Analysis of cellular mechanoreceptors by atomic force microscopy

Guillaume Charras and Michael Horton

Department of Medicine, University College London,
London WC1E 6JJ, UK.

Atomic Force Microscopy (AFM) can be used to apply forces onto cells as well as to determine cellular material properties and strains. Bone adapts its structure to its mechanical usage. The detection of the strain applied to bones is effected by cells of the osteoblast lineage. An increase in intracellular calcium concentration is one of the earliest events in the detection of mechanical stimuli by cells. We used AFM (ThermoMicroscopes Explorer AFM) to study mechano-transduction in primary osteoblasts. While forces were applied onto cells via the AFM, intracellular calcium levels were monitored in cells loaded with the calcium sensitive dye, Fluo 3, via an integrated Bio-Rad Radiance 2000 confocal microscope. The material properties of the cells were calculated and the strains applied derived. The cells responded to strains in a dose-dependent manner. Mechano-sensitive channels, phospholipase C, intracellular calcium stores and voltage activated calcium channels played an important role in the signal transduction pathway. Cellular reactions could also be modulated by using cytoskeleton disrupting drugs such as cytochalasin B, jasplakinolide, nocodazole, taxol, acrylamide and diamide. In conclusion, AFM has been used to examine the behaviour of osteoblasts in response to strain, a process of key importance in our understanding of the mechanism by which the adaptation of the skeleton to mechanical forces is regulated.