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Mol Cell Endocrinol. 2012 Mar 5;350(1):10-9. doi: 10.1016/j.mce.2011.11.017. Epub 2011 Nov 25.

Optimized amplification and single-cell analysis identify GnRH-mediated activation of Rap1b in primary rat gonadotropes.

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  • 1Center for Translational Systems Biology and Department of Neurology, Mount Sinai School of Medicine, New York, NY 10029, United States.

Abstract

Identifying the early gene program induced by GnRH would help understand how GnRH-activated signaling pathways modulate gonadotrope secretory response. We previously analyzed GnRH-induced early genes in LβT2 cells, however these lack GnRH self-potentiation, a physiological attribute of gonadotropes. To minimize cellular heterogeneity, rat primary pituitary cultures were enriched for gonadotropes by 40-60% using a sedimentation gradient. Given the limited number of gonadotropes, RNA was amplified prior to microarray analysis. Thirty-three genes were up-regulated 40 min after GnRH stimulation. Real-time PCR confirmed regulation of several transcripts including fosB, c-fos, egr-2 and rap1b, a small GTPase and member of the Ras family. GnRH stimulated rap1b gene expression in gonadotropes, measured by a sensitive single cell assay. Immunocytochemistry revealed increased Rap1 protein in GnRH-stimulated gonadotropes. These data establish rap1b as a novel gene rapidly induced by GnRH and a candidate to modulate gonadotropin secretion in rat gonadotropes.

Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

PMID:
22127306
[PubMed - indexed for MEDLINE]
PMCID:
PMC3919063
Free PMC Article

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