%0 Journal Article %1 Hayn2010 %A von Hayn, K. %A Werthmann, R. C. %A Nikolaev, V. O. %A Hommers, L. G. %A Lohse, M. J. %A Bunemann, M. %D 2010 %J Am J Physiol Cell Physiol %K & *Calcium 1/antagonists AMP/*metabolism Adenylate Adrenergic Animals Aorta/enzymology Biosensing Cultured Cyclase/antagonists Cyclic Dose-Response Drug Energy Factors Fluorescence GTP-Binding Gq-G11/*metabolism Humans Inhibitors/pharmacology Interference Isoenzymes/antagonists Isoproterenol/pharmacology Mice Muscle, Muscle/drug Myocytes, Nucleotide Phosphodiesterase Phosphodiesterases, Protein Purinergic/metabolism RNA Relationship, Resonance Signaling/drug Smooth Smooth, Subunits, Techniques Time Transfection Transfer Triphosphate/metabolism Type Uridine Vascular/drug Vasoconstriction Vasodilation Xanthines/pharmacology alpha beta-Agonists/pharmacology effects effects/*enzymology inhibitors/genetics/*metabolism inhibitors/metabolism Receptor Cell %N 2 %P C324-32 %T Gq-mediated Ca2+ signals inhibit adenylyl cyclases 5/6 in vascular smooth muscle cells %U http://www.ncbi.nlm.nih.gov/pubmed/19889965 %V 298 %X cAMP and Ca(2+) are antagonistic intracellular messengers for the regulation of vascular smooth muscle tone; rising levels of Ca(2+) lead to vasoconstriction, whereas an increase of cAMP induces vasodilatation. Here we investigated whether Ca(2+) interferes with cAMP signaling by regulation of phophodiesterases (PDEs) or adenylyl cyclases (ACs). We studied regulation of cAMP concentrations by Ca(2+) signals evoked by endogenous purinergic receptors in vascular smooth muscle cells (VSMCs). The fluorescence resonance energy transfer (FRET)-based cAMP sensor Epac1-camps allowed the measurement of cAMP levels in single-living VSMCs with subsecond temporal resolution. Moreover, in vitro calibration of Epac1-camps enabled us to estimate the absolute cytosolic cAMP concentrations. Stimulation of purinergic receptors decreased cAMP levels in the presence of the beta-adrenergic agonist isoproterenol. Simultaneous imaging of cAMP with Epac1-camps and of Ca(2+) with Fura 2 revealed a rise of intracellular Ca(2+) in response to purinergic stimulation followed by a decline of cAMP. Chelation of intracellular Ca(2+) and overexpression of Ca(2+)-independent AC4 antagonized this decline of cAMP, whereas pharmacological inhibition of Ca(2+)-activated PDE1 had no effect. AC assays with VSMC membranes revealed a significant attenuation of isoproterenol-stimulated cAMP production by the presence of 2 muM Ca(2+). Furthermore, small interfering RNA (siRNA) knockdown of AC5 and AC6 (the two ACs known to be inhibited by Ca(2+)), significantly reduced the decrease of cAMP upon purinergic stimulation of isoproterenol-prestimulated VSMCs. Taken together, these results implicate a Ca(2+)-mediated inhibition of AC5 and 6 as an important mechanism of purinergic receptor-induced decline of cAMP and show a direct cross talk of these signaling pathways in VSMCs.

