title: Mechanism of Ca(2+) overload in endothelial cells exposed to simulated ischemia.
abstract: OBJECTIVE
  Several studies have shown that myocardial ischemia leads to functional failure of endothelial cells (EC) whereby disturbance of Ca(2+) homeostasis may play an important role. The mechanisms leading to Ca(2+) disbalance in ischemic EC are not fully understood. The aim of this study was to test effects of different components of simulated ischemia (glucose deprivation, anoxia, low extracellular pH (pH(o)) and lactate) on Ca(2+) homeostasis in EC.
  
  
  METHODS
  Cytosolic Ca(2+) (Ca(i)), cytosolic pH (pH(i)) and ATP content were measured in cultured rat coronary EC.
  
  
  RESULTS
  In normoxic cells 60 min glucose deprivation at pH(o) 7.4 had no effect on pH(i). It only slightly increased Ca(i) and decreased ATP content. Reduction of pH(o) to 6.5 under these conditions led to marked cytosolic acidosis and Ca(i) overload, but had no effect on ATP content. Anoxia at pH(o) 6.5 had no additional effect on Ca(i) overload, but significantly reduced cellular ATP. Addition of 20 mmol/l lactate to anoxia at pH(o) 6.5 accelerated Ca(i) overload due to faster cytosolic acidification. Acidosis-induced Ca(i) overload was prevented by inhibition of Ca(2+) release channels of endoplasmic reticulum (ER) with 3 micromol/l ryanodine or by pre-emptying the ER with thapsigargin. Re-normalisation of pH(o) for 30 min led to recovery of pH(i), but not of Ca(i).
  
  
  CONCLUSIONS
  The ischemic factors leading to cytosolic acidosis (low pH(o) and lactate) cause Ca(i) overload in endothelial cells, while anoxia and glucose deprivation play only a minor role. The ER is the main source for this Ca(i) rise. Ca(i) overload is not readily reversible.
authors.affiliation: Physiologisches Institut, Justus-Liebig-Universität, Giessen, Germany. yury.ladilov@physiologie.med.uni-giessen.de
authors.forename: Y
authors.initials: Y
authors.lastname: Ladilov
authors.forename: C
authors.initials: C
authors.lastname: Schäfer
authors.forename: A
authors.initials: A
authors.lastname: Held
authors.forename: M
authors.initials: M
authors.lastname: Schäfer
authors.forename: T
authors.initials: T
authors.lastname: Noll
authors.forename: H M
authors.initials: HM
authors.lastname: Piper
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000704:Analysis of Variance; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D048429:Cell Size / Q000187:drug effects; D002478:Cells, Cultured; D003331:Coronary Vessels; D003600:Cytosol / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006146:Guanidines / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D019344:Lactic Acid / Q000494:pharmacology; D008297:Male; D008345:Manganese / Q000378:metabolism; D017202:Myocardial Ischemia / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D012433:Ryanodine / Q000494:pharmacology; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors; D013450:Sulfones / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology
chemical_list: D002121:Calcium Channel Blockers; D004791:Enzyme Inhibitors; D006146:Guanidines; D017923:Sodium-Hydrogen Exchangers; D013450:Sulfones; D012433:Ryanodine; D019344:Lactic Acid; D008345:Manganese; D019284:Thapsigargin; C093373:cariporide; D000255:Adenosine Triphosphate; D002118:Calcium