Obesity and hunger are two closely related physiological states involving complex neural and hormonal regulatory mechanisms. In studying these metabolic mechanisms, NPY/AgRP neurons that are sensitive to hunger signals play an important role. Today, we are introducing the Agrp-IRES-CreERT2-P2A-tdTomato cre inducible fluorescent reporter mouse model (Product Number: C001558), specifically designed to target the arcuate nucleus of the hypothalamus (ARH), a.k.a. the hypothalamic arcuate nucleus (ARC).

NPY/AgRP Neurons in Hunger Regulation

NPY/AgRP neurons in the brain are found exclusively in the ARC, where they are highly active when hungry because they express receptors for leptin and insulin and are inhibited by the presence of these hormones. These neurons effectively stimulate appetite by increasing neuropeptide Y (NPY) signaling and also by releasing Agouti-related protein (AgRP), which inhibits melanocortin signaling to produce peptide substances.[1] The Agouti-related protein (AgRP), encoded by the AGRP gene, is a neuropeptide primarily expressed in the ARC, as well as in the kidneys and adrenal glands. Within the ventral part of the hypothalamic ARC, AgRP is synthesized solely in NPY-containing cell bodies and is co-expressed with NPY, playing a role in increasing appetite, reducing metabolism, and lowering energy expenditure. These factors make AgRP one of the most effective and long-lasting appetite stimulants.

Figure 1. NPY/AgRP neurons are located within the arcuate nucleus of the hypothalamus (ARC).[2]

Agrp-IRES-CreERT2-P2A-tdTomato Mouse Model: Genetic Background and Functionality

Implementing our gene editing technologies, Cyagen has developed the Agrp-IRES-CreERT2-P2A-tdTomato Cre conditional fluorescent reporter mouse model (Product Number: C001558), which expresses tamoxifen-inducible CreERT2 recombinase under the control of endogenous Agrp mouse gene regulatory elements. Additionally, the CreERT2 recombinase expression element in this strain is followed by a red fluorescent protein (tdTomato) expression element, enabling lineage tracing of Agrp-positive cells.

In the absence of tamoxifen treatment, the CreERT2 recombinase remains in the cytoplasm. Following tamoxifen treatment, the CreERT2 recombinase enters the nucleus and exerts its recombinase function. When Agrp-IRES-CreERT2-P2A-tdTomato mice are crossed with mice carrying loxP sites, tamoxifen induction can trigger Cre recombinase mediated cassette exchange (RMCE) between loxP sites in AgRP-positive neurons in the offspring. This approach enables the development of conditional CreERT2 inducible, fluorescently-labeled mouse models for obesity research related to NPY/AgRP.

Successful Induction of Fluorescent Protein Expression in the Hypothalamus

In breeding experiments, Agrp-IRES-CreERT2-P2A-tdTomato mice were crossed with LSL_ZsGreen mice, which conditionally expresses ZsGreen fluorescent protein. Following tamoxifen induction, hypothalamic tissues were collected from the offspring and examined for autofluorescence of ZsGreen protein using frozen sections. The results showed that in tamoxifen-treated Agrp-IRES-CreERT2-P2A-tdTomato; LSL_ZsGreen double-positive mice, spontaneous green fluorescence signals were observed specifically in the arcuate nucleus (ARC) region of the hypothalamus.

Figure 2. Expression of Cre recombinase in the hypothalamus.

Co-localized Expression of tdTomato and ZsGreen Fluorescent Proteins

Immunofluorescence (IF) staining for tdTomato protein expression was performed to identify the cell types specificity of this cross-bred mouse model. The results showed that in tamoxifen-treated Agrp-IRES-CreERT2-P2A-tdTomato; LSL_ZsGreen double-positive mice, both tdTomato (red fluorescent protein) and ZsGreen (green fluorescent protein) signals were detected in the hypothalamus. The Cre recombinase-induced ZsGreen signal was co-localized with the endogenous Agrp-regulated tdTomato fluorescence, indicating that the expression pattern of Cre recombinase in this model aligns with the endogenous Agrp expression in mice.

Figure 3. Co-localization of ZsGreen (green fluorescent protein) and tdTomato (red fluorescent protein) expression in the hypothalamus.

Conclusion

The Agrp-IRES-CreERT2-P2A-tdTomato mouse model (Catalog No.: C001558) enables precise Cre recombinase expression within the arcuate nucleus of the hypothalamus (ARC). Tamoxifen-induced double-fluorescent protein expression in cross-breeding experiments confirms the specificity of the mouse model, making it an excellent tool for obesity research focused on the AgRP gene and NPY/AgRP neurons.

Comprehensive Cre Driver and KO/cKO Mouse Models

Beyond this inducible model, Cyagen is committed to expanding our library of pre-developed rodent models to serve as a comprehensive resource for researchers. Cyagen’s Knockout Catalog Models Repository (eBank) provides a selection of knockout (KO) and conditional knockout (cKO) lines available for diverse research applications. For more information on Cyagen’s extensive selection of KO/cKO mouse models, explore our Knockout Catalog Models Repository (eBank).

In addition to our KO/cKO mouse model eBank, we offer a robust lineup of specialized Cre driver mouse lines, including iCre & CreERT2, to support precise genetic modifications in various tissues. >>Checking our Cre mouse lines

 

 

References:

[1] Morton GJ, Schwartz MW. The NPY/AgRP neuron and energy homeostasis. Int J Obes Relat Metab Disord. 2001 Dec;25 Suppl 5:S56-62.

[2] Han J, Liang X, Guo Y, Wu X, Li Z, Hong T. Agouti-related protein as the glucose signaling sensor in the central melanocortin circuits in regulating fish food intake. Front Endocrinol (Lausanne). 2022 Nov 1;13:1010472.

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