Xm. Wang et al., KINETICS OF THE SARCOLEMMAL LACTATE CARRIER IN SINGLE HEART-CELLS USING BCECF TO MEASURE PH(I), American journal of physiology. Heart and circulatory physiology, 36(5), 1994, pp. 1759-1769
The pH-sensitive fluorescent indicator 2',7'-bis(carboxyethyl)-5(6)car
boxyfluorescein (BCECF) was used to measure lactate transport in singl
e cardiac myocytes. Addition of lactate externally caused a rapid fall
of intracellular pH (pH(i)), which was largely inhibited by 5 mM alph
a-cyano-4-hydroxycinnamate (CHC), a specific inhibitor of the lactate
carrier. Stilbene disulfonates such as 4,4'-dibenzamidostilbene-2,2'-d
isulfonate (DBDS) only partially inhibited the response, with inhibiti
on being greater in guinea pig than rat myocytes. The data are consist
ent with two isoforms of the lactate carrier, one sensitive and one in
sensitive 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 c
onstant (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-lactat
e 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 maxim
al rate values for efflux were both threefold higher than for influx a
nd were related to each other and the transmembrane pH gradient as pre
dicted by the Haldane relationship. It is suggested that under hypoxic
conditions, the carrier may be the rate-limiting factor for lactate e
xtrusion.