Elsevier

Neuroscience

Volume 173, 26 January 2011, Pages 37-56
Neuroscience

Cellular and Molecular Neuroscience
Research Paper
Characterization of Kiss1 neurons using transgenic mouse models

https://doi.org/10.1016/j.neuroscience.2010.11.022Get rights and content

Abstract

Humans and mice with loss-of-function mutations of the genes encoding kisspeptins (Kiss1) or kisspeptin receptor (Kiss1r) are infertile due to hypogonadotropic hypogonadism. Within the hypothalamus, Kiss1 mRNA is expressed in the anteroventral periventricular nucleus (AVPV) and the arcuate nucleus (Arc). In order to better study the different populations of kisspeptin cells we generated Kiss1-Cre transgenic mice. We obtained one line with Cre activity specifically within Kiss1 neurons (line J2–4), as assessed by generating mice with Cre-dependent expression of green fluorescent protein or β-galactosidase. Also, we demonstrated Kiss1 expression in the cerebral cortex and confirmed previous data showing Kiss1 mRNA in the medial nucleus of amygdala and anterodorsal preoptic nucleus. Kiss1 neurons were more concentrated towards the caudal levels of the Arc and higher leptin-responsivity was observed in the most caudal population of Arc Kiss1 neurons. No evidence for direct action of leptin in AVPV Kiss1 neurons was observed. Melanocortin fibers innervated subsets of Kiss1 neurons of the preoptic area and Arc, and both populations expressed melanocortin receptors type 4 (MC4R). Specifically in the preoptic area, 18–28% of Kiss1 neurons expressed MC4R. In the Arc, 90% of Kiss1 neurons were glutamatergic, 50% of which also were GABAergic. In the AVPV, 20% of Kiss1 neurons were glutamatergic whereas 75% were GABAergic. The differences observed between the Kiss1 neurons in the preoptic area and the Arc likely represent neuronal evidence for their differential roles in metabolism and reproduction.

Research Highlights

▶Generation and validation of Kiss1-Cre mouse lines. ▶Kiss1 neurons in the arcuate nucleus (only) are direct target of leptin. ▶Kiss1 neurons coexpress melanocortin 4 receptor (MC4R). ▶Most of Kiss1 neurons in the AVPV/PeN (75%) are GABAergic. ▶Kiss1 neurons in the arcuate nucleus (95%) are glutamatergic.

Section snippets

Subjects

Adult male and female Kiss1-Cre, LepR-IRES-Cre/LacZ, MC4R-GFP and C57BL/6 mice were housed in the University of Texas Southwestern Medical Center Animal Resource Center, in a light- (12 h on/12 h off) and temperature- (21–23 °C) controlled environment. They were fed standard chow diet (Harlan Teklad Global Diet, Harlan Laboratories Inc., Indianapolis, IN, USA), unless otherwise mentioned and had free access to water. All experiments were carried out in accordance with the guidelines established

Production and validation of Kiss1-Cre mouse model

To generate transgenic mice that express Cre-recombinase driven by Kiss1 regulatory elements (Kiss1-Cre mouse model), we constructed two different Kiss1-Cre transgene-containing BACs (Fig. 1). We were successful in generating 14 different potential Kiss1-Cre founder mice. After crossing with reporter mice, we identified three lines with Cre activity in nuclei previously shown to express Kiss1 mRNA (Gottsch et al., 2004) (Fig. 2A, C, E, G). All three lines were originated from founders generated

Discussion

In the present study, we describe the generation and validation of a Kiss1-Cre transgenic mouse line (J2–4) which displayed Cre activity in areas known to express Kiss1 mRNA. We then generated Kiss1-reporter mice which allowed us to identify kisspeptin neurons in the mouse brain. We validated our mouse model through colocalization of the reporter gene and Kiss1 mRNA in the preoptic area (AVPV and Pe) and Arc, and confirmed the expression of Kiss1 in other sites, such as the medial nucleus of

Conclusion

In the present study we describe the generation of a novel Kiss1-Cre mouse model. This mouse model enabled us to generate reporters for Kiss1 neurons and to determine the chemical profiles of different populations of Kiss1 neurons. Future studies will take advantage of the Kiss1-specific expression of Cre recombinase to manipulate gene expression in Kiss1 neurons. This will allow us to further elucidate the mechanisms by which these neurons are influenced by changing metabolic states and by

Acknowledgments

We acknowledge the assistance of Robert Hammer and the UTSW Medical Center Transgenic Core Facility in generating the Kiss1-Cre transgenic animals, as well as of Angela Mobley and the UTSW Flow Cytometry Core Facility for the FACS procedure. We also thank Dr. Joyce Repa (UTSW) for Kiss1, Cyclophilin and GAD67 primers. We thank Dr. Jeffrey Friedman (Rockefeller University, New York) for kindly providing the LepR-IRES-Cre mice and Dr. Joel K. Elmquist (UTSW Medical Center, Dallas-TX) for kindly

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    JMZ and CFE contributed equally to this work.

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