Time course of action of antagonists of mitochondrial Ca uptake in intact ventricular myocytes

Zhuan Zhou, Donald M Bers

Research output: Contribution to journalArticle

12 Scopus citations

Abstract

Mitochondrial Ca uptake is important for ATP production and intracellular Ca buffering. Several agents are often used to interfere with mitochondrial Ca transport, but the use of these agents in intact cells is complicated by lack of knowledge about how rapidly these agents gain access to intracellular mitochondria. We have developed a novel method to assess the time course with which extracellularly applied inhibitors gain access to mitochondria in intact ventricular myocytes preloaded with the fluorescent Ca indicator indo-1-AM. Cell contraction (assessed as the normalized change in cell length ΔLn) was employed as an index of the cytosolic [Ca] ([Ca]c), and was compared with the indo-1 ratio Rn, which reflects both mitochondrial [Ca] ([Ca]m) and [Ca]c. Upon abrupt plasma membrane damage in control cells, the delay (tk) between the rise in Ln and Rn was <10 s (reflecting the time lag between the change in [Ca]c and that in [Ca]m). Exposure of cells to 50 μM ruthenium red (RR) increased tk as a monotonic function of preincubation time. In contrast, 10 μM Ru360, a selective and more potent Ca uniporter blocker (Ki ∼0.2 nM) reached a comparable maximal tk after only 10 min, making it practical to use in intact cells. Carbonylcyanide p-(trifluoromethoxy)phenylhydrazone (FCCP) and carbonylcyanide m-chlorophenylhydrazone (CCCP) produced smaller maximal effects on tk, but did so almost immediately. These results are the first quantitative data on the time course of blockade of mitochondrial Ca uptake by the four most widely used mitochondrial Ca uptake antagonists in single ventricular myocytes.

Original languageEnglish (US)
Pages (from-to)132-138
Number of pages7
JournalPflugers Archiv European Journal of Physiology
Volume445
Issue number1
DOIs
StatePublished - Oct 1 2002
Externally publishedYes

Keywords

  • Biosensor
  • Ca uptake
  • Cardiac cells
  • CCCP
  • FCCP
  • Indo-1
  • Ru360
  • Ruthenium red

ASJC Scopus subject areas

  • Physiology

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