Distribution and dynamics of fluorochromed actin in living stages of Physarum polycephalum.

Isolated muscle and Physarum actins were labeled with various fluorochromes and microinjected into living stages of Physarum polycephalum (caffeine-droplets, endoplasmic drops, thin-spread macroplasmodia). Subsequent analysis of the intracellular redistribution by fluorescence microscopy, video-enhancement and digital image processing revealed RITC (rhodamineisothiocyanate) actin to be the most reliable molecular probe for the marking of microfilaments. In relaxed caffeine-droplets, the RITC-actin first diffuses randomly and then is locally incorporated into a thin cortical layer at the internal face of the plasma membrane. During Ca2+-induced contraction the fluorescent layer starts to detach from the plasma membrane, thus causing separation of central granuloplasm from peripheral hyaloplasm. Thin sections of both, relaxed and contracted specimens demonstrated that the RITC-actin layer in living droplets exactly coincides with a sheath of more or less oriented microfilaments. In contrast, RITC-bovine serum albumin (BSA) injected as control is excluded from those regions which show intense fluorescence with RITC-actin and the presence of an actin network with EM. Successful incorporation of the molecular probe into stages of Physarum polycephalum other than caffeine droplets was not yet achieved. The results obtained by fluorescent analog cytochemistry (FAC) are discussed with regard to the spatial organization of the actin system in acellular slime molds.