-PCR amplification of the RNA from single-cells dissected from supraoptic nuclei of lactating rats to produce cDNAs from identified OT or VP MCNs, which were used to construct OT- and VP-MCN-specific cDNA libraries. These cDNA libraries were then screened using labeled probes from the OT- and VP-cells’ amplified cDNAs. Differentially hybridized colonies were isolated and characterized by slot blot hybridization, Southern blot hybridization, DNA sequencing, and in situ hybridization histochemistry. Using this approach, several novel cell-specific mRNAs were identified in the MCNs. One cell-specific clone, phosphofructokinase-C, was isolated from the OT-cell library, and five cell-specific clones were isolated from the VP-cell library. These were identified as paternally expressed gene (Peg)5/neuronatin, metallothionein III, Peg3, synaptotagmin V, and a 3′-phosphoadenosine 5′-phosphosulfate synthase 2-related mRNA. None of these genes would have been predicted to be differentially expressed in OT and VP MCNs, based on our current knowledge; and hence, this single cell differential gene expression approach has begun to further define the MCN phenotypes by identifying selectively expressed molecules in them.
