Effect of monensin on rumen metabolism in vitro.

The effect of Monensin (Rumensin, Eli Lilly & Co.) in incubations with mixed rumen microorganisms metabolizing carbohydrate or protein substrates was investigated. Monensin partly inhibited methanogenesis and increased propionate production, although the effect was not always statistically significant. Incubations with substrates specific for methane bacteria suggest that inhibition of methanogenesis by Monensin was not due to a specific toxic action on the methanogenic flora, but rather to an inhibition of hydrogen production from formate. Total and net microbial growth were considerably decreased by addition of Monensin, although the amount of substrate fermented was not altered, resulting in lowered values of microbial growth efficiency. In incubations with casein, Monensin lowered protein degradation in line with a lowered ammonia production, whereas a slight accumulation of alpha-amino nitrogen was observed. The results suggest that besides an influence of Monensin on the rumen carbohydrate fermentation pattern, another reason for the beneficial effects observed in vivo might be decreased food protein degradation in the rumen, altering the final site of protein digestion in the animal. Also, the possibility of a decrease in rumen microbial growth efficiency has to be considered when using Monensin as a food additive.     

On the lack of inotropy of cardiac glycosides on skeletal muscle: a comparison of Na+, K+-ATPases from skeletal and cardiac muscle.

Comparison of Na,K-ATPase from skeletal and cardiac muscle revealed that, although the skeletal muscle enzyme was only slightly less sensitive to inhibition by ouabain, the rates of [³H]ouabain binding to, and dissociation from, the skeletal enzyme were much faster than the corresponding rates for the cardiac enzyme. The skeletal muscle enzyme required higher concentrations of potassium to stabilize the ouabainenzyme complex and to stimulate the K⁺-phosphatase activity. The K⁺-phosphatase activity was only 8% of the Na,K-ATPase activity of the skeletal muscle enzyme, compared to 22% for the cardiac preparation. The glycoprotein subunit found in Na,K-ATPases from cardiac and many other tissues appeared to be absent in the enzyme from skeletal muscle. The differences in binding and dissociation rates for ouabain suggest that there may be significant differences in the structure of the digitalis receptor in the two enzymes. The I₅₀ for ouabain inhibition of the skeletal muscle Na,K-ATPase was, however, only slightly higher than for the cardiac enzyme, suggesting that the lack of an inotropic effect of cardiac glycosides on skeletal muscle could not be due to failure of the digitalis drugs to bind to and inhibit the membrane-linked sodium pump.     

Distribution and some properties of NADPH and NADH oxidase in parenchymal and nonparenchymal liver cells

The activities of NADPH and NADH oxidase were determined in homogenates of isolated pure parenchymal and nonparenchymal rat liver cells at neutral (7.4) and acid (5.5) pH. The NADPH oxidase at pH 7.4 is about equally active in parenchymal and nonparenchymal cells and in both cell types is rather insensitive to KCN (1 mm) inhibition. By lowering the pH to 5.5, the NADPH oxidase of the nonparenchymal cells is stimulated (twofold) while the activity in parenchymal cells is decreased. The NADH consumption at neutral pH in parenchymal cells is 75% inhibited by KCN, while this activity in nonparenchymal cells is relatively insensitive to KCN. The NADH oxidase in both parenchymal and nonparenchymal liver cells is less active when the pH is lowered from 7.4 to 5.5. The distribution of NAD(P)H oxidases between parenchymal and nonparenchymal liver cells and the effect of pH on their activities suggest that in the nonparenchymal cells, the NADPH oxidase might play a role in the synthesis of H₂O₂ within the phagocytic vacuole. A scheme is proposed which describes the metabolic events involved in H₂O₂ formation and catabolism of endo(phago)cytosed particles in nonparenchymal liver cells.     

The protein-mediated net transfer of phosphatidylinositol in model systems

The phospholipid monolayer technique has been used to study the transfer activity of the phospholipid exchange protein from beef brain. In measuring the transfer between a monolayer consisting of equimolar amounts of phosphatidylcholine and phosphatidylinositol and liposomes consisting of 98 mol% phosphatidylcholine and 2 mol% phosphatidylinositol, the beef brain protein demonstrates an 8-fold higher transfer activity for phosphatidylinositol than for phosphatidylcholine. Under similar conditions the phosphatidylcholine exchange protein from beef liver showed a great preference for phosphatidylcholine. Phosphatidylcholine liposomes devoid of phosphatidylinositol still functioned as receptors of phosphatidylinositol when the beef brain exchange protein was present. This indicates that this protein can catalyse a net transfer of phosphatidylinopsitol. Binding of both phosphatidylinositol and phosphatidylcholine to the beef brain protein was shown.     

Miscibility properties of binary phosphatidylcholine mixtures. A calorimetric study

From data obtained by differential scanning calorimetry phase diagrams were constructed, using a thermodynamically based fitting method. The following binary mixtures of phosphatidylcholines in water were studied: 14:0/ 14:0-glycerophosphocholine/16:0/16:0-glycerophosphocholine, 14:0/14:0-glycerophosphocholine /18:0/18:0-glycerophosphocholine, 12:0/12:0-glycerophosphocholine /16:0/16:0-glycerophosphocholine, 18:1_t/18:1_t-glycerophosphocholine /14:0/14:0-glycerophosphocholine and 18:1_t/18:1_t-glycerophosphocholine /16:0/16:0-glycerophosphocholine.A comparison is made of the present results with those obtained using probe techniques and the differences are discussed.     

Aggregative properties of different cell types of the fresh-water spongeEphydatia fluviatilis isolated on ficoll gradients

Summary Cell suspensions of the fresh-watersponge Ephydatia fluviatilis have been fractionated by means ofFicoll gradient centrifugation. Three fractions were isolated. The densest contains archeocyte-like cells only; the intermediate fraction is very rich in choanocytes, and the lightest is a mixture of cell types. Earch fraction shows specificaggregative properties and potentialities to reconstitute functional sponges.It appears that the sequence of reconstitution events can be selectively altered by certain disequilibria in the cell populationThese preliminary results constitute a first approach to the analysis ofcell type specificity in sponges.     

Voltage-dependent orientation of membrane proteins

In order to study the influence of electrostatic forces on the disposition of proteins in membranes, we have examined the interaction of a receptor protein and of a membrane-active peptide with black lipid membranes. In the first study we show that the hepatic asialoglycoprotein receptor can insert spontaneously into lipid bilayers from the aqueous medium. Under the influence of a trans-positive membrane potential, the receptor, a negatively charged protein, appears to change its disposition with respect to the membrane. In the second study we consider melittin, an amphipathic peptide containing a generally hydrophobic stretch of 19 amino acids followed by a cluster of four positively charged residues at the carboxy terminus. The hydrophobic region contains two positively charged residues. In response to trans-negative electrical potential, melittin appears to assume a transbilayer position. These findings indicate that electrostatic forces can influence the disposition, and perhaps the orientation, of membrane proteins. Given the inside-negative potential of most or all cells, we would expect transmembrane proteins to have clusters of positively charged residues adjacent to the cytoplasmic ends of their hydrophobic transmembrane segments, and clusters of negatively charged residues just to the extracytoplasmic side. This expectation has been borne out by examination of the few transmembrane proteins for which there is sufficient information on both sequence and orientation. Surface and dipole potentials may similarly affect the orientation of membrane proteins.