Regulation of the L-type Ca2+ channel during cardiomyogenesis: switch from NO to adenylyl cyclase-mediated inhibition.
In adult mammalian cardiomyocytes, stimulation of muscarinic receptors counterbalances the beta-adrenoceptor-mediated increase in myocardial contractility and heart rate by decreasing the L-type Ca2+ current (ICa) (1, 2). This effect is mediated via inhibition of adenylyl cyclase and subsequent reduction of cAMP-dependent phosphorylation of voltage-dependent L-type Ca2+ channels (3). Little is known, however, about the nature and origin of this pivotal inhibitory pathway. Using embryonic stem cells as an in vitro model of cardiomyogenesis, we found that muscarinic agonists depress ICa by 58 +/-3% (n=34) in early stage cardiomyocytes lacking functional beta-adrenoceptors. The cholinergic inhibition is mediated by the nitric oxide (NO)/cGMP system since it was abolished by application of NOS inhibitors (L-NMA, L-NAME), an inhibitor of the soluble guanylyl cyclase (ODQ), and a selective phosphodiesterase type II antagonist (EHNA). The NO/cGMP-mediated ICa depression was dependent on a reduction of cAMP/protein kinase A (PKA) levels since application of the catalytic subunit of PKA or of the PKA inhibitor PK) prevented the carbachol effect. In late development stage cells, as reported for ventricular cardiomyocytes (2, 4), muscarinic agonists had no effect on basal ICa but antagonized beta-adrenoceptor-stimulated ICa by 43 +/-4% (n=16). This switch in signaling pathways during development is associated with distinct changes in expression of the two NO-producing isoenzymes, eNOS and iNOS, respectively. These findings indicate a fundamental role for NO as a signaling molecule during early embryonic development and demonstrate a switch in the signaling cascades governing ICa regulation.
Ji
G J
GJ
Institute of Neurophysiology, University of Cologne, Cologne, Germany.
Fleischmann
B K
BK
Bloch
W
W
Feelisch
M
M
Andressen
C
C
Addicks
K
K
Hescheler
J
J
D000225:Adenine / Q000031:analogs & derivatives / Q000494:pharmacology; D000067956:Adenylyl Cyclase Inhibitors; D000818:Animals; D015220:Calcium Channels / Q000502:physiology*; D004594:Electrophysiology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006321:Heart / Q000196:embryology* / Q000502:physiology*; D015640:Ion Channel Gating / Q000187:drug effects; D051379:Mice; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000502:physiology*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D013234:Stem Cells / Q000502:physiology; D019323:omega-N-Methylarginine / Q000494:pharmacology