BAC transgenic mice to study the expression of P2X2 and P2Y1 receptors

Purinergic Signalling, vol. 17, pp. 449–465

Abstract

Abstract Extracellular purines are important signaling molecules involved in numerous physiological and pathological processes via the activation of P2 receptors. Information about the spatial and temporal P2 receptor (P2R) expression and its regulation remains crucial for the understanding of the role of P2Rs in health and disease. To identify cells carrying P2X2Rs in situ, we have generated BAC transgenic mice that express the P2X2R subunits as fluorescent fusion protein (P2X2-TagRFP). In addition, we generated a BAC P2Y1R TagRFP reporter mouse expressing a TagRFP reporter for the P2RY1 gene expression. We demonstrate expression of the P2X2R in a subset of DRG neurons, the brain stem, the hippocampus, as well as on Purkinje neurons of the cerebellum. However, the weak fluorescence intensity in our P2X2R-TagRFP mouse precluded tracking of living cells. Our P2Y1R reporter mice confirmed the widespread expression of the P2RY1 gene in the CNS and indicate for the first time P2RY1 gene expression in mouse Purkinje cells, which so far has only been described in rats and humans. Our P2R transgenic models have advanced the understanding of purinergic transmission, but BAC transgenic models appeared not always to be straightforward and permanent reliable. We noticed a loss of fluorescence intensity, which depended on the number of progeny generations. These problems are discussed and may help to provide more successful animal models, even if in future more versatile and adaptable nuclease-mediated genome-editing techniques will be the methods of choice.

Authors 14

1. Marcus Grohmann

   Leipzig University

   Affiliation as printed

   HTK Hygiene Technologie Kompetenzzentrum GmbH, Bamberg, Germany

   Rudolf Boehm Institute of Pharmacology and Toxicology, University of Leipzig, Leipzig, Germany

2. Michaela Schumacher Aachen Institute of Clinical Pharmacology

   RWTH Aachen University

   Affiliation as printed

   Institute of Clinical Pharmacology, RWTH Aachen University, Wendlingweg 2, 52074, Aachen, Germany

   Institute of Clinical Pharmacology, RWTH Aachen University, Aachen, Germany

3. Janka Günther Aachen Institute of Clinical Pharmacology

   RWTH Aachen University

   Affiliation as printed

   Institute of Clinical Pharmacology, RWTH Aachen University, Wendlingweg 2, 52074, Aachen, Germany

   Institute of Clinical Pharmacology, RWTH Aachen University, Aachen, Germany

4. Stefan M. Singheiser Aachen Institute of Clinical Pharmacology

   RWTH Aachen University

   Affiliation as printed

   Institute of Clinical Pharmacology, RWTH Aachen University, Wendlingweg 2, 52074, Aachen, Germany

   Institute of Clinical Pharmacology, RWTH Aachen University, Aachen, Germany

5. Tanja Nussbaum Aachen Institute of Clinical Pharmacology

   RWTH Aachen University

   Affiliation as printed

   Institute of Clinical Pharmacology, RWTH Aachen University, Wendlingweg 2, 52074, Aachen, Germany

   Institute of Clinical Pharmacology, RWTH Aachen University, Aachen, Germany

6. Florian Wildner

   Charité - Universitätsmedizin Berlin

   Affiliation as printed

   Institute of Neurophysiology, Charité-Universitätsmedizin Berlin, Berlin, Germany

7. Zoltán Gerevich

   Charité - Universitätsmedizin Berlin

   Affiliation as printed

   Institute of Neurophysiology, Charité-Universitätsmedizin Berlin, Berlin, Germany

8. Ronald Jabs

   University of Bonn

   Affiliation as printed

   Institute of Cellular Neurosciences, Medical Faculty, University of Bonn, Bonn, Germany

9. Daniela Hirnet

   Universität Hamburg

   Affiliation as printed

   Division of Neurophysiology, University of Hamburg, Hamburg, Germany

10. Christian Lohr

    Universität Hamburg

    Affiliation as printed

    Division of Neurophysiology, University of Hamburg, Hamburg, Germany

11. Péter Illés

    Leipzig University

    Affiliation as printed

    Rudolf Boehm Institute of Pharmacology and Toxicology, University of Leipzig, Leipzig, Germany

12. Günther Schmalzing Aachen Institute of Clinical Pharmacology

    RWTH Aachen University

    Affiliation as printed

    Institute of Clinical Pharmacology, RWTH Aachen University, Wendlingweg 2, 52074, Aachen, Germany

    Institute of Clinical Pharmacology, RWTH Aachen University, Aachen, Germany

13. Heike Franke

    Leipzig University

    Affiliation as printed

    Rudolf Boehm Institute of Pharmacology and Toxicology, University of Leipzig, Leipzig, Germany

14. Ralf Hausmann corresponding Aachen Institute of Clinical Pharmacology

    RWTH Aachen University

    Affiliation as printed

    Institute of Clinical Pharmacology, RWTH Aachen University, Wendlingweg 2, 52074, Aachen, Germany. rhausmann@ukaachen.de

    Institute of Clinical Pharmacology, RWTH Aachen University, Aachen, Germany