AJP - Cell Physiology, Vol 256, Issue 6 C1138-C1144, Copyright © 1989 by American Physiological Society
Effects of intracellular free Ca and rate of Ca influx on the Ca pump
Y. C. Yang and D. R. Yingst
Department of Physiology, Wayne State University, School of Medicine, Detroit, Michigan 48201.
The activity of the Ca pump of human red blood cells was studied in
resealed ghosts as a function of intracellular free Ca (fCai). Resealed
ghosts were made by the agarose column method to contain, in addition to
other constituents, less than 0.1 microM fCai, 100 microM arsenazo III, and
either 1 mM ATP plus an ATP regenerating system (active ghosts) or no added
ATP and no regenerating system (passive ghosts). The rate of Ca influx into
these ghosts was manipulated by suspending them in solutions containing
various combinations of free Ca (1-30 microM) and the Ca ionophore A23187
(0.1-0.7 microM). Entering Ca increased the fCai and stimulated the pump in
active ghosts. In passive ghosts, all the Ca movement could be described by
a single rate constant. The activity of the Ca pump was calculated from the
rate of net Ca uptake in the active ghosts, using the rate constant for
passive Ca movement as determined in the passive ghosts. fCai and the rates
of Ca transport in both active and passive ghosts were calculated from the
absorbance of entrapped arsenazo III. In general, increasing fCai from 1 to
10 microM activated the pump. Higher fCai caused an inhibition compared
with peak activity. The maximum rate of pumping was 80 microM/min. The
major new finding is that the rate of active transport at a given fCai
appeared to vary with the rate of fCai accumulation.(ABSTRACT TRUNCATED AT
250 WORDS)
