Lipid membrane perturbations, fluorescence

The modulation of synaptosomal plasma membranes (SPMs) by adriamycin and the resultant effects on the activity of membrane-bound enzymes have been reported [58]. Again DPH was used as fluorescence probe. Adriamycin increased the lipid fluidity of the membrane labeled with DPH, as indicated by the steady-state fluorescence anisotropy. The lipid-phase separation of the membrane at 23.3 °C was perturbed by adriamycin so that the transition temperature was reduced to 16.2 °C. At the same time it was found that the Na+,K+-stimulated ATPase activity exhibits a break point at 22.8 °C in control SPMs. This was reduced to 15.8 °C in adriamydn-treated SPMs. It was proposed that adriamycin achieves this effect through asymmetric perturbation of the lipid membrane structure and that this change in the membrane fluidity may be an early key event in adriamycin-induced neurotoxicity. 

Often, experimental studies of lipid systems are based on spectroscopic approaches, which in turn frequently employ probes for enhancement of sensitivity and resolution. For example, in NMR, hydrogen atoms of lipids are replaced with deuterium, and in fluorescence spectroscopy and imaging, native lipid molecules are replaced with lipids in which one of the hydrocarbon chains is linked covalently to a fluorescent marker such as pyrene or diphenylhexatriene. Fluorescent markers allow one to follow numerous cellular processes in real time, such as intracellular trafficking of molecules and formation of domains within a biomembrane, see Fig. 3. The downside is that the probes tend to perturb their environment and affect the thermodynamic state of the system. Experiments have shown, for example, that probes may change the main transition temperature of a lipid membrane, and that the dynamics of probes may deviate considerably from the dynamics of corresponding native molecules (see discussion in Reference 27). Therefore, we wish to pose several questions. What is the range of perturbations induced by the probe How significant are these perturbations actually ... 

The polarization properties of light in combination with fluorescence can be used as a powerful tool for determining motional properties of membranes. This is possible due to the fact that the time scale of interest for membrane lipids falls within the time frame of the fluorescence decay phenomena (0-100+ ns). This, coupled with high sensitivity, low perturbing properties of fluorescent probes, and the large number of available probes, makes the fluorescence approach the method of choice for membrane motional studies. 

Jelinek and co-workers demonstrated that the changes in fluorescence of PDA can be used to detect the interactions of drug molecules with the membranes of live cells. Diacetylene patches containing 10,12-tricosadiynoic acid were transferred from diacetylene vesicles to the membranes of leukemia cells and were polymerized with UV radiation. This fusion efficiency was found to depend on the lipid composition of the liposomes and the presence of cholesterol on the plasma membrane. The interactions of these cells with lidocaine (a local anesthetic), polymixin-B (an antibiotic), and oleic acid were studied by confocal fluorescence microscopy. The PDA patch on the live cells showed bright red fluorescence when the cell membranes were perturbed. The blue to red color change in the presence of oleic acid was apparent when the cells were sedimented by centrifugation. 

Effects of HO radicals on biological membranes include alterations in membrane proteins, peroxidation of unsaturated lipids accompanied by perturbation of the lipid bilayer polarity. Berroud et al. (1996) have measured radiation-induced membrane modifications using two fluorescent lipophilic membrane probes (TMA-DPH and DPH) by the technique of fluorescence polarization of Chinese hamster ovary K1 and lymphoblastic RPMI 1788 cell lines, ylrradiation from a Co source with dose rates of 0.1 and 1 Gy/min for a final dose of 4 and 8 Gy induced a dose-dependent decrease of... 

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