Knock-in mouse models for studying somatostatin and cholecystokinin expressing cells

Marta Balog; Allison Anderson; Channabasavaiah B Gurumurthy; Rolen M Quadros; Zeljka Korade; Karoly Mirnics

doi:10.1016/j.jneumeth.2022.109704

Knock-in mouse models for studying somatostatin and cholecystokinin expressing cells

J Neurosci Methods. 2022 Nov 1:381:109704. doi: 10.1016/j.jneumeth.2022.109704. Epub 2022 Sep 5.

Authors

Marta Balog¹, Allison Anderson², Channabasavaiah B Gurumurthy³, Rolen M Quadros⁴, Zeljka Korade⁵, Karoly Mirnics⁶

Affiliations

¹ Munroe-Meyer Institute for Genetics and Rehabilitation, University of Nebraska Medical Center Omaha, NE, USA; Department of Medical Biology and Genetics, Faculty of Medicine, University of Osijek, Osijek, Croatia.
² Munroe-Meyer Institute for Genetics and Rehabilitation, University of Nebraska Medical Center Omaha, NE, USA.
³ Mouse Genome Engineering Core Facility, University of Nebraska Medical Center, Omaha, NE, USA; Department of Pharmacology and Experimental Neuroscience, College of Medicine, University of Nebraska Medical Center, Omaha, NE, USA.
⁴ Mouse Genome Engineering Core Facility, University of Nebraska Medical Center, Omaha, NE, USA.
⁵ Department of Pediatrics, University of Nebraska Medical Center Omaha, NE, USA; Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center Omaha, NE, USA; Child Health Research Institute, University of Nebraska Medical Center Omaha, NE, USA. Electronic address: zeljka.korade@unmc.edu.
⁶ Munroe-Meyer Institute for Genetics and Rehabilitation, University of Nebraska Medical Center Omaha, NE, USA; Department of Pharmacology and Experimental Neuroscience, College of Medicine, University of Nebraska Medical Center, Omaha, NE, USA; Department of Pediatrics, University of Nebraska Medical Center Omaha, NE, USA; Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center Omaha, NE, USA; Child Health Research Institute, University of Nebraska Medical Center Omaha, NE, USA. Electronic address: karoly.mirnics@unmc.edu.

PMID: 36070817
DOI: 10.1016/j.jneumeth.2022.109704

Abstract

Background: Somatostatin (SST) and cholecystokinin (CCK) are peptide hormones that regulate the endocrine system, cell proliferation and neurotransmission.

New method: We utilized the novel Easi-CRISPR system to generate two knock-in mouse strains with Cre recombinase in SST- and CCK-expressing cells and validated their utility in the developing and adult brain tissues.

Results: The full nomenclature for the newly generated strains are C57BL/6-Sst^{em1(P2A-iCre-T2A-mCherry)Mirn} and C57BL/6-Cck^{em1(iCre-T2A-mCherry-P2A)Mirn}. For the Sst locus, a P2A-iCre-T2A-mCherry cassette was inserted immediately upstream of the stop codon (C terminus fusion). For the Cck locus, iCre-P2A-mCherry-T2A cassette was inserted at the start codon (N terminus fusion). Knock-in mice were generated using the Easi-CRISPR method. Developmental and adult SST and CCK expressions were preserved and showed an appropriate expression pattern in both models, with an active fluorescent tag in both animal lines.

Comparison with existing methods: Knock-in mouse models to study cell types that produce these critically important molecules are limited to date. The knock-in mice we generated can be used as reporters to study development, physiology, or pathophysiology of SST and CCK expressing cells - without interference with native expression of SST and CCK. In addition, they can be used as Cre driver models to conditionally delete floxed genes in SST and CCK expressing cells across various tissues.

Conclusions: These two mouse models serve as valuable tools for in vitro and in vivo research studies related to SST and CCK biology across the lifespan and across different tissue types.

Keywords: CRISPR/Cas9; Cholecystokinin; Cre; Easi-CRISPR; Somatostatin; ZsGreen1.

Publication types

Research Support, N.I.H., Extramural

MeSH terms

Animals
Cholecystokinin* / genetics
Codon, Initiator
Codon, Terminator
Mice
Mice, Inbred C57BL
Mice, Transgenic
Somatostatin* / genetics

Substances

Codon, Initiator
Codon, Terminator
Somatostatin
Cholecystokinin

Grants and funding

R01 MH067234/MH/NIMH NIH HHS/United States