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Single Cell Reverse Transcription-Polymerase Chain Reaction Analysis of Rat Supraoptic Magnocellular Neurons: Neuropeptide Phenotypes and High Voltage-Gated Calcium Channel Subtypes

Laboratory of Neurochemistry, Basic Neuroscience Program, National Institute of Neurological Disorders and Stroke (E.G., K.K., H.C., H.G.), In Situ Facility, BNP, NINDS (E.M.), and the Section on Neural Gene Expression, National Institute of Mental Health (W.S.Y.), Bethesda, Maryland 20892

Address all correspondence and requests for reprints to: Harold Gainer, Ph.D., Laboratory of Neurochemistry, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Building 36, Room 4D20, Bethesda, Maryland 20892. E-mail: hgatnih{at}codon.nih.gov

Magnocellular neurosecretory cells (MNCs) in the hypothalamo-neurohypophysial system that express and secrete the nonapeptides oxytocin (OT) and vasopressin (VP) were evaluated for the expression of multiple genes in single magnocellular neurons from the rat supraoptic nucleus using a single cell RT-PCR protocol. We found that all cells representing the two major phenotypes, the OT and VP MNCs, express a small, but significant, amount of the other nonapeptide’s messenger RNA (mRNA). In situ hybridization histochemical analyses confirmed this observation. A third phenotype, containing equivalent amounts of OT and VP mRNA, was detected in about 19% of the MNCs from lactating female supraoptic nuclei. Analyses of these phenotypes for other coexisting peptide mRNAs (e.g. CRH, cholecystokinin, galanin, dynorphin, and the calcium-binding protein, calbindin) generally confirmed expectations from the literature, but revealed cell to cell variation in their coexpression. Our results also show that the high voltage-activated calcium channel subunit genes, 1A-D, 2, and ß1–4 are expressed in virtually all MNCs. However, the 1E subunit gene is not expressed at detectable levels in these cells. The expression of all of the ß-subunit genes in each MNC may account for the variations in physiological and pharmacological properties of the high voltage-activated channels found in these neurons.

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