Physical properties of biological membranes determined by the fluorescence of the calcium ionophore A23187

Abstract

Interactions between the divalent cation ionophore, A23187, and the divalent cations Ca 2+, Mg 2+, and Mn 2+ were studied in sarcoplasmic reticulum and mitochondria. Conductance measurements suggest that A23187 facilitates the movement of divalent cations across bilayer membranes via a primarily electroneutral process, although a cationic form of A23187 does carry some current. On the basis of fluorescence excitation spectra, A23187 can form either a 1:1 or 2:1 complex with Ca 2+ in organic solvents. However, in biological membranes, only the 1:1 complexes with Ca 2+, Mg 2+, or Mn 2+ are detected. A23187 produces fluorescent transients under conditions of Ca 2+ uptake in sarcoplasmic reticulum, which appear to represent changes in intramembrane Ca 2+ content. Changes in A23187 fluorescence due to mitochondrial Ca 2+ accumulation are much smaller by comparison and fluorescence transients are not detected. Studies of A23187 fluorescence polarization and lifetimes in biological membranes allow a determination of the rotational correlation time (ρh) of the ionophore. In mitochondria at 22 °C, ρh is 11 nsec in the presence of Ca 2+ and Mg 2+, and less than 2 nsec in the presence of excess EDTA. The present results are consistent with a model of ionophore-mediated cation transport in which free M 2+ binds with A23187 at the membrane surface to form the complex M(A23187) +. Reaction of this complex with another molecule of A23187 at the membrane surfaces results in the formation of electrically neutral M(A23187) 2, which carries the divalent cation through the membrane. These results are discussed in terms of physical properties of biological membranes in regions in which divalent cation transport occurs.