Quantitative analyses of topography and elasticity of living and fixed astrocytes

The topography and elasticity of living and fixed astrocytes cultured from the rat cerebra were studied quantitatively by atomic force microscopy (AFM ). Ridge-like structures reflecting F-actin beneath the cell membrane were prominent in the contact-mode images of living astrocytes. Many of these ri dges became unclear after fixation (2%, glutaraldehyde). In addition, the r idge-like structures were invisible in the topography of living cells obser ved at zero-loading force in the force mapping mode, which is considered to show the real cell surface not pressed down by an AFM tip. The topography of fixed cells observed both in the contact mode and at zero-loading force in the force mapping mode was similar to that of living cells observed at z ero-loading force in the force mapping mode, although some deformed areas w ere detected in the fixed cells. The elasticity map images of living astroc ytes showed that the cell membrane above the nucleus was softer (2-3 kPa) t han the surroundings, and that the cell membrane above F-actin was stiffer (10-20 kPa) than the surroundings. In the elasticity map images of fixed as trocytes, on the other hand, the elasticity of the cells was found to be re latively uniform (200-700 kPa) irrespective of the inner structures of cell s. These results show that images observed by AFM should be carefully exami ned in consideration of the force introduced to specimens and the elasticit y of specimens to find out the real surface topography.