Quantification of [Ca2+](i) in perfused hearts. Critical evaluation of the 5F-BAPTA and nuclear magnetic resonance method as applied to the study of ischemia and reperfusion

E. Marban, M. Kitakaze, Y. Koretsune, D. T. Yue, V. P. Chacko, M. M. Pike

Research output: Contribution to journalArticle

180 Scopus citations

Abstract

Calcium has been implicated as a mediator of cell injury in ischemia and reperfusion, but direct measurements of Ca2+ are required to refine this idea. We used nuclear magnetic resonance spectroscopy and the Ca2+ indicator 5F-BAPTA to measure [Ca2+](i) in perfused ferret hearts. Several lines of evidence are presented to show that loading with the acetoxymethyl ester of 5F-BAPTA is not significantly complicated by accumulation of partially de-esterified metabolites, compartmentalization into mitochondria, or disproportionate uptake into endothelial cells. During 20 minutes of total global ischemia at 30°C, time-averaged [Ca2+](i) increased significantly, reaching peak values roughly three times control at 15-20 minutes. Reperfusion resulted in a persistent elevation of [Ca2+](i) during the first 5 minutes, but not afterward. Although the nonlinear response of 5F-BAPTA to [Ca2+] leads to underestimation of the true time-averaged [Ca2+](i), the measured alterations of intracellular Ca2+ homeostasis during ischemia are large compared with the likely errors in quantification. Phosphorus nuclear magnetic resonance spectroscopy of 5F-BAPTA-loaded hearts reveals changes during ischemia similar to those recorded previously in hearts not containing a Ca2+ indicator. Developed pressure recovers to only 50% of control values during reflow, indicating that the presence of 5F-BAPTA in the cytosol does not protect against stunning, at least when the extracellular calcium concentration has been raised to 8 mM. We conclude that 5F-BAPTA provides useful measurements that reveal that time-averaged [Ca2+](i) rises during ischemia and returns to control levels soon after reperfusion.

Original languageEnglish (US)
Pages (from-to)1255-1267
Number of pages13
JournalCirculation research
Volume66
Issue number5
DOIs
StatePublished - Jan 1 1990
Externally publishedYes

Keywords

  • cardiac function
  • excitation-contraction coupling
  • nuclear magnetic resonance spectroscopy
  • stunned myocardium

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine

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