Increased lysophosphatidylcholine content induced by thrombin receptor stimulation in adult rabbit cardiac ventricular myocytes.
OBJECTIVE
The aims were (1) to determine whether thrombin, which is increased in the presence of coronary thrombosis, can directly stimulate the production of lysophosphatidylcholine, which has arrhythmogenic properties, in ventricular myocytes; (2) whether the effect is dependent upon extracellular [Ca2+]; and (3) whether it is mediated directly through stimulation of the thrombin receptor.


METHODS
Lipids were extracted from isolated adult rabbit ventricular myocytes and lysophosphatidylcholine was isolated by HPLC and quantified using a recently developed radiometric assay employing 3H-acetic anhydride.


RESULTS
Thrombin (0.05 U.ml-1) stimulation of ventricular myocytes resulted in a nearly sixfold increase in lysophosphatidylcholine levels [0.26(SEM 0.03) to 1.61(0.42) nmol.mg-1 protein] within 1 min. The increase in myocytic lysophosphatidylcholine content was prevented by preincubation of thrombin with the proteolytic site inhibitors phenyly-prolyl-arginyl-chloromethyl ketone (PPACK) and dansylarginine N-(3-ethyl-1,5-pentanediyl)amide. The increase in lysophosphatidylcholine content in response to thrombin was not present at an extracellular calcium concentration ([Ca2+)]o) = 500 microM, but was marked at a physiological level of [Ca2+]o = 1.8 mM. Stimulation of myocytes with the thrombin receptor activating peptide SFLLRNPNDKYEPF (100 microM for 1 min) resulted in a similar increase in lysophosphatidylcholine content [1.61(0.27) nmol.mg-1 protein].


CONCLUSIONS
The marked increase in lysophosphatidylcholine content in cardiac myocytes in response to thrombin has important implications as an arrhythmogenic mechanism during early myocardial ischaemia.
Cardiovascular Division, Washington University School of Medicine, St Louis, Missouri 63110.
T H
TH
Park
J
J
McHowat
R A
RA
Wolf
P B
PB
Corr
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D001120:Arginine / Q000031:analogs & derivatives / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D003619:Dansyl Compounds / Q000494:pharmacology; D005260:Female; D008244:Lysophosphatidylcholines / Q000378:metabolism*; D008297:Male; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D010446:Peptide Fragments / Q000494:pharmacology; D011817:Rabbits; D011956:Receptors, Cell Surface; D018179:Receptors, Thrombin / Q000187:drug effects*; D013268:Stimulation, Chemical; D013917:Thrombin / Q000494:pharmacology*
D000590:Amino Acid Chloromethyl Ketones; D003619:Dansyl Compounds; D008244:Lysophosphatidylcholines; D010446:Peptide Fragments; D011956:Receptors, Cell Surface; D018179:Receptors, Thrombin; C072032:thrombin receptor peptide (42-55); C019162:dansylarginine N-(3-ethyl-1,5-pentanediyl)amide; D001120:Arginine; D013917:Thrombin; C040308:phenylalanyl-prolyl-arginine-chloromethyl ketone; D002118:Calcium