A set of different biophysical approaches has been used to explore the phase behavior of palmitoylsphingomyelin (pSM)/cholesterol (Chol) model membranes in the presence and absence of palmitoylceramide (pCer). Fluorescence spectroscopy of di-4-ANEPPDHQ-stained pSM/Chol vesicles and atomic force microscopy of supported planar bilayers show gel Lb/liquid-ordered (Lo) phase coexistence within the range XChol = 0-0.25 at 22°C. At the latter compositional point and beyond, a single Lo pSM/Chol phase is detected. In ternary pSM/Chol/pCer mixtures, differential scanning calorimetry of multilamellar vesicles and confocal fluorescence microscopy of giant unilamellar vesicles concur in showing immiscibility, but no displacement, between Lo cholesterol-enriched (pSM/Chol) and gel-like ceramide-enriched (pSM/pCer) phases at high pSM/(Chol + pCer) ratios. At higher cholesterol content, pCer is unable to displace cholesterol at any extent, even at XChol < 0.25. It is interesting that an opposite strong cholesterol-mediated pCer displacement from its tight packing with pSM is clearly detected, completely abolishing the pCer ability to generate large microdomains and giving rise instead to a single ternary phase. These observations in model membranes in the absence of the lipids commonly used to form a liquid-disordered phase support the role of cholesterol as the key determinant in controlling its own displacement from Lo domains by ceramide upon sphingomyelinase activity. © 2010 by the Biophysical Society.
Busto, J. V., Sot, J., Requejo-Isidro, J., Goni, F. M., & Alonso, A. (2010). Cholesterol displaces palmitoylceramide from its tight packing with Palmitoylsphingomyelin in the Absence of a Liquid-Disordered Phase. Biophysical Journal, 99(4), 1119–1128. https://doi.org/10.1016/j.bpj.2010.05.032