Effect of protein kinase A on calcium sensitivity of force and its sarcomere length dependence in human cardiomyocytes.
OBJECTIVE
We investigated whether the Frank-Starling mechanism is absent or preserved in end-stage failing human myocardium and if phosphorylation of contractile proteins modulates its magnitude through the sarcomere length-dependence of calcium sensitivity of isometric force development.


METHODS
The effect of phosphorylation of troponin I and C-protein by the catalytic subunit of protein kinase A (3 microg/ml; 40 min at 20 degrees C) was studied in single Triton-skinned human cardiomyocytes isolated from donor and end-stage failing left ventricular myocardium at sarcomere lengths measured at rest of 1.8, 2.0 and 2.2 microm. Isometric force development was studied at various free-calcium concentrations before and after protein kinase A incubation at 15 degrees C (pH 7.1).


RESULTS
Maximal isometric tension at 2.2 microm amounted to 39.6+/-10.4 and 33.7+/-3.5 kN/m2 in donor and end-stage failing cardiomyocytes, respectively. The midpoints of the calcium sensitivity curves (pCa50) of donor and end-stage failing hearts differed markedly at all sarcomere lengths (mean delta pCa50=0.22). A reduction in sarcomere length from 2.2 to 1.8 microm caused reductions in maximum isometric force to 64% and 65% and in pCa50 by 0.10 and 0.08 pCa units in donor and failing cardiomyocytes, respectively. In donor tissue, the effect of protein kinase A treatment was rather small, while in end-stage failing myocardium it was much larger (delta pCa50=0.24) irrespective of sarcomere length.


CONCLUSIONS
The data obtained indicate that the Frank-Starling mechanism is preserved in end-stage failing myocardium and suggest that sarcomere length dependence of calcium sensitivity and the effects of phosphorylation of troponin I and C-protein are independent.
van der Velden
J
J
Laboratory for Physiology, Institute for Cardiovascular Research (ICaR-VU), Free University, Amsterdam, The Netherlands. j.van_der_velden.physiol@med.vu.nl
de Jong
J W
JW
Owen
V J
VJ
Burton
P B
PB
Stienen
G J
GJ
D002118:Calcium / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000494:pharmacology*; D006333:Heart Failure / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D007537:Isometric Contraction; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D010766:Phosphorylation; D012518:Sarcomeres / Q000187:drug effects*; D019210:Troponin I / Q000378:metabolism
D002352:Carrier Proteins; D019210:Troponin I; C020723:citrate-binding transport protein; D017868:Cyclic AMP-Dependent Protein Kinases; D002118:Calcium