Endogenous heavy metal ions perturb fura-2 measurements of basal and hormone-evoked Ca2+ signals

Vladislav A. Snitsarev, Tracy J. McNulty, Colin W. Taylor

Research output: Contribution to journalArticle

31 Scopus citations

Abstract

Using the membrane-permeant chelator of heavy metal ions, N,N,N',N'- tetrakis(2-pyridylmethyl)ethylene diamine (TPEN), we demonstrate that in pancreatic acinar cells, hepatocytes, and a variety of mammalian cell lines, endogenous heavy metal ions bind to cytosolic fura-2 causing basal cytosolic free [Ca2+] ([Ca2+](i)) to be overestimated. TPEN had most effect in cells lightly loaded with fura-2, suggesting the presence of a limited pool of heavy metal ions (≤12 μM in pancreatic acinar cells) that does not rapidly exchange across the plasma membrane. In fura-2-loaded hepatocytes, vasopressin failed to evoke a detectable change in fluorescence, but after preincubation of cells with TPEN, it caused fluorescence changes characteristic of an increase in [Ca2+](i). We conclude that in many mammalian cells, a slowly exchanging mixture of cytosolic heavy metal ions binds to fura-2 both to quench its fluorescence and to mimic the effects of Ca2+ binding, thereby causing basal [Ca2+](i) to be overestimated. By chelating endogenous heavy metal ions, TPEN allows basal [Ca2+](i) to be accurately measured and, by preventing competition between heavy metal ions and Ca2+ for binding to fura-2, unmasks the full effect of agonists in increasing [Ca2+](i).

Original languageEnglish
Pages (from-to)1048-1056
Number of pages9
JournalBiophysical Journal
Volume71
Issue number2
DOIs
StatePublished - Aug 1996

    Fingerprint

Cite this