Structure-function studies of canine cardiac sarcolemmal membranes. II. Structural organization of the sarcolemmal membrane as determined by electron microscopy and lamellar X-ray diffraction

Abstract

The morphological and ultrastructural properties of highly purified canine cardiac sarcolemmal vesicles, prepared by a modification (Colvin, R.A., Ashavaid, T.F. and Herbette, L.G. (1985) Biochim. Biophys. Acta 812, 601-608) of the method of Jones et al. (Jones L.R., Madlock, S.W. and Besch, H.R. (1980) J. Biol. Chem. 255, 9971-9980), were examined by several techniques. Thin-section electron microscopy showed predominantly intact unilamellar vesicles with little staining beyond the lipid bilayer boundaries. Freeze-fracture electron microscopy demonstrated that the majority of particles are approx. 90 A diameter and present at a density of 780 +/- 190 micrometers-2 (+/- S.D.). If it is assumed that some of these particles represent the (Na+ + K+)-ATPase, the finding that they are largely confined to the convex fracture face suggests a predominant right-side-out orientation of these sarcolemmal vesicles that is consistent with biochemical assays. The sarcolemmal membrane width measured by electron microscopy (unhydrated membrane width of 50-70 A) is consistent with the unit cell dimensions of 56-77 A determined by lamellar X-ray diffraction (hydrated membrane width). A unit cell dimension of 56-62 A was also found by X-ray diffraction for sarcolemmal lipids extracted from these preparations, indicating that the isolated sarcolemmal preparations do not contain a significant surface coat (glycocalyx). As both cardiac and skeletal sarcoplasmic reticulum membranes have a 80-100 A membrane width, these findings demonstrate that the purified sarcolemmal membrane is structurally distinct from both cardiac and skeletal sarcoplasmic reticulum. In contrast to the protein-rich skeletal sarcoplasmic reticulum membrane, which contains a single essential protein responsible for the regulation of cytosolic Ca2+ concentration, the sarcolemma is a lipid-rich membrane that contains a variety of proteins associated with many regulatory functions served by this membrane in cardiac muscle.