Increased GH responsiveness to dopamine receptor stimulation in alcohol addicts during the late withdrawal syndrome

In humans the release of growth hormone (GH) elicited by dopamine (DA) and DA agonists may represent a reliable model to assess change in sensitivity of DA receptors. We now report that in chronic alcoholics, 4–7 days after the suspension of alcohol consumption, the increase of GH response to DA infusion was higher than that seen in non alcoholic volunteers. The specificity of this GH response to DA administration was demonstrated by the use of domperidone, a novel peripheral antagonist of DA receptors. These results suggest the development of hyper-responsiveness of DA receptors involved in the control of GH secretion in chronic alcoholics during the later phases of the “withdrawal syndrome”.     

Temperature-related divergence in experimental populations of Drosophila melanogaster. II. Correlation between fitness and body dimensions

From a laboratory stock of Drosophila melanogaster (Oregon), reared for more than 20 years at 18° C, a new population was derived and maintained at 28° C for 8 years. The chromosomal and cytoplasmic contribution to genetic divergence between the two populations was estimated. Six body traits and reproductive fitness were taken into account. The third chromosome is responsible for the adaptive difference for temperature between the two lines. Temperature-selected genes which control body size are located on the second and third chromosomes, although the contribution of each chromosome depends on the environment in which the flies develop. The correlation between the chromosomal and cytoplasmic contributions to different traits and fitness, changes with temperature. At 28° C the correlation between fitness and each body trait is proportional to the response to selection exhibited by each of them, but this is not true at 18° C. Body size has, therefore, an adaptive significance in relation to temperature, which is expressed only in the environment where selection occurs. Cytoplasmic genes affect almost all characters to an extent similar to that of chromosomal genes. Inter-chromosomal and nucleo-cytoplasmic interactions are present and also change with temperature. In general, genes selected in a given environment produce greater phenotypic changes in that environment than in another. The population that experienced both temperatures is fitter in both environments, suggesting that the capacity to adapt to warm temperatures depends on genes other than those which are involved in the adaptation to cold.     

Electrical conductivity dispersion as a probe of membrane modifications in mouse polyomavirus infected cells in culture

In this report we investigate the inhibition of membrane conductivity, due to the murine polyomavirus infection in permissive cells in culture. We define experimental conditions to have reproducible results and demonstrate that the intensity of the effects on the cell membrane, depends upon the virus titer used in the infection. Finally, the virus dependent effects disappear if the infection is performed in the presence of a drug that inhibits polymavirus DNA replication.     

Changes in drinking water selenium and mortality for coronary disease in a residential cohort

In a part of the municipal territory of Reggio Emilia, northern Italy, selenium in drinking water decreased from 7 μg/L to less than 1 μg/L. In a cohort of 4419 individuals, previously exposed for at least 5 yr to the drinking water with higher selenium content, the 7-yr temporal distribution of deaths for coronary disease and for stroke was analyzed to examine a possible relationship with changes in drinking water selenium. From January 1986 until August 1988, when tap water selenium was 7 μg/L, deaths for coronary disease were one in males and two in females. After the decrease in drinking water selenium, 21 and 10 coronary deaths were observed, respectively, in males and in females from September 1988 to December 1992. No significant difference in the temporal distribution of stroke deaths was observed both in males and in females. Even if an effect of chance and aging in the temporal distribution of coronary deaths may not be excluded, findings of the study seem to be consistent with the hypothesis of a beneficial effect of selenium on coronary disease mortality.     

Intergenic spacers of rRNA genes in three species of theCynareae (Asteraceae)

Intergenic spacers of the rRNA genes of three species of theCynareae tribe:Cynara cardunculus subsp.scolymus (artichoke),Onopordum acanthium, andO. illyricum were cloned in the plasmid pGEM-7zf(+). Detailed restriction mapping and partial sequencing of the IGSs were carried out. The structural analysis showed a clear diversity betweenCynara andOnopordum, while a high degree of homology was found between the twoOnopordum spp. In all three species a fragment of about 450 bp from the 5 end of 18S to the Acc I site with a high sequence homology was present. Nucleotide sequences upstream from the above mentioned Acc I site show a gradual decrease of homology betweenCynara andOnopordum.     

Mouse polyomavirus mediated effects on the infected cell membrane studied by dielectric spectroscopy

The effect of polyomavirus on the infected cell has been investigated by dieclectric spectroscopy. This technique has a great potential in the study of the ion transport properties of the cell membrane. The results presented in this communication suggest a correlation between progression of the viral infection and dielectric features of the infected, cell plasma membrane.     

Pore formation by the sea anemone cytolysin equinatoxin II in red blood cells and model lipid membranes

Summary The interaction ofActinia equina equinatoxin II (EqT-II) with human red blood cells (HRBC) and with model lipid membranes was studied. It was found that HRBC hemolysis by EqT-II is the result of a colloid-osmotic shock caused by the opening of toxin-induced ionic pores. In fact, hemolysis can be prevented by osmotic protectants of adequate size. The functional radius of the lesion was estimated to be about 1.1 nm. EqT-II increased also the permeability of calcein-loaded lipid vesicles comprised of different phospholipids. The rate of permeabilization rised when sphingomyelin was introduced into the vesicles, but it was also a function of the pH of the medium, optimum activity being between pH 8 and 9; at pH 10 the toxin became markedly less potent. From the dose-dependence of the permeabilization it was inferred that EqT-II increases membrane permeability by forming oligomeric channels comprising several copies of the cytolysin monomer. The existence of such oligomers was directly demonstrated by chemical cross-linking. Addition of EqT-II to one side of a planar lipid membrane (PLM) increases the conductivity of the film in discrete steps of defined amplitude indicating the formation of cation-selective channels. The conductance of the channel is consistent with the estimated size of the lesion formed in HRBC. High pH and sphingomyelin promoted the interaction even in this system. Chemical modification of lysine residues or carboxyl groups of this protein changed the conductance, the ion selectivity and the current-voltage characteristic of the pore, suggesting that both these groups were present in its lumen.     

