%0 Journal Article %1 Nikolaev2006 %A Nikolaev, V. O. %A Bunemann, M. %A Schmitteckert, E. %A Lohse, M. J. %A Engelhardt, S. %D 2006 %J Circ Res %K AMP/genetics/*metabolism Adrenergic Animals Biosensing Cardiac/cytology/*metabolism Channels/biosynthesis/genetics/metabolism Cyclic Diester Energy Fluorescence Humans Hydrolases/metabolism Ion Isoenzymes/metabolism Mice Models, Molecular Myocytes, Phosphoric Protein Resonance Signal Structure, Techniques/methods Tertiary Transduction Transfer/methods Transgenic beta-1/*metabolism beta-2/*metabolism beta-Agonists/pharmacology Receptor %N 10 %P 1084-91 %T Cyclic AMP imaging in adult cardiac myocytes reveals far-reaching beta1-adrenergic but locally confined beta2-adrenergic receptor-mediated signaling %U http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17038640 %V 99 %X Beta(1)- and beta(2)-adrenergic receptors (betaARs) are known to differentially regulate cardiomyocyte contraction and growth. We tested the hypothesis that these differences are attributable to spatial compartmentation of the second messenger cAMP. Using a fluorescent resonance energy transfer (FRET)-based approach, we directly monitored the spatial and temporal distribution of cAMP in adult cardiomyocytes. We developed a new cAMP-FRET sensor (termed HCN2-camps) based on a single cAMP binding domain of the hyperpolarization activated cyclic nucleotide-gated potassium channel 2 (HCN2). Its cytosolic distribution, high dynamic range, and sensitivity make HCN2-camps particularly well suited to monitor subcellular localization of cardiomyocyte cAMP. We generated HCN2-camps transgenic mice and performed single-cell FRET imaging on freshly isolated cardiomyocytes. Whole-cell superfusion with isoproterenol showed a moderate elevation of cAMP. Application of various phosphodiesterase (PDE) inhibitors revealed stringent control of cAMP through PDE4>PDE2>PDE3. The beta(1)AR-mediated cAMP signals were entirely dependent on PDE4 activity, whereas beta(2)AR-mediated cAMP was under control of multiple PDE isoforms. beta(1)AR subtype-specific stimulation yielded approximately 2-fold greater cAMP responses compared with selective beta(2)-subtype stimulation, even on treatment with the nonselective PDE inhibitor 3-isobutyl-1-methylxanthine (IBMX) (DeltaFRET, 17.3+/-1.3% beta(1)AR versus 8.8+/-0.4% beta(2)AR). Treatment with pertussis toxin to inactivate G(i) did not affect cAMP production. Localized beta(1)AR stimulation generated a cAMP gradient propagating throughout the cell, whereas local beta(2)AR stimulation did not elicit marked cAMP diffusion. Our data reveal that in adult cardiac myocytes, beta(1)ARs induce far-reaching cAMP signals, whereas beta(2)AR-induced cAMP remains locally confined.

@article{Nikolaev2006, abstract = {Beta(1)- and beta(2)-adrenergic receptors (betaARs) are known to differentially regulate cardiomyocyte contraction and growth. We tested the hypothesis that these differences are attributable to spatial compartmentation of the second messenger cAMP. Using a fluorescent resonance energy transfer (FRET)-based approach, we directly monitored the spatial and temporal distribution of cAMP in adult cardiomyocytes. We developed a new cAMP-FRET sensor (termed HCN2-camps) based on a single cAMP binding domain of the hyperpolarization activated cyclic nucleotide-gated potassium channel 2 (HCN2). Its cytosolic distribution, high dynamic range, and sensitivity make HCN2-camps particularly well suited to monitor subcellular localization of cardiomyocyte cAMP. We generated HCN2-camps transgenic mice and performed single-cell FRET imaging on freshly isolated cardiomyocytes. Whole-cell superfusion with isoproterenol showed a moderate elevation of cAMP. Application of various phosphodiesterase (PDE) inhibitors revealed stringent control of cAMP through PDE4>PDE2>PDE3. The beta(1)AR-mediated cAMP signals were entirely dependent on PDE4 activity, whereas beta(2)AR-mediated cAMP was under control of multiple PDE isoforms. beta(1)AR subtype-specific stimulation yielded approximately 2-fold greater cAMP responses compared with selective beta(2)-subtype stimulation, even on treatment with the nonselective PDE inhibitor 3-isobutyl-1-methylxanthine (IBMX) (DeltaFRET, 17.3+/-1.3% [beta(1)AR] versus 8.8+/-0.4% [beta(2)AR]). Treatment with pertussis toxin to inactivate G(i) did not affect cAMP production. Localized beta(1)AR stimulation generated a cAMP gradient propagating throughout the cell, whereas local beta(2)AR stimulation did not elicit marked cAMP diffusion. Our data reveal that in adult cardiac myocytes, beta(1)ARs induce far-reaching cAMP signals, whereas beta(2)AR-induced cAMP remains locally confined.}, added-at = {2010-12-14T18:12:02.000+0100}, author = {Nikolaev, V. O. and Bunemann, M. and Schmitteckert, E. and Lohse, M. J. and Engelhardt, S.}, biburl = {https://www.bibsonomy.org/bibtex/2f7f26dcb586619ba8757bdaafc2afc4c/pharmawuerz}, endnotereftype = {Journal Article}, interhash = {2abfdc37014776a34cc57d1da1b63eb0}, intrahash = {f7f26dcb586619ba8757bdaafc2afc4c}, issn = {1524-4571 (Electronic) 1524-4571 (Linking)}, journal = {Circ Res}, keywords = {AMP/genetics/*metabolism Adrenergic Animals Biosensing Cardiac/cytology/*metabolism Channels/biosynthesis/genetics/metabolism Cyclic Diester Energy Fluorescence Humans Hydrolases/metabolism Ion Isoenzymes/metabolism Mice Models, Molecular Myocytes, Phosphoric Protein Resonance Signal Structure, Techniques/methods Tertiary Transduction Transfer/methods Transgenic beta-1/*metabolism beta-2/*metabolism beta-Agonists/pharmacology Receptor}, month = {Nov 10}, note = {Nikolaev, Viacheslav O Bunemann, Moritz Schmitteckert, Eva Lohse, Martin J Engelhardt, Stefan Research Support, Non-U.S. Gov't United States Circulation research Circ Res. 2006 Nov 10;99(10):1084-91. Epub 2006 Oct 12.}, number = 10, pages = {1084-91}, shorttitle = {Cyclic AMP imaging in adult cardiac myocytes reveals far-reaching beta1-adrenergic but locally confined beta2-adrenergic receptor-mediated signaling}, timestamp = {2010-12-14T18:22:41.000+0100}, title = {Cyclic AMP imaging in adult cardiac myocytes reveals far-reaching beta1-adrenergic but locally confined beta2-adrenergic receptor-mediated signaling}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17038640}, volume = 99, year = 2006 }