Equilibration and Exchange of Fluorescently Labeled Molecules in Skeletal Muscle Fibers Studied Using Confocal Microscopy

Abstract

Diffusion of molecules into skinned muscle fibers is often necessary when studying muscle contraction and its regulation. Usually, it is assumed that diffusion of molecules is fast also in the structured system of a muscle fiber, and that equilibration is reached within minutes or at most hours. One method to study not only equilibration, but also the time dependent distribution and the binding dynamics of fluorescently labeled molecules inside a muscle fiber is confocal laser fluorescence microscopy. Because of the principle of confocal imaging, one can obtain optical sections of a muscle fiber even under physiological conditions and with reasonable time and spatial resolution. In the present study, we used confocal microscopy to follow the equilibration of several fluorescently labeled substances in chemically skinned single muscle fibers of rabbit psoas. The investigation concerned mainly two questions: * What causes the equilibration time for molecules of the same size and type to be different from minutes to days? * Is the high affinity binding ofsome muscle-specific molecules readily reversible? Originally published Biophysical Journal, Vol. 68, No. 4 suppl, Apr 1995