A junctional cAMP compartment regulates rapid Ca2+ signaling in atrial myocytes

Year: 2022

Authors: Brandenburg S., Pawlowitz J., Steckmeister V., Subramanian H., Uhlenkamp D., Scardigli M., Mushtaq M., Amlaz S.I., Kohl T., Wegener J.W., Arvanitis D.A., Sanoudou D., Sacconi L., Hasenfuss G., Voigt N., Nikolaev V.O., Lehnart S.E.

Autors Affiliation: Cellular Biophysics and Translational Cardiology Section, Department of Cardiology & Pneumology, Heart Research Center Gottingen, University Medical Center Gottingen, Gottingen, Germany; DZHK (German Centre for Cardiovascular Research), partner site Gottingen, Germany; Cluster of Excellence “Multiscale Bioimaging: from Molecular Machines to Networks of Excitable Cells” (MBExC), University of Gottingen, Germany; Institute of Pharmacology and Toxicology, Heart Research Center Gottingen, University Medical Center Gottingen, Gottingen, Germany; Institute of Experimental Cardiovascular Research, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; Department of Physics and Astronomy, University of Florence, Florence, Italy; European Laboratory for Non-Linear Spectroscopy and National Institute of Optics (INO-CNR), Sesto Fiorentino, Italy; Molecular Biology Division, Biomedical Research Foundation, Academy of Athens, Athens, Greece; Institute for Experimental Cardiovascular Medicine, University Heart Center Freiburg, Faculty of Medicine, University of Freiburg, Freiburg im Breisgau, Germany; DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/L

Abstract: Axial tubule junctions with the sarcoplasmic reticulum control the rapid intracellular Ca2+-induced Ca2+ release that initiates atrial contraction. In atrial myocytes we previously identified a constitutively increased ryanodine receptor (RyR2) phosphorylation at junctional Ca2+ release sites, whereas non-junctional RyR2 clusters were phosphorylated acutely following beta-adrenergic stimulation. Here, we hypothesized that the baseline synthesis of 3?,5?-cyclic adenosine monophosphate (cAMP) is constitutively augmented in the axial tubule junctional compartments of atrial myocytes. Confocal immunofluorescence imaging of atrial myocytes revealed that junctin, binding to RyR2 in the sarcoplasmic reticulum, was densely clustered at axial tubule junctions. Interestingly, a new transgenic junctin-targeted FRET cAMP biosensor was exclusively co-clustered in the junctional compart-ment, and hence allowed to monitor cAMP selectively in the vicinity of junctional RyR2 channels. To dissect local cAMP levels at axial tubule junctions versus subsurface Ca2+ release sites, we developed a confocal FRET imaging technique for living atrial myocytes. A constitutively high adenylyl cyclase activity sustained increased local cAMP levels at axial tubule junctions, whereas beta-adrenergic stimulation overcame this cAMP compartmentation resulting in additional phosphorylation of non-junctional RyR2 clusters. Adenylyl cyclase inhibition, however, abolished the junctional RyR2 phosphorylation and decreased L-type Ca2+ channel currents, while FRET imaging showed a rapid cAMP decrease. In conclusion, FRET biosensor imaging identified compartmentalized, consti-tutively augmented cAMP levels in junctional dyads, driving both the locally increased phosphorylation of RyR2 clusters and larger L-type Ca2+ current density in atrial myocytes. This cell-specific cAMP nanodomain is maintained by a constitutively increased adenylyl cyclase activity, contributing to the rapid junctional Ca2+- induced Ca2+ release, whereas beta-adrenergic stimulation overcomes the junctional cAMP compartmentation through cell-wide activation of non-junctional RyR2 clusters.


Volume: 165      Pages from: 141  to: 157

More Information: This work was supported by Deutsche Forschungsgemeinschaft to SB (SFB1002 project A09), SEL (SFB1002 projects A09 and S02; SFB1190 project P03), GH (SFB1002 project D01), NV (SFB1002 project A13, VO 1568/3-1, VO 1568/4-1, IRTG1816), VON (SFB1002 project A01); and under Germany´s Excellence Strategy -EXC 2067/1-390729940. VON was supported by the Gertraud und Heinz Rose-Foundation. NV received funding from the Else-Kr oner-Fresenius Foundation (EKFS 2016_A20). JP was supported by the German Cardiac Society (Otto-Hess-Scholarship). SB received financial support through a DZHK (German Centre for Cardiovascular Research) postdoc start-up grant (81X3300108), and the clinician scientist program “Translational Medicine” of the University Medical Center Gottingen. GH, NV, SEL, VON are principal investigators of the DZHK (German Centre for Cardiovascular Research).
KeyWords: Adenylyl cyclase; Atrial myocyte; cAMP; Ca2+ signaling; FRET biosensor; Ryanodine receptor
DOI: 10.1016/j.yjmcc.2022.01.003

Citations: 6
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