Identifying the efferent projections of leptin-responsive neurons in the dorsomedial hypothalamus using a novel conditional tracing approach

Laurent Gautron, Michael Lazarus, Michael M. Scott, Clifford B. Saper, Joel K. Elmquist

Research output: Contribution to journalArticle

56 Citations (Scopus)

Abstract

Tracing the axonal projections of selected neurons is labor intensive and inherently limited by currently available neuroanatomical methods. We developed an adeno-associated virus (AAV) that can be used for efficiently tracing identified neuronal populations. The virus encodes a humanized Renilla green fluorescent protein (hrGFP) that is transcriptionally silenced by a neo cassette flanked by LoxH/LoxP sites (AAV-lox-Stop-hrGFP). Thus, hrGFP is expressed only in neurons with Cre recombinase activity. To demonstrate the utility of this approach, the virus was injected unilaterally into the dorsomedial hypothalamus (DMH) of mice that express Cre in neurons expressing the leptin receptor. Animals with DMH injections showed robust hrGFP expression in DMH neurons, as visualized by its endogenous fluorescence or following immunolabeling. We found that hrGFP was expressed in approximately one-third to one-half of Cre-expressing neurons at the site of injection, but not in non-Cre-expressing neurons. The expression of GFP allowed us to identify the projection fields of DMH leptin-responsive neurons. Our results show hrGFP-positive axonal projections and terminals in the paraventricular nucleus of the hypothalamus, arcuate nucleus, preoptic area, bed nucleus of the stria terminalis, paraventricular thalamus, periaqueductal gray, and precoeruleus. The aforementioned pattern of projections was similar to DMH projections determined by injections of biotinylated dextran amine in the mouse DMH. Interestingly, some hrGFP-positive terminals were seen contacting the ependymal layer of the third and fourth ventricles. In summary, this approach is an effective tool for tracing axonal projections of chemically identified neurons, including leptin-responsive neurons.

Original languageEnglish (US)
Pages (from-to)2090-2108
Number of pages19
JournalJournal of Comparative Neurology
Volume518
Issue number11
DOIs
StatePublished - Jun 1 2010

Fingerprint

Renilla
Leptin
Hypothalamus
Green Fluorescent Proteins
Neurons
Dependovirus
Injections
Viruses
Fourth Ventricle
Leptin Receptors
Septal Nuclei
Periaqueductal Gray
Arcuate Nucleus of Hypothalamus
Third Ventricle
Preoptic Area
Paraventricular Hypothalamic Nucleus
Thalamus
Fluorescence

Keywords

  • Adeno-associated virus
  • Cre-lox
  • Green fluorescent protein
  • Mouse

ASJC Scopus subject areas

  • Neuroscience(all)

Cite this

Identifying the efferent projections of leptin-responsive neurons in the dorsomedial hypothalamus using a novel conditional tracing approach. / Gautron, Laurent; Lazarus, Michael; Scott, Michael M.; Saper, Clifford B.; Elmquist, Joel K.

In: Journal of Comparative Neurology, Vol. 518, No. 11, 01.06.2010, p. 2090-2108.

Research output: Contribution to journalArticle

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