The cytosolic phospholipase A2 pathway, a safeguard of beta2-adrenergic cardiac effects in rat.
We have recently demonstrated that in human heart, beta2-adrenergic receptors (beta2-ARs) are biochemically coupled not only to the classical adenylyl cyclase (AC) pathway but also to the cytosolic phospholipase A2 (cPLA2) pathway (Pavoine, C., Behforouz, N., Gauthier, C., Le Gouvello, S., Roudot-Thoraval, F., Martin, C. R., Pawlak, A., Feral, C., Defer, N., Houel, R., Magne, S., Amadou, A., Loisance, D., Duvaldestin, P., and Pecker, F. (2003) Mol. Pharmacol. 64, 1117-1125). In this study, using Fura-2-loaded cardiomyocytes isolated from adult rats, we showed that stimulation of beta2-ARs triggered an increase in the amplitude of electrically stimulated [Ca2+]i transients and contractions. This effect was abolished with the PKA inhibitor, H89, but greatly enhanced upon addition of the selective cPLA2 inhibitor, AACOCF3. The beta2-AR/cPLA2 inhibitory pathway involved G(i) and MSK1. Potentiation of beta2-AR/AC/PKA-induced Ca2+ responses by AACOCF3 did not rely on the enhancement of AC activity but was associated with eNOS phosphorylation (Ser1177) and L-NAME-sensitive NO production. This was correlated with PKA-dependent phosphorylation of PLB (Ser16). The constraint exerted by the beta2-AR/cPLA2 pathway on the beta2-AR/AC/PKA-induced Ca2+ responses required integrity of caveolar structures and was impaired by Filipin III treatment. Immunoblot analyses demonstrated zinterol-induced translocation of cPLA and its cosedimentation with MSK1, eNOS, PLB, and sarcoplasmic reticulum Ca2+ pump (SERCA) 2a in a low density caveolin-3-enriched membrane fraction. This inferred the gathering of beta2-AR signaling effectors around caveolae/sarcoplasmic reticulum (SR) functional platforms. Taken together, these data highlight cPLA as a cardiac beta2-AR signaling pathway that limits beta2-AR/AC/PKA-induced Ca2+ responses in adult rat cardiomyocytes through the impairment of eNOS activation and PLB phosphorylation.
Ait-Mamar
Bouziane
B
Inserm, U581, University of Paris, XII-Val de Marne, Créteil F-94010, France.
Cailleret
Michel
M
Rucker-Martin
Catherine
C
Bouabdallah
Anissa
A
Candiani
Gabriele
G
Adamy
Christophe
C
Duvaldestin
Philippe
P
Pecker
Francoise
F
Defer
Nicole
N
Pavoine
Catherine
C
D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D051242:Caveolin 1; D051244:Caveolin 3; D022461:Caveolins / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D003600:Cytosol / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004983:Ethanolamines / Q000494:pharmacology; D015151:Immunoblotting; D007150:Immunohistochemistry; D007211:Indoles / Q000494:pharmacology; D007546:Isoquinolines / Q000494:pharmacology; D008245:Lysophospholipase / Q000378:metabolism; D018613:Microscopy, Confocal; D009206:Myocardium / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000378:metabolism*; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D037342:Pertussis Toxin / Q000494:pharmacology; D010741:Phospholipases A / Q000378:metabolism / Q000502:physiology*; D054467:Phospholipases A2; D010766:Phosphorylation; D051381:Rats; D018343:Receptors, Adrenergic, beta-2 / Q000378:metabolism*; D012519:Sarcoplasmic Reticulum / Q000378:metabolism; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D012694:Serine / Q000737:chemistry; D013449:Sulfonamides / Q000494:pharmacology; D013997:Time Factors
C495537:Cav1 protein, rat; C495549:Cav3 protein, rat; D051242:Caveolin 1; D051244:Caveolin 3; D022461:Caveolins; D004791:Enzyme Inhibitors; D004983:Ethanolamines; D007211:Indoles; D007546:Isoquinolines; D018343:Receptors, Adrenergic, beta-2; D013449:Sulfonamides; D012694:Serine; C002904:zinterol; D037342:Pertussis Toxin; D017868:Cyclic AMP-Dependent Protein Kinases; D010741:Phospholipases A; D054467:Phospholipases A2; D008245:Lysophospholipase; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000262:Adenylyl Cyclases; D000252:Calcium-Transporting ATPases; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide; D002118:Calcium; D019331:NG-Nitroarginine Methyl Ester; C064758:Ro 31-8220