Induction of phagocytic behaviour in human epithelial cells by Escherichia coli cytotoxic necrotizing factor type1

Cytotoxic necrotizing factor type 1 (CNF1) from strains of pathogenic Escherichia coli induces in human epithelial HEp-2 cells, a profound reorganization of the actin cytoskeleton into prominent stress fibres and membrane ruffles. We report here that this process is associated with induction of phagocytic-like activity. CNF1-treated cells acquired the ability to ingest latex beads as well as non-invasive bacteria such as Listeria innocua, which were taken as a model system. Uptake of bacteria was similar to pathogen-induced phagocytosis, since L. innocua transformed with DNA coding for the pore-forming toxin listeriolysln O behaved, with respect to intracellular growth, like the invasive, pathogenic species L. monocytogenes. Our results raise the possibility that, in vivo, pathogenic CNF1 -producing E. coli may invade epithelia by this novel induced phagocytic-like mechanism.     

The oxygenase component of phenol hydroxylase from Acinetobacter radioresistens S13.

Phenol hydroxylase (PH) from Acinetobacter radioresistens S13 represents an example of multicomponent aromatic ring monooxygenase made up of three moieties: a reductase (PHR), an oxygenase (PHO) and a regulative component (PHI). The function of the oxygenase component (PHO), here characterized for the first time, is to bind molecular oxygen and catalyse the mono-hydroxylation of substrates (phenol, and with less efficiency, chloro- and methyl-phenol and naphthol). PHO was purified from extracts of A. radioresistens S13 cells and shown to be a dimer of 206 kDa. Each monomer is composed by three subunits: alpha (54 kDa), beta (38 kDa) and gamma (11 kDa). The gene encoding PHO alpha (named mopN) was cloned and sequenced and the corresponding amino acid sequence matched with that of functionally related oxygenases. By structural alignment with the catalytic subunits of methane monooxygenase (MMO) and alkene monooxygenase, we propose that PHO alpha contains the enzyme active site, harbouring a dinuclear iron centre Fe-O-Fe, as also suggested by spectral analysis. Conserved hydrophobic amino acids known to define the substrate recognition pocket, are also present in the alpha-subunit. The prevalence of alpha-helices (99.6%) as studied by CD confirmed the hypothized structural homologies between PHO and MMO. Three parameters (optimum ionic strength, temperature and pH) that affect kinetics of the overall phenol hydroxylase reaction were further analyzed with a fixed optimal PHR/PHI/PHO ratio of 2/1/1. The highest level of activity was evaluated between 0.075 and 0.1 m of ionic strength, the temperature dependence showed a maximum of activity at 24 degrees C and finally the pH for optimal activity was determined to be 7.5.     

Synthesis and biological activity of tachykinin analogs containing the adamantane moiety

Summary The adamantane moiety was introduced in the tachykinin NK₂ receptor-selective agonist [-Ala⁸]-NKA(4–10) (H-Asp-Ser-Phe-Val--Ala-Leu-Met-NH₂, MEN 10210) and in different positions of the NK₂ receptor antagonist MEN 10376 (H-Asp-Tyr-d-Trp-Val-d-Trp-d-Trp-Lys-NH₂) in order to investigate how this substitution affects their biological activity at tachykinin NK₁, NK₂ and NK₃ receptors. 1-Adamantaneacetic acid (1-Ada-CH₂COOH) was directly conjugated in the solid phase as the preformed OBt active ester to the N-terminal position of MEN 10210, obtaining MEN 10586 (1-Ada-CH₂CO-Asp-Ser-Phe-Val--Ala-Leu-Met-NH₂). The Pfp ester of adamantaneacetic acid (1) was prepared and used for the acylation of the N-terminal position of MEN 10376, yielding MEN 10606 (1-Ada-CH₂CO-Asp-Tyr-d-Trp-Val-d-Trp-d-Trp-Lys-NH₂). Compound 1 was then used to obtain the building block Fmoc-Lys(1-Ada-CH₂CO)-OH as a modified amino acid for the synthesis of MEN 10818 [H-Asp-Tyr-d-Trp-Val-d-Trp-d-Trp-Lys(1-Ada-CH₂CO)-NH₂]. In order to investigate the biological activity of the peptide bearing the adamantane group together with the free N-terminal amino function, we synthesised MEN 10676 [H-Asp(O-2-Ada)-Tyr-d-Trp-Val-d-Trp-d-Trp-Lys-NH₂] using Fmoc-Asp(O-2-Ada)-OH, in which 2-adamantanole was the protecting group of the aspartate -COOH moiety during the peptide synthesis and survived the final peptide cleavage and deprotection carried out under controlled conditions. MEN 10586 showed an agonist activity comparable to that of the parent compound MEN 10210 at NK₁ and NK₂ receptors of guinea pig ileum, rabbit isolated pulmonary artery and hamster isolated trachea preparations, while it showed a 25-fold higher agonist activity at NK₃ receptors of rat isolated portal vein. The three modified antagonist analogs displayed similar or reduced affinity at NK₁, NK₂ and NK₃ receptors as compared to MEN 10376. The drop was particularly evident (>2 log units) at the NK₂ receptors of the rabbit isolated pulmonay artery.