Effects of culture and incubation conditions on membrane fluidity in monolayers of cultured cells measured as fluorescence anisotropy using trimethylammoniumdiphenylhexatriene (TMA-DPH).
Membrane fluidity of coverslip attached living cells was measured as fluorescence anisotropy using 5 microM trimethylammoniumdiphenylhexatriene (TMA-DPH) as fluorescent probe. Fluorescence anisotropy is inversely related to membrane fluidity. Cells were grown on glass coverslips that were inserted and directly incubated in quarz cuvettes. The coverslips were fixed with special holders at an angle of 30 degrees in respect to the incident light. Effects of incubation temperature, of cell growth and densities and of the ionic and nonionic composition of the incubation medium on membrane fluorescence anisotropy were measured. Membranes of growing cells were more fluid than those of stationary cells, while cell densities had no effect except at very low cell numbers. Calcium concentrations increasing from 0 to 8 mmol/l in the incubation medium proportionally decreased membrane fluidity. Hypotonicity of the incubation media increased membrane fluidity while hypertonicity compared to normotonicity had no effect. Differentiated human fibroblasts from different origins exhibited similar membrane fluidities. They were, however, different from those of rat cells. Membrane fluidity of rat brain tumor cells increased with age in culture while membrane fluidity of primary differentiating rat brain cells decreased in with age in culture. Measurement of fluorescence anisotropy in living cells attached to glass coverslips is a convenient tool to study effects of culture--as well as of environmental--conditions on membrane fluidity.
Department of Pediatrics, University of Bern, Switzerland.
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H
Toplak
V
V
Batchiulis
A
A
Hermetter
T
T
Hunziker
U E
UE
Honegger
U N
UN
Wiesmann
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002452:Cell Count; D002455:Cell Division; D004161:Diphenylhexatriene / Q000031:analogs & derivatives*; D005347:Fibroblasts / Q000502:physiology; D005454:Fluorescence Polarization; D005456:Fluorescent Dyes; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007038:Hypotonic Solutions; D008560:Membrane Fluidity; D051381:Rats; D014407:Tumor Cells, Cultured
D005456:Fluorescent Dyes; D007038:Hypotonic Solutions; D004161:Diphenylhexatriene; C033628:1-(4-(trimethylamino)phenyl)-6-phenylhexa-1,3,5-triene; D002118:Calcium