Phosphorylation of phospholamban in the intact heart. A study on the physiological role of the Ca(2+)-calmodulin-dependent protein kinase system.
The aim of the present study was to further elucidate the physiological role of the calcium-calmodulin (Ca(2+)-Cm)-dependent protein kinase system on phospholamban phosphorylation in the intact functioning heart. The effect of increasing extracellular calcium concentration [Ca]o on phospholamban phosphorylation (PHPL) was studied under different experimental conditions: (a) regular twitches and ryanodine induced-tetani both in the presence and in the absence of 3 x 10(-8) M isoproterenol and (b) Post-stimulation potentiation (PSP), i.e. the potentiation of contractility that follows a period of rapid repetitive stimulation. In the regular twitch, the increase in [Ca]o enhanced contractility both, in the absence and in the presence of beta-stimulation without changing basal or isoproterenol stimulated cAMP levels respectively. This increase in contractility was accompanied by a significant enhancement of PHPL-from 90.6 +/- 16.4 to 216 +/- 35.2 pmols 32Pi/mg protein at 0.25 and 3.85 mM [Ca]o respectively-only when isoproterenol was present. The calmodulin antagonist W-7 significantly decreased the isoproterenol-induced phosphorylation of phospholamban at [Ca]o 1.35 mM. Similar results were obtained under tetanic conditions. When myocardial contractility was enhanced by PSP up to ten-times with respect to the regular twitch, no detectable effect in PHPL was observed. Indirect evidence obtained from skinned rat cardiac trabeculae suggested that the failure of the cAMP-independent mechanisms to phosphorylate phospholamban is not related to a deficient increase in intracellular calcium. The results support the notion that the increase in intracellular calcium induces an increase in PHPL only at high intracellular cAMP levels.
Napolitano
R
R
Centro de Investigaciones Cardiovasculares, Facultad de Ciencias Médicas, Universidad Nacional de La Plata, Argentina.
Vittone
L
L
Mundiña
C
C
Chiappe de Cingolani
G
G
Mattiazzi
A
A
D000818:Animals; D002118:Calcium / Q000502:physiology*; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002147:Calmodulin / Q000502:physiology*; D000242:Cyclic AMP / Q000378:metabolism; D004558:Electric Stimulation; D066298:In Vitro Techniques; D007545:Isoproterenol / Q000494:pharmacology; D009200:Myocardial Contraction / Q000187:drug effects; D009206:Myocardium / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011494:Protein Kinases / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D000099211:Phospholamban
D002118:Calcium; D002135:Calcium-Binding Proteins; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002147:Calmodulin; D000242:Cyclic AMP; D007545:Isoproterenol; D011494:Protein Kinases; D000099211:Phospholamban