Diffusion of a small molecule in the cytoplasm of mammalian cells

Electron spin resonance was used to measure the diffusion of a small (M(r) 170) spin label in the aqueous cytoplasm of mammalian cells. Translational and rotational motion were determined from the same spectra. Based on measurements made in model systems, it was hypothesized that calculations of the apparent viscosity from either rotational or translational motion would distinguish between the effects of cytoplasmic viscosity or cytoplasmic structure on diffusion. The diffusion coefficient calculated from spin label collision frequency, averaged 3.3 x 10^-6 cm²/sec in several cell lines. It was greater in cells and in cells treated with cytochalasin B than in quiescent cells. The viscosity of the cytoplasm calculated from the translational diffusion coefficient or the rotational correlation time was 2.0-3.0 centipoise (1 P = 0.1 Pa·sec), about 2-3 times that of the spin label in water. Therefore, over the dimensions measured by the technique, 50-100 Å, solvent viscosity appears to be the major determinant of particle movement in cells under physiological conditions. However, when cells were subjected to hypertonic conditions, the translational motion decreased by 67%, while the rotational motion changed less than 20%. These data suggested that the decrease in cell volume under hypertonic conditions was accompanied by an increase in cytoplasmic barriers and a decrease in the spacing between existing components. In addition, a comparison of reported values for diffusion of a variety of molecules in water and in cells indicates that cytoplasmic structure plays an important role in the diffusion of proteins such as bovine serum albumin.