@article{Hayn2010, abstract = {cAMP and Ca(2+) are antagonistic intracellular messengers for the regulation of vascular smooth muscle tone; rising levels of Ca(2+) lead to vasoconstriction, whereas an increase of cAMP induces vasodilatation. Here we investigated whether Ca(2+) interferes with cAMP signaling by regulation of phophodiesterases (PDEs) or adenylyl cyclases (ACs). We studied regulation of cAMP concentrations by Ca(2+) signals evoked by endogenous purinergic receptors in vascular smooth muscle cells (VSMCs). The fluorescence resonance energy transfer (FRET)-based cAMP sensor Epac1-camps allowed the measurement of cAMP levels in single-living VSMCs with subsecond temporal resolution. Moreover, in vitro calibration of Epac1-camps enabled us to estimate the absolute cytosolic cAMP concentrations. Stimulation of purinergic receptors decreased cAMP levels in the presence of the beta-adrenergic agonist isoproterenol. Simultaneous imaging of cAMP with Epac1-camps and of Ca(2+) with Fura 2 revealed a rise of intracellular Ca(2+) in response to purinergic stimulation followed by a decline of cAMP. Chelation of intracellular Ca(2+) and overexpression of Ca(2+)-independent AC4 antagonized this decline of cAMP, whereas pharmacological inhibition of Ca(2+)-activated PDE1 had no effect. AC assays with VSMC membranes revealed a significant attenuation of isoproterenol-stimulated cAMP production by the presence of 2 muM Ca(2+). Furthermore, small interfering RNA (siRNA) knockdown of AC5 and AC6 (the two ACs known to be inhibited by Ca(2+)), significantly reduced the decrease of cAMP upon purinergic stimulation of isoproterenol-prestimulated VSMCs. Taken together, these results implicate a Ca(2+)-mediated inhibition of AC5 and 6 as an important mechanism of purinergic receptor-induced decline of cAMP and show a direct cross talk of these signaling pathways in VSMCs.}, added-at = {2010-12-14T18:12:02.000+0100}, author = {von Hayn, K. and Werthmann, R. C. and Nikolaev, V. O. and Hommers, L. G. and Lohse, M. J. and Bunemann, M.}, biburl = {https://www.bibsonomy.org/bibtex/213f6a36466b98c5c55fe479047213a0f/pharmawuerz}, endnotereftype = {Journal Article}, groups = {private}, interhash = {f026b4ac62780ff20c37dfefb08d5c90}, intrahash = {13f6a36466b98c5c55fe479047213a0f}, issn = {1522-1563 (Electronic) 0363-6143 (Linking)}, journal = {Am J Physiol Cell Physiol}, keywords = {& *Calcium 1/antagonists AMP/*metabolism Adenylate Adrenergic Animals Aorta/enzymology Biosensing Cultured Cyclase/antagonists Cyclic Dose-Response Drug Energy Factors Fluorescence GTP-Binding Gq-G11/*metabolism Humans Inhibitors/pharmacology Interference Isoenzymes/antagonists Isoproterenol/pharmacology Mice Muscle, Muscle/drug Myocytes, Nucleotide Phosphodiesterase Phosphodiesterases, Protein Purinergic/metabolism RNA Relationship, Resonance Signaling/drug Smooth Smooth, Subunits, Techniques Time Transfection Transfer Triphosphate/metabolism Type Uridine Vascular/drug Vasoconstriction Vasodilation Xanthines/pharmacology alpha beta-Agonists/pharmacology effects effects/*enzymology inhibitors/genetics/*metabolism inhibitors/metabolism Receptor Cell}, month = Feb, note = {von Hayn, Kathrin Werthmann, Ruth C Nikolaev, Viacheslav O Hommers, Leif G Lohse, Martin J Bunemann, Moritz Research Support, Non-U.S. Gov't United States American journal of physiology. Cell physiology Am J Physiol Cell Physiol. 2010 Feb;298(2):C324-32. Epub 2009 Nov 4.}, number = 2, pages = {C324-32}, shorttitle = {Gq-mediated Ca2+ signals inhibit adenylyl cyclases 5/6 in vascular smooth muscle cells}, timestamp = {2010-12-14T18:20:49.000+0100}, title = {Gq-mediated Ca2+ signals inhibit adenylyl cyclases 5/6 in vascular smooth muscle cells}, url = {http://www.ncbi.nlm.nih.gov/pubmed/19889965}, volume = 298, year = 2010 }