MOVEMENT OF AXOPLASMIC ORGANELLES ON ACTIN-FILAMENTS FROM SKELETAL-MUSCLE

It was recently shown that, in addition to the well-established microt ubule-dependent mechanism, fast transport of organelles in squid giant axons also occurs in the presence of actin filaments [Kuznetsov et al ., 1992, Nature 356:722-725]. The objectives of this study were to obt ain direct evidence of axoplasmic organelle movement on actin filament s and to demonstrate that these organelles are able to move on skeleta l muscle actin filaments. Organelles and actin filaments were visualiz ed by video-enhanced contrast differential interference contrast (AVEC -DIC) microscopy and by video intensified fluorescence microscopy. Act in filaments, prepared by polymerization of monomeric actin purified f rom rabbit skeletal muscle, were stabilized with rhodamine-phalloidin and adsorbed to cover slips. When axoplasm was extruded on these cover slips in the buffer containing cytochalasin B that prevents the forma tion of endogenous axonal actin filaments, organelles were observed to move at the fast transport rate. Also, axoplasmic organelles were obs erved to move on bundles of actin filaments that were of sufficient th ickness to be detected directly by AVEC-DIC microscopy. The range of a verage velocities of movement on the muscle actin filaments was not st atistically different from that on axonal filaments. The level of moti le activity (number of organelles moving/min/field) on the exogenous f ilaments was less than on endogenous filaments probably due to the ent anglement of filaments on the cover slip surface. We also found that c almodulin (CaM) increased the level of motile activity of organelles o n actin filaments. In addition, CaM stimulated the movement of elongat ed membranous organelles that appeared to be tubular elements of smoot h endoplasmic reticulum or extensions of prelysosomes. These studies p rovide the first direct evidence that organelles from higher animal ce lls such as neurons move on biochemically defined actin filaments. (C) 1994 Wiley-Liss, Inc.