Actin dynamics in growth cones.
The mechanism of actin incorporation and turnover in the nerve growth cone was examined by immunoelectron microscopy and low-light-level video microscopy of cultured neurons injected with biotin-labeled actin or fluorescently labeled actin. We first determined the sites of actin incorporation into the cytoskeleton of growth cones by immunoelectron microscopy of cultured neurons injected with biotin-labeled actin and reacted with an anti-biotin antibody and a gold-labeled secondary antibody. Shortly after the injection, biotin-actin molecules incorporated into the cytoskeleton were localized in the distal part of actin bundles in the filopodia and at the membrane-associated fringe of the actin filament network. With longer incubation, most actin polymers in the growth cones were labeled uniformly, suggesting that actin subunits are added preferentially at the membrane-associated ends of preexisting actin filaments. We then determined whether actin filaments translocate within the growth cones by low-light-level video microscopy of living neurons injected with fluorescently labeled actin and photobleached with a laser beam. When actin fluorescence at the leading edge of a growth cone was bleached, a rearward translocation of the bleached spot toward the base of the growth cone was observed. This observation suggests the presence of a rearward flow of actin polymers within growth cones. Taken together, these results indicate that there is a continuous addition of actin monomers at the leading edge of the growth cone and a successive rearward translocation of the assembled filaments.
Okabe
S
S
Department of Anatomy and Cell Biology, School of Medicine, University of Tokyo, Japan.
Hirokawa
N
N
D000199:Actins / Q000378:metabolism*; D000818:Animals; D001370:Axonal Transport; D001369:Axons / Q000378:metabolism* / Q000648:ultrastructure; D001710:Biotin; D002460:Cell Line; D003599:Cytoskeleton / Q000378:metabolism / Q000648:ultrastructure; D019793:Fluorescein; D005452:Fluoresceins; D005455:Fluorescent Antibody Technique; D005727:Ganglia, Spinal / Q000648:ultrastructure; D006046:Gold; D007700:Kinetics; D051379:Mice; D008856:Microscopy, Fluorescence; D016253:Microscopy, Immunoelectron; D009474:Neurons / Q000648:ultrastructure*; D010777:Photochemistry; D011108:Polymers / Q000378:metabolism
D000199:Actins; D005452:Fluoresceins; D011108:Polymers; D001710:Biotin; D006046:Gold; D019793:Fluorescein