Isolation of Raja erinacea basolateral liver plasma membranes: characterization of lipid composition and fluidity

Developing a method for isolating skate (Raja erinacea) basolateral liver plasma membranes, as well as characterizing the lipid composition and fluidity of these membranes, was the primary purpose of this study. Membranes were isolated using self-generating Percoll gradients. Marker enzyme studies indicate that this preparation is highly enriched in the basolateral domain of the liver plasma membrane and largely free of contamination by intracellular organelles or canalicular membranes. Further, these membranes contain the agency responsible for Na(+)-dependent alanine transport. This finding indicates that this membrane preparation will be useful for the study of skate liver plasma membrane transport processes. The lipid composition and fluidity (as assessed by the fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene) of the skate basolateral liver plasma membrane shows little variation among preparations. Further, DPH anisotropy plotted as a function of temperature yields a straight line (r = 0.99) which indicates that there is no lipid phase change in these membranes from 4 degrees to 37 degrees C. The membrane preparation does contain substantial phospholipase A2 activity. The function of this enzyme is, in part, to modify membrane lipid composition and fluidity in response to temperature variations; therefore, this finding suggests that in situ lipid metabolizing enzymes may play a central role in the adaptation of skate basolateral liver plasma membranes to changes in the ambient temperature.