cAMP- and Ca2+/calmodulin-dependent protein kinases mediate inotropic, lusitropic and arrhythmogenic effects of urocortin 2 in mouse ventricular myocytes

Authors

  • Li-Zhen Yang,

    1. Molecular Neuroendocrinology Group, Max Planck Institute for Experimental Medicine, Goettingen, Germany
    2. Division of Endocrinology and Metabolism, School of Medicine, University of California, San Diego, CA, USA
    3. Specialized Neuroscience Research Program 2 of the John A. Burns School of Medicine of the University of Hawaii at Manoa, Honolulu, HI, USA
    4. Division of Endocrinology, Department of Internal Medicine, Shanghai Ninth People's Hospital of Shanghai Jiaotong University, Shanghai, China
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    • These authors contributed equally to this work.

  • Jens Kockskämper,

    1. Division of Cardiology, Medical University of Graz, Auenbruggerplatz, Graz, Austria
    2. Institute of Pharmacology and Clinical Pharmacy, Philipps-University of Marburg, Marburg, Germany
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    • These authors contributed equally to this work.

  • Shelina Khan,

    1. Division of Cardiology, Medical University of Graz, Auenbruggerplatz, Graz, Austria
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  • Jorge Suarez,

    1. Division of Endocrinology and Metabolism, School of Medicine, University of California, San Diego, CA, USA
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  • Stefanie Walther,

    1. Division of Cardiology, Medical University of Graz, Auenbruggerplatz, Graz, Austria
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  • Bernhard Doleschal,

    1. Division of Cardiology, Medical University of Graz, Auenbruggerplatz, Graz, Austria
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  • Gregor Unterer,

    1. Division of Cardiology, Medical University of Graz, Auenbruggerplatz, Graz, Austria
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  • Mounir Khafaga,

    1. Division of Cardiology, Medical University of Graz, Auenbruggerplatz, Graz, Austria
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  • Heinrich Mächler,

    1. Division of Cardiac Surgery, Medical University of Graz, Auenbruggerplatz, Graz, Austria
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  • Frank R. Heinzel,

    1. Division of Cardiology, Medical University of Graz, Auenbruggerplatz, Graz, Austria
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  • Wolfgang H. Dillmann,

    1. Division of Endocrinology and Metabolism, School of Medicine, University of California, San Diego, CA, USA
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  • Burkert Pieske,

    Corresponding author
    1. Division of Cardiology, Medical University of Graz, Auenbruggerplatz, Graz, Austria
      Professor Dr Burkert Pieske, Medical University of Graz, Division of Cardiology, Auenbruggerplatz 15, A-8036 Graz, Austria. E-mail: burkert.pieske@medunigraz.at
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    • These authors contributed equally to this work.

  • Joachim Spiess

    1. Molecular Neuroendocrinology Group, Max Planck Institute for Experimental Medicine, Goettingen, Germany
    2. Specialized Neuroscience Research Program 2 of the John A. Burns School of Medicine of the University of Hawaii at Manoa, Honolulu, HI, USA
    3. Sanford Burnham Medical Research Institute, La Jolla, CA, USA
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    • These authors contributed equally to this work.


Professor Dr Burkert Pieske, Medical University of Graz, Division of Cardiology, Auenbruggerplatz 15, A-8036 Graz, Austria. E-mail: burkert.pieske@medunigraz.at

Abstract

BACKGROUND AND PURPOSE Urocortin 2 is beneficial in heart failure, but the underlying cellular mechanisms are not completely understood. Here we have characterized the functional effects of urocortin 2 on mouse cardiomyocytes and elucidated the underlying signalling pathways and mechanisms.

EXPERIMENTAL APPROACH Mouse ventricular myocytes were field-stimulated at 0.5 Hz at room temperature. Fractional shortening and [Ca2+]i transients were measured by an edge detection and epifluorescence system respectively. Western blots were carried out on myocyte extracts with antibodies against total phospholamban (PLN) and PLN phosphorylated at serine-16.

KEY RESULTS Urocortin 2 elicited time- and concentration-dependent positive inotropic and lusitropic effects (EC50: 19 nM) that were abolished by antisauvagine-30 (10 nM, n= 6), a specific antagonist of corticotrophin releasing factor (CRF) CRF2 receptors. Urocortin 2 (100 nM) increased the amplitude and decreased the time constant of decay of the underlying [Ca2+]i transients. Urocortin 2 also increased PLN phosphorylation at serine-16. H89 (2 µM) or KT5720 (1 µM), two inhibitors of protein kinase A (PKA), as well as KN93 (1 µM), an inhibitor of Ca2+/calmodulin-dependent protein kinase II (CaMKII), suppressed the urocortin 2 effects on shortening and [Ca2+]i transients. In addition, urocortin 2 also elicited arrhythmogenic events consisting of extra cell shortenings and extra [Ca2+]i increases in diastole. Urocortin 2-induced arrhythmogenic events were significantly reduced in cells pretreated with KT5720 or KN93.

CONCLUSIONS AND IMPLICATIONS Urocortin 2 enhanced contractility in mouse ventricular myocytes via activation of CRF2 receptors in a cAMP/PKA- and Ca2+/CaMKII-dependent manner. This enhancement was accompanied by Ca2+-dependent arrhythmogenic effects mediated by PKA and CaMKII.

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