Analysis of Transcription Factor mRNAs in Identified Oxytocin and Vasopressin Magnocellular Neurons Isolated by Laser Capture Microdissection |
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| Authors: | Madison Humerick Jeffrey Hanson Jaime Rodriguez-Canales Daniel Lubelski Omar M. Rashid Yasmmyn D. Salinas YiJun Shi Todd Ponzio Raymond Fields Michael R. Emmert-Buck Harold Gainer |
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| Affiliation: | 1. Laboratory of Neurochemistry, National Institute of Neurological Disease and Stroke, National Institutes of Health, Bethesda, Maryland, United States of America.; 2. Laser Microdissection Core, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.; Pisces Therapeutics, United States of America, |
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| Abstract: | The oxytocin (Oxt) and vasopressin (Avp) magnocellular neurons (MCNs) in the hypothalamus are the only neuronal phenotypes that are present in the supraoptic nucleus (SON), and are characterized by their robust and selective expression of either the Oxt or Avp genes. In this paper, we take advantage of the differential expression of these neuropeptide genes to identify and isolate these two individual phenotypes from the rat SON by laser capture microdissection (LCM), and to analyze the differential expression of several of their transcription factor mRNAs by qRT-PCR. We identify these neuronal phenotypes by stereotaxically injecting recombinant Adeno-Associated Viral (rAAV) vectors which contain cell-type specific Oxt or Avp promoters that drive expression of EGFP selectively in either the Oxt or Avp MCNs into the SON. The fluorescent MCNs are then dissected by LCM using a novel Cap Road Map protocol described in this paper, and the purified MCNs are extracted for their RNAs. qRT-PCR of these RNAs show that some transcription factors (RORA and c-jun) are differentially expressed in the Oxt and Avp MCNs. |
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