Calcium transport and monovalent cation and proton fluxes in sarcoplasmic
reticulum vesicles.
J. Biol. Chem. 256 (2): 636--643 (January 1981)
Abstract
ATP-dependent Ca$^2+$ uptake by rabbit skeletal muscle sarcoplasmic
reticulum vesicles has been studied in the presence and absence of
artificially generated pH gradients and membrane potentials. H$^+$
and K$^+$ diffusion potentials were generated via the H$^+$
and K,Na channels of sarcoplasmic reticulum by transfer of vesicles
from low to high pH, or from high to low K$^+$. Membrane potentials
were measured using the voltage-sensitive fluorescent dye 3,3'-dipentyl-2,2'-oxacarbocyanine.
The initial rate of Ca$^2+$ uptake was found to be increased
in the presence of a pH gradient and membrane potential (negative
inside). In turn, the rates of decay of K$^+$- or H$^+$-induced
membrane potentials were accelerated during Ca$^2+$ transport,
suggesting that active Ca$^2+$ uptake stimulated the release
of K$^+$ and H$^+$ from the vesicles. The ratio of K$^+$
(or H$^+$) release to Ca$^2+$ transport was near two. Release
of K$^+$ did not appear to be directly catalyzed by the Ca$^2+$-ATPase.
Evidence against a directly coupled ATP-mediated 2 K$^+$-Ca$^2+$
or K$^+$-Ca$^2+$ exchange reaction was that (i) similar results
were obtained when K$^+$ was substituted by Na$^+$ or by
organic cations which could rapidly permeate through the channel
of K$^+$,Na$^+$-permeable vesicles and (ii) Ca$^2+$ transport
did not result in an equivalent release of 86Rb+ or 22Na$^+$
from K$^+$,Na$^+$-impermeable vesicles. These studies are
in support of an electrogenic Ca$^2+$ transport system in sarcoplasmic
reticulum. The results further suggest that during Ca$^2+$ transport
development of a membrane potential (positive inside) is likely nullified
by the countermovement of the permeant cations K$^+$, Na$^+$,
and H$^+$.
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