Ca(2+) -stimulated basal adenylyl cyclase activity localization in membrane lipid microdomains of cardiac sinoatrial nodal pacemaker cells.
Spontaneous, rhythmic subsarcolemmal local Ca(2+) releases driven by cAMP-mediated, protein kinase A (PKA)-dependent phosphorylation are crucial for normal pacemaker function of sinoatrial nodal cells (SANC). Because local Ca(2+) releases occur beneath the cell surface membrane, near to where adenylyl cyclases (ACs) reside, we hypothesized that the dual Ca(2+) and cAMP/PKA regulatory components of automaticity are coupled via Ca(2+) activation of AC activity within membrane microdomains. Here we show by quantitative reverse transcriptase PCR that SANC express Ca(2+)-activated AC isoforms 1 and 8, in addition to AC type 2, 5, and 6 transcripts. Immunolabeling of cell fractions, isolated by sucrose gradient ultracentrifugation, confirmed that ACs localize to membrane lipid microdomains. AC activity within these lipid microdomains is activated by Ca(2+) over the entire physiological Ca(2+) range. In intact SANC, the high basal AC activity produces a high level of cAMP that is further elevated by phosphodiesterase inhibition. cAMP and cAMP-mediated PKA-dependent activation of ion channels and Ca(2+) cycling proteins drive sarcoplasmic reticulum Ca(2+) releases, which, in turn, activate ACs. This feed forward "fail safe" system, kept in check by a high basal phosphodiesterase activity, is central to the generation of normal rhythmic, spontaneous action potentials by pacemaker cells.
Younes
Antoine
A
Laboratory of Cardiovascular Science, Gerontology Research Center, NIA, Intramural Research Program, NIH, Baltimore, MD 21224, USA.
Lyashkov
Alexey E
AE
Graham
David
D
Sheydina
Anna
A
Volkova
Maria V
MV
Mitsak
Megan
M
Vinogradova
Tatiana M
TM
Lukyanenko
Yevgeniya O
YO
Li
Yue
Y
Ruknudin
Abdul M
AM
Boheler
Kenneth R
KR
van Eyk
Jennifer
J
Lakatta
Edward G
EG
D000262:Adenylyl Cyclases / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology*; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004789:Enzyme Activation; D005786:Gene Expression Regulation; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D008563:Membrane Lipids / Q000378:metabolism*; D021962:Membrane Microdomains / Q000187:drug effects / Q000201:enzymology*; D012333:RNA, Messenger / Q000235:genetics; D011817:Rabbits; D012849:Sinoatrial Node / Q000166:cytology* / Q000201:enzymology*
D007527:Isoenzymes; D008563:Membrane Lipids; D012333:RNA, Messenger; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D000262:Adenylyl Cyclases; D002118:Calcium