Guanosine 3':5'cyclic monophosphate and activators of guanylate cyclase inhibit secretagogue-induced corticotropin release by rat anterior pituitary cells

The secretion of corticotropin by perfused rat anterior pituitary cell columns was studied. Forty-