Effect of thrombin on calcium homeostasis in chick embryonic heart cells. Receptor-operated calcium entry with inositol trisphosphate and a pertussis toxin-sensitive G protein as second messengers.
Thrombin increases intracellular calcium ([Ca++]i) in several cell types and causes a positive inotropic effect in the heart. We examined the mechanism of the thrombin-induced [Ca++]i increase in chick embryonic heart cells loaded with the fluorescent calcium indicator, indo-1. Thrombin (1 U/ml) increased both systolic and diastolic [Ca++]i from 617 +/- 62 and 324 +/- 46 to 1041 +/- 93 and 587 +/- 38 nM, respectively. An initial rapid [Ca++]i increase was followed by a more sustained increase. There were associated increases in contraction strength, beat frequency, and action potential duration. The [Ca++]i increase was not blocked by tetrodotoxin or verapamil, but was blocked by pretreatment with pertussis toxin (100 ng/ml). The thrombin-induced [Ca++]i increase was partly due to intracellular calcium release, since it persisted after removal of external calcium. The [Ca++]i increase in zero calcium was more transitory than in normal calcium and was potentiated by 10 mM Li+. Thrombin also induced influx of calcium across the surface membrane, which could be monitored using Mn++ ions, which quench indo-1 fluorescence when they enter the cell. Thrombin-induced Mn++ entry was insensitive to verapamil, but was blocked by 2 mM Ni++. Thrombin increased inositol trisphosphates by 180% at 90 s and this effect was also blocked by pretreatment with pertussis toxin.


CONCLUSIONS
thrombin promotes calcium entry and release in embryonic heart cells even when action potentials are inhibited. Both modes of [Ca++]i increase may be coupled to the receptor by pertussis toxin-sensitive G proteins.
Chien
W W
WW
Division of Cardiology, Falk Cardiovascular Research Center, Stanford University School of Medicine, California 94305.
Mohabir
R
R
Clusin
W T
WT
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism*; D002642:Chick Embryo; D005456:Fluorescent Dyes; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006352:Heart Ventricles / Q000187:drug effects / Q000378:metabolism; D006706:Homeostasis; D007211:Indoles; D007294:Inositol / Q000378:metabolism; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism*; D007700:Kinetics; D009206:Myocardium / Q000378:metabolism*; D037342:Pertussis Toxin; D015290:Second Messenger Systems / Q000187:drug effects*; D013917:Thrombin / Q000494:pharmacology*; D016276:Ventricular Function; D014700:Verapamil / Q000494:pharmacology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology*
D015220:Calcium Channels; D005456:Fluorescent Dyes; D007211:Indoles; D010566:Virulence Factors, Bordetella; D007294:Inositol; D015544:Inositol 1,4,5-Trisphosphate; D014700:Verapamil; D037342:Pertussis Toxin; D013917:Thrombin; D019204:GTP-Binding Proteins; C048960:indo-1; D002118:Calcium