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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 μ m −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 Ca 2+ 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.

Six mutants of Salmonella typhimurium LT2 with defects in the heptose region of the lipopolysaccharide (LPS) ("rough" mutants) were more sensitive to the growth-inhibitory effects of erythromycin, bacitracin, vancomycin, novobiocin, kanamycin, and cloxacillin and of deoxycholate than smooth strains, but less sensitive to tetracycline and ampicillin. In general, growth of the three rough mutants of chemotype Rd2, which lack the distal but not the proximal heptose unit in the LPS, was less inhibited than the three mutants of chemotype Re, which are heptose-deficient. In addition, inhibition of uracil-1-14C incorporation in the presence of actinomycin D and spheroplast formation in the presence of lysozyme occurred in the rough mutants without ethylenediaminetetraacetate (EDTA) treatment of the cells, while actinomycin D and lysozyme were effective on smooth strains only after EDTA treatment. Since the major part of the LPS is in the outer membrane of the cell envelope, and since the target of the toxic agents used is located inside this layer, these data indicate that the carbohydrate part of the LPS component of the outer membrane is an essential part of a barrier layer preventing penetration of large molecules.

Three structurally distinct amphiphiles palmitic acid, oleic acid, and palmityl carnitine were studied to determine their effects on sodium dependent calcium uptake by purified cardiac sarcolemmal vesicles (PSL). Sodium dependent calcium uptake by PSL when studied over a 20 min reaction period was composed of an initial rapid uptake (20.9 +/- 0.93 nmol/mg X 30 s, mean +/- S.E. n = 20) a plateau in calcium content (42.4 +/- 3.2 nmol/mg, mean +/- S.E. n = 20) and a slow spontaneous release characterized by a first order rate constant of 0.68 +/- 0.08/h (mean +/- S.E. n = 18). Both palmityl carnitine and palmitic acid inhibited, whereas oleic acid stimulated initial calcium uptake. All three amphiphiles shortened the time to peak calcium content, inhibited peak calcium content and increased the rate constant for calcium release. All these effects were observed at fatty acid: membrane phospholipid mole ratios of 0.67 : 1 to 1.67 : 1 for oleic acid and palmityl carnitine and 0.02 : 1 to 0.42 : 1 for palmitic acid. These effects do not reflect disruption of membrane vesicle structure and may be explained, at least in part, by amphiphile induced increases in sarcolemmal membrane ion permeability. Although amphiphile accumulation has been implicated in the pathogenesis of cellular abnormalities in the ischemic myocardium, this study has shown that large amounts of amphiphile relative to membrane lipid are required to alter sarcolemmal membrane function in vitro.