title: Transgenic mice for real-time visualization of cGMP in intact adult cardiomyocytes.
abstract: BACKGROUND
  3',5'-Cyclic guanosine monophosphate (cGMP) is an important second messenger that regulates cardiac contractility and protects the heart from hypertrophy. However, because of the lack of real-time imaging techniques, specific subcellular mechanisms and spatiotemporal dynamics of cGMP in adult cardiomyocytes are not well understood.
  
  
  OBJECTIVE
  Our aim was to generate and characterize a novel cGMP sensor model to measure cGMP with nanomolar sensitivity in adult cardiomyocytes.
  
  
  RESULTS
  We generated transgenic mice with cardiomyocyte-specific expression of the highly sensitive cytosolic Förster resonance energy transfer-based cGMP biosensor red cGES-DE5 and performed the first Förster resonance energy transfer measurements of cGMP in intact adult mouse ventricular myocytes. We found very low (≈10 nmol/L) basal cytosolic cGMP levels, which can be markedly increased by natriuretic peptides (C-type natriuretic peptide >> atrial natriuretic peptide) and, to a much smaller extent, by the direct stimulation of soluble guanylyl cyclase. Constitutive activity of this cyclase contributes to basal cGMP production, which is balanced by the activity of clinically established phosphodiesterase (PDE) families. The PDE3 inhibitor, cilostamide, showed especially strong cGMP responses. In a mild model of cardiac hypertrophy after transverse aortic constriction, PDE3 effects were not affected, whereas the contribution of PDE5 was increased. In addition, after natriuretic peptide stimulation, PDE3 was also involved in cGMP/cAMP crosstalk.
  
  
  CONCLUSIONS
  The new sensor model allows visualization of real-time cGMP dynamics and pharmacology in intact adult cardiomyocytes. Förster resonance energy transfer imaging suggests the importance of well-established and potentially novel PDE-dependent mechanisms that regulate cGMP under physiological and pathophysiological conditions.
authors.affiliation: From Emmy Noether Group of the DFG, Department of Cardiology and Pneumology, European Heart Research Institute Göttingen, Georg August University Medical Center, University of Göttingen, Göttingen, Germany (K.R.G., J.U.S., R.K.P., J.H.S., S.E.L., V.O.N.); Institute of Physiology, University of Würzburg, Würzburg, Germany (M.K.); Department of Cardiovascular Sciences, National Heart and Lung Institute, Imperial College, London, United Kingdom (J.G.); and Institut de Recherche Experimentale et Clinique (IREC), Pole of Pharmacology and Therapeutics (FATH), University Catholique de Louvain, and Department of Medicine, Cliniques Universitaires Saint-Luc, Brussels, Belgium (J.-L.B.).
authors.forename: Konrad R
authors.initials: KR
authors.lastname: Götz
authors.forename: Julia U
authors.initials: JU
authors.lastname: Sprenger
authors.forename: Ruwan K
authors.initials: RK
authors.lastname: Perera
authors.forename: Julia H
authors.initials: JH
authors.lastname: Steinbrecher
authors.forename: Stephan E
authors.initials: SE
authors.lastname: Lehnart
authors.forename: Michaela
authors.initials: M
authors.lastname: Kuhn
authors.forename: Julia
authors.initials: J
authors.lastname: Gorelik
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Balligand
authors.forename: Viacheslav O
authors.initials: VO
authors.lastname: Nikolaev
mesh_terms: D000818:Animals; D015374:Biosensing Techniques / Q000379:methods; D000242:Cyclic AMP / Q000378:metabolism; D006152:Cyclic GMP / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D051379:Mice; D008822:Mice, Transgenic; D023421:Models, Animal; D032383:Myocytes, Cardiac / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D058987:Phosphodiesterase 3 Inhibitors / Q000494:pharmacology; D015363:Quinolones / Q000494:pharmacology
chemical_list: D058987:Phosphodiesterase 3 Inhibitors; D015363:Quinolones; C021294:cilostamide; D000242:Cyclic AMP; D006152:Cyclic GMP
keywords: biosensing techniques; fluorescence resonance energy transfer; hypertrophy; mice, transgenic; myocytes, cardiac; phosphodiesterase inhibitors