Physiological assay of liposome-mediated transport of a drug across Xenopus intestine: cell-liposome interaction

(1) The antagonistic effect of atropine methyl bromide entrapped in liposomes on contraction of Xenopus intestine in vitro induced by acetylcholine was studied. The results provided some insight into cell-liposome interaction. (2) Acetylcholine (0.1 mM) was added to the medium in the bath (serosal solution), while liposomes containing atropine methyl bromide in their internal and external phases were added on the mucosal side of the intestine. Large multilamellar liposomes were prepared from egg lecithin (phosphatidylcholine, PC) and cholesterol in various molar ratios. Atropine methyl bromide had most effect in liposomes composed of PC and cholesterol in a ratio of 7:3, less in those with a ratio of 4:5, and none in those with a ratio of 9:1. These effects were parallel with the sizes of these liposomes, determined by quasi-elastic light-scattering; that is, the larger the liposomes, the greater was their effect. Addition (to the liposomes) of phosphatidic acid, the negative charge of which increases the distance between the lamellar layers, increased the effect, indicating that atropine methyl bromide in the space between lamellar layers was effective. Another type of liposomes in which atropine methyl bromide was present only in the external phase of liposomes was as effective as liposomes in which atropine methyl bromide was present in both the internal and external phases. (3) From these results the following new model for liposome-mediated stimulation of transport of atropine methyl bromide is proposed. Large multilamellar liposomes have structural defects in their external lamellae through which atropine methyl bromide in the mucosal solution can penetrate into the space between the external lamellar layers and move into intestinal cells through regions of fusion between the outermost layers of the liposomes and the cell membrane.     

Acid proteinase activity in fish--I. Comparative study of extraction of cathepsins B and D from Mujil auratus

Acid catheptic activity was measured in crude extracts of muscle, liver, heart, spleen and gonads from the fishes Mujil auratus, Sparus aurata and Lightonatus mormyrus. The spleen was the organ which showed the highest activity. A comparative study of the seven most commonly used extraction methods was made. Some were modified to account for the characteristics of the fish organs and the activity extracted from them. The Siebert method resulted as the best extraction method only if 1 mM EDTA was present in the medium. The activity from Mujil auratus muscle was strongly inhibited by iodoacetate, N-ethylmaleimide, p-hydroxy mercuribenzoate, and diazo-acetyl-DL-norleucine methyl ester. The results indicated the presence of a carboxyl-proteinase and a thiol-proteinase. According to inhibition studies, the levels of proteinase and amidase activities shown by different organs of Mujil auratus were re-examined. The spleen extract showed the maximum activity for both cathepsins, but muscle extract accounted for more than 95% of total catheptic activity.     

Some Observations on Factors Controlling Apical Dominance in the 'Rogue' Tomato

The rogue tomato differs from the normal plant in that it exhibitsa lesser degree of apical dominance. Grafting techniques andmeasurements of the endogenous levels of growth substances inthe two types have been used in order to establish whether thisdifference is due to an altered hormonal balance in the roguetype. The results suggest that root-produced cytokinins play no rolein the control of apical dominance in the tomato plant, andthat lateral bud out-growth is influenced by a balance betweenapically-produced auxin, abscisic acid produced at the sitesof bud development and cytokinins synthesized within the budsthemselves. Lycopersicon esculentum L., tomato, apical dominance, abscisic acid, auxins, cytokinins, growth regulation     

Composition and peptide maps of cross-linked human choriogonadotropin-receptor complexes on porcine granulosa cells

