title: Dynamics of leading lamellae of living fibroblasts visualized by high-speed scanning probe microscopy.
abstract: In this study, we aimed at improving the temporal resolution of scanning probe microscopy (SPM) for observing living cells by introducing soft cantilevers, low feedback-gain operations, and cantilever deflection imaging. We achieved visualization of the mechanical architecture in leading lamellae of living fibroblasts at a temporal resolution of around 10 s, which is higher than that of conventional contact-mode SPM. Time-lapse SPM could be used to monitor not only cytoskeletal dynamics but also the dynamics of numerous microgranules. Statistical analysis of microgranular motion revealed that the microgranules have superdiffusive behaviors and significant directional order of motion. We also found that the direction of their motion is correlated with the direction of growing actin stress fibers. The combination of SPM with fluorescence microscopy showed that vinculin, a component of cell-substratum adhesion sites, localizes at the microgranules. Our experimental data provides a new insight into the intracellular mechanical architecture and its structural dynamics, suggesting that high-speed live-cell SPM has great potential for investigating the structural origin of cellular dynamics.
authors.affiliation: Division of Biological Sciences, Graduate School of Science, Hokkaido University, Kita-ku, Sapporo, Japan. tam@mail.sci.hokudai.ac.jp
authors.forename: Kazushi
authors.initials: K
authors.lastname: Tamura
authors.forename: Takeomi
authors.initials: T
authors.lastname: Mizutani
authors.forename: Hisashi
authors.initials: H
authors.lastname: Haga
authors.forename: Kazushige
authors.initials: K
authors.lastname: Kawabata
mesh_terms: D000577:Amides / Q000494:pharmacology; D000818:Animals; D002470:Cell Survival; D002478:Cells, Cultured; D015638:Cytochalasin D / Q000494:pharmacology; D005347:Fibroblasts / Q000166:cytology* / Q000187:drug effects / Q000648:ultrastructure*; D051379:Mice; D008856:Microscopy, Fluorescence; D020527:Microscopy, Scanning Probe / Q000379:methods*; D041681:NIH 3T3 Cells; D011554:Pseudopodia / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure*; D011725:Pyridines / Q000494:pharmacology
chemical_list: D000577:Amides; D011725:Pyridines; C108830:Y 27632; D015638:Cytochalasin D