Kinetics of the sarcolemmal lactate carrier in single heart cells using BCECF to measure pH(i)

Xuemin Wang, A. J. Levi, A. P. Halestrap

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Abstract

The pH-sensitive fluorescent indicator 2',7'-bis(carboxyethyl)-5(6)- carboxyfluorescein (BCECF) was used to measure lactate transport in single cardiac myocytes. Addition of lactate externally caused a rapid fall of intracellular pH (pH(i)), which was largely inhibited by 5 mM α-cyano-4- hydroxycinnamate (CHC), a specific inhibitor of the lactate carrier. Stilbene disulfonates such as 4,4'-dibenzamidostilbene-2,2'-disulfonate (DBDS) only partially inhibited the response, with inhibition being greater in guinea pig than rat myocytes. The data are consistent with two isoforms of the lactate carrier, one sensitive and one insensitive to DBDS, coexisting within a single myocyte and both having a stoichiometry of 1 lactate:1 proton. The initial rate of pH(i) fall was used to determine carrier kinetics. Rat myocytes had a Michaelis constant (K(m)) for external L-lactate of 2.74 mM and a K(m) for external pyruvate of 0.2 mM. Guinea pig cells had a K(m) for external L-lactate of 2.2 mM. Kinetics of lactate efflux were also evaluated using the rate of pH(i) recovery on removing external lactate. The K(m) and maximal rate values for efflux were both threefold higher than for influx and were related to each other and the transmembrane pH gradient as predicted by the Haldane relationship. It is suggested that under hypoxic conditions, the carrier may be the rate-limiting factor for lactate extrusion.

Original languageEnglish
JournalAmerican Journal of Physiology - Heart and Circulatory Physiology
Volume267
Issue number5 36-5
Publication statusPublished - 1994
Externally publishedYes

Keywords

  • cardiac myocyte
  • carrier stoichiometry
  • glycolysis
  • hypoxia
  • ischemia
  • lactic acid
  • lactic acid transport
  • stilbene disulfonate

ASJC Scopus subject areas

  • Physiology
  • Agricultural and Biological Sciences(all)

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