Radioiodinated human choriogonadotropin was affinity-cross-linked with a cleavable (nondisulfide) homobifunctional reagent to the hormone receptor on porcine granulosa cells and the solubilized sample was electrophoresed. Cross-linked samples revealed four additional bands of slower electrophoretic mobility in addition to the hormone alpha, beta, and alpha beta dimer bands. The four bands corresponded to masses of 68, 74, 102, and 136 kDa whereas the alpha beta dimer band corresponded to 50 kDa. Formation of the four bands requires the 125I-hormone to bind specifically to the receptor with subsequent cross-linking. Binding can be prevented by excess of native hormone but not by follitropin. A monofunctional analog of the cross-linking reagent failed to produce the four bands. They were also produced by cross-linking Triton X-100-solubilized hormone-receptor complexes. Reagent concentration-dependent cross-linking revealed that their formation was sequential; smaller complexes formed first and then larger ones. When gels of the cross-linked sample were treated with reagents that cleave covalent cross-links and then electrophoresed in a second dimension gel, 18-, 24-, 28-, and 34-kDa components were released, in addition to the alpha and beta subunits of the native hormone. Simultaneous peptide mapping of the cross-linked complexes in the gel matrix with Staphylococcus V8 protease or papain revealed progressive proteolysis to generate terminal fragments of 30 or 27 kDa, respectively. These fragments were unique to and commonly present in the 74-, 102-, and 136-kDa hormone-receptor complexes but were not produced by proteolysis of the cross-linked human choriogonadotropin (hCG) alpha beta dimer or the hCG alpha subunit. Apparently, the radioactively labeled segment(s) of the alpha subunit of 125I-hCG was cross-linked to the 24-kDa component. The results demonstrate the protein nature of the receptor and suggest that 125I-hCG was initially cross-linked to the 24-kDa component to generate the 74-kDa complex, then the 28- and 34-kDa components were sequentially cross-linked to the 24-kDa component in the 74-kDa complex to generate the 102- and 134-kDa complexes.     

Environmental stress and bacterial infection in channel catfish, Ictalurus punctatus Rafinesque

Channel catfish, Ictalurus punctatus , were injected intraperitoneally with a sublethal dose of Aerornonas hydrophila and then stressed for 144 h by being maintained either in a dissolved oxygen concentration of 1·5 mg/1, 1·2 mg/1 total ammonia, and/or 6·5 mg/1 free CO₂ with a continuous inflow of water. A significant difference in percentage of mortality was noted between treatments ( P < 0·05). The trunk kidneys of surviving stressed fish had significantly higher total bacterial counts than non-stressed controls. A. hydrophila was isolated from 67% of the stressed fish and 9% of the control fish. Edwardsiella tarda , apparently endemic in the population, was isolated from 43% of the stressed fish and 7% of the control fish. Histopathological lesions were in the gills, liver, spleen, trunk kidney, and head kidney of stressed fish, but not control fish.     

Liver microsome phospholipids and cytochrome P.450 conceptration in phenobarbital treated rats fed on different diets

Liver microsomal concentration of cytochrome P.450 is increased in animals which are fed diets rich in polyunsaturated fatty acids. On the other hand, the effects of phenobarbital are more important when the dietary fat is more unsaturated. The unsaturation index in liver microsomal phosphatidylcholines depends on the unsaturation of the dietary fats. The treatment with phenobarbital constantly results in a decrease of the unsaturation index of fatty acids both in lecithins and cephalins. The importance of the liver microsomal cytochrome P.450 increase and the importance of the unsaturation index decrease in liver microsomal lecithins, both promoted by phenobarbital, are in good agreement.     

In vivo phosphorylation of isocitrate lyase inEscherichia coli andAcinetobacter calcoaceticus

Isocitrate lyase inEscherichia coli and inAcinetobacter calcoaceticus is phosphorylated when the cells are grown with acetate as the sole carbon source in low-phosphate mineral salts medium containing³²P inorganic phosphate. The level of³²P incorporation into the enzyme in both microorganisms appears to be constant throughout the entire growth cycle. Further, theresults of immunoblots and rocket immunoelectrophoresis suggest that the amount of isocitrate lyase protein, although at different levels in each microorganism, also remains constant throughout the growth cycle.     

Effect of tween 80 on exploratory behavior and locomotor activity in rats

Exploratory behavior and locomotor activity is enhanced in male rat pups (aged 10 to 20 days) whose dams received a chronic dose (1.25 ml/l) of Tween 80 via their drinking water. This enhancement manifests itself during the diurnal period of the day. These data suggest that Tween 80 has an effect on the CNS which could lead to misinterpretation of results in toxicology studies that use this compound as a dosage vehicle.