Utilization of fluorescent lanthanide ions for the study of cation binding to extracellular sites in frog skeletal muscle

The binding of Tb³⁺ ions to frog muscle has been studied, using fluorescence measurements. Kinetic evidence suggests that Tb³⁺ binds to the muscle at multiple sites. Two major classes can be distinguished: One having relatively fast kinetics and saturating within a few minutes; the second type being either very slow (hours) or consisting of an infinite pool. Binding to the first class of sites is readily reversible and is probably extracellular. It is shown that part of the slow type of binding is also reversible, and is also associated with extracellular binding sites. The binding of terbium is antagonized by other metal ions, their order of effectiveness being: La³⁺, Eu³⁺ > Mn²⁺, Hg²⁺ ≫ Ca²⁺ > Mg²⁺. A comparison with other studies suggests that part of the binding sites are located at the surface membrane of the muscle. Our experiments raise the possibility that part of the slowly exchanged calcium ions may well be of extracellular origin rather than intracellular one. The limitations and advantages of such studies, as a means to probe ion interactions in complex biological systems are discussed.