Multiple microscopic techniques for the measurement of plasma membrane lipid structure during hypoxia

Abstract

Alterations in plasma membrane structure and function are considered of primary importance in the pathogenesis of cell injury. Multiple microscopic techniques are employed to detail alterations in plasma membrane lipid structure during hypoxic injury in individual rat hepatocytes. Multiparameter digitized video microscopy, fluorescence quenching imaging, and fluorescence resonance energy transfer imaging are used to measure and monitor lipid domain formation and topography; laser tweezers are used to monitor the plasma membrane viscoelasticity. These microscopic techniques indicate that hypoxic injury in hepatocytes leads to alterations in plasma membrane lipid topography with the eventual formation of lipid domains. In concert with previous data generated with digitized fluorescence polarization microscopy and fluorescence recovery after photobleaching (FRAP), a model is proposed where formation of the distinct lipid domains promotes loss of the plasma membrane permeability barrier and cell death.