Heterogeneous calcium and adenosine triphosphate distribution in calcium‐permeabilized human red cells.

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Abstract
1. Calcium permeabilization of inosine-fed human red cells using the divalent cation ionophore A23187 induces pump-leak steady states in which the mean total calcium content of the cells may be held below electrochemical equilibrium for hours. A new method developed to detect and separate cells with different calcium contents revealed a striking heterogeneity of calcium contents in subpopulations of cells in pump-leak steady state (García-Sancho & Lew, 1988a). Most of the mean total cell calcium was found within a fraction of cells rendered dense by the separation procedure (H cells), with relatively little within the remaining light cells (L cells). The experiments in this paper were designed to study the nature and origin of the observed heterogeneity. 2. The fraction of steady-state H cells increased, and the mean ATP content of the cells fell, both linearly, as calcium influx was increased. The H/L divide is therefore the result of a continuous variation in cell properties. When calcium influx was above about 30 mmol/(l cells.h), all cells became dense, calcium distribution was at or near equilibrium, and cell ATP was 0.1-0.2 mmol/l cells. 3. Inosine-fed cells, subjected to ionophore-mediated net calcium influx of 13-15 mmol/(l cells.h), attained a steady state with mean calcium contents far below equilibrium. After ionophore removal and reincubation in calcium-free media, the initial calcium efflux was only a fraction of that required to sustain the previous steady state (less than 25% for H cells, and less than 2% for L cells). The ATP content of L cells was normal whereas that of H cells was irreversibly reduced. These results revealed a paradoxical discrepancy between leak influx and calcium pump efflux in H and L cells which were supposed to have been in steady-state pump-leak balance. 4. The changes in cell calcium and ATP were followed in time after calcium permeabilization to characterize the development of steady-state heterogeneity. Calcium influx triggered a sharp peak in the H cell fraction within 15 s of permeabilization. The mean calcium content of H cells increased towards steady-state values as their fraction decreased; most other cells transferred from H to L density fractions (HL cells) within the first 5 min of permeabilization. 5. In substrate-starved cells calcium influx triggered an immediate fall in cell ATP, steeper in H cells than in L cells. The initial calcium and density transients were unattected.(ABSTRACT TRUNCATED AT 400 WORDS)

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