A functional in vitro model for studies of intracellular motility in digitonin-permeabilized erythrophores.

Abstract

Phase contrast cine results demonstrate that erythrophores maintain saltatory particle motion for hours after permeabilization with 0.001% digitonin in a cytoskeletal stabilizing solution at 23 degrees C. High voltage electron microscopy (HVEM) studies reveal that cytoskeletal elements are retained intact, except in immediate subplasmalemmal regions where the plasma membrane is punctured by digitonin. During digitonin treatments, cells are permeable to ions, small molecules, and antibodies. We find that motion is Ca2+ and ATP-sensitive, and optimal in PIPES buffer (pH 7.2 containing 1 mM Mg2+/ATP and EGTA-CA2+ (10(-7) M Ca2+) at 37 degrees C. Experiments testing the inhibitory effects of vanadate (0.4-10 microM), ouabain (100-600 microM), N-ethyl maleimide, and the cytochalasins B and D indicate that a dyneinlike ATPase may provide the motive force for driving saltatory pigment motion in erythropores.