Real-time imaging of drug-membrane interactions by atomic force microscopy

Understanding drug-biomembrane interactions at high resolution is a key issue in current biophysical and pharmaceutical research. Here we used real-time atomic force microscopy (AFM) imaging to visualize the interaction of the antibiotic azithromycin with lipid domains in model biomembranes. Various supported lipid bilayers were prepared by fusion of unilamellar vesicles on mica and imaged in buffer solution. Phase-separation was observed in the form of domains made of dipalmitoylphosphatidylcholine (DPPC), sphingomyelin (SM), or SM/cholesterol (SM/Chl) surrounded by a fluid matrix of dioleoylphosphatidylcholine (DOPC). Time-lapse images collected following addition of 1 mM azithromycin revealed progressive erosion and disappearance of DPPC gel domains within 60 min. We attribute this effect to the disruption of the tight molecular packing of the DPPC molecules by the drug, in agreement with earlier biophysical experiments. By contrast, SM and SM-Chl domains were not modified by azithromycin. We suggest that the higher membrane stability of SM-containing domains results from stronger intermolecular interactions between SM molecules. This work provides direct evidence that the perturbation of lipid domains by azithromycin strongly depends on the lipid nature and opens the door for developing new applications in membrane biophysics and pharmacology.