Do rat cardiac myocytes release ATP on contraction?
ATP is released by numerous cell types in response to mechanical strain. It then acts as a paracrine or autocrine signaling molecule, inducing a variety of biological responses. In this work, we addressed the question whether mechanical force acting on the membranes of contracting cardiomyocytes during periodic longitudinal shortening can stimulate the release of ATP. Electrically stimulated isolated adult rat cardiomyocytes as well as spontaneously contracting mouse cardiomyocytes derived from embryonic stem (ES) cells were assayed for ATP release with the use of luciferase and a sensitive charge-coupled device camera. Sensitivity of soluble luciferase in the supernatant of cardiomyocytes was 100 nM ATP, which is approximately 10-fold below the EC(50) values for most purinergic receptors expressed in the heart (1.5-20 microM). Light intensities were not different between resting or contracting adult rat cardiomyocytes. Similar results were obtained with ES-cell-derived contracting mouse cardiomyocytes. ATP release was measurable only from obviously damaged or permeabilized cells. To increase selectivity and sensitivity of ATP detection we have targeted a recombinant luciferase to the sarcolemmal membrane using a wheat germ agglutinin-IgG linker. Contraction of labeled adult rat cardiomyocytes was not associated with measurable bioluminescence. However, when human umbilical vein endothelial cells were targeted with membrane-bound luciferase, shear stress-induced ATP release could be clearly detected, demonstrating the sensitivity of the detection method. In the present study, we did not detect ATP release from contracting cardiomyocytes on the single cell level, despite adequate sensitivity of the detection system. Thus deformation of the contracting cardiomyocyte is not a key stimulus for the release of cellular ATP.
Gödecke
Stefanie
S
Institut für Herz- und Kreislaufphysiologie, Heinrich-Heine-Universität, Düsseldorf, Germany. stefanie.goedecke@uni-duesseldorf.de
Stumpe
Thomas
T
Schiller
Hilmar
H
Schnittler
Hans-J
HJ
Schrader
Jürgen
J
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000367:Age Factors; D000818:Animals; D002478:Cells, Cultured; D004558:Electric Stimulation; D004730:Endothelium, Vascular / Q000166:cytology; D049454:Firefly Luciferin; D006801:Humans; D049408:Luminescent Agents; D051379:Mice; D008853:Microscopy / Q000295:instrumentation / Q000379:methods; D009200:Myocardial Contraction / Q000502:physiology*; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D013234:Stem Cells / Q000166:cytology; D014470:Umbilical Cord / Q000166:cytology
D049408:Luminescent Agents; D049454:Firefly Luciferin; D000255:Adenosine Triphosphate