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The Journal of Neuroscience, April 20, 2005, 25(16):4181-4188; doi:10.1523/JNEUROSCI.0158-05.2005

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Cellular/Molecular
Use of Laser-Capture Microdissection for the Identification of Marker Genes for the Ventromedial Hypothalamic Nucleus

Jeremy P. Segal,¹ Nancy R. Stallings,² Charlotte E. Lee,³ Liping Zhao,² Nicholas Socci,⁵ Agnes Viale,⁴ Thomas M. Harris,⁶ Marcelo B. Soares,⁷ Geoffrey Childs,⁶ Joel K. Elmquist,³ Keith L. Parker,² and Jeffrey M. Friedman¹

¹The Rockefeller University, New York, New York 10021, ²University of Texas Southwestern Medical Center at Dallas, Dallas, Texas 75390-8857, ³Beth Israel Deaconess Medical Center, Boston, Massachusetts 02215, ⁴Department of Molecular Biology, ⁵Memorial Sloan-Kettering Cancer Center, New York, New York 10021, ⁶Albert Einstein College of Medicine, Bronx, New York 10461, and ⁷Department of Pediatrics, University of Iowa Health Care, Iowa City, Iowa 52242

The ventromedial hypothalamic nucleus (VMH) plays an important role in the control of feeding and energy homeostasis. In contrast to other hypothalamic nuclei that are also known to regulate energy balance, there is a paucity of nucleus-specific marker genes for the VMH, limiting the application of molecular approaches for analyzing VMH information processing, function, and circuitry. Here, we report the use of laser-capture microdissection to isolate a set of cDNAs that are enriched in the VMH relative to two adjacent hypothalamic nuclei, the arcuate and dorsomedial hypothalamus. The relative expression levels of nine of the 12 most robustly expressed VMH-enriched genes were confirmed by real-time PCR analysis using separate RNAs from these three nuclei. Three of these VMH-enriched genes were further characterized by in situ hybridization histochemistry, including pituitary adenylate cyclase activating polypeptide, cerebellin 1, and an expressed sequence tag named LBH2. Finally, to test whether some of these genes were coordinately regulated, we monitored their expression in steroidogenic factor 1 (SF-1) knock-out mice. SF-1 is a transcription factor that controls the development of the VMH. The RNA levels for four of these genes were reduced in these knock-out animals, further suggesting that they are direct or indirect targets of this orphan nuclear receptor. The VMH-enriched genes identified here provide a basis for a functional analysis of VMH neuronal subpopulations via the use of bacterial artificial chromosome transgenics and related technologies. These results also demonstrate the utility of laser-capture microdissection coupled with microarray technology to identify nucleus-specific transcriptional networks.

Key words: ventromedial; hypothalamus; marker; VMH; expression; microarray; LCM; microdissection

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Received Jan 12, 2005; revised March 8, 2005; accepted March 9, 2005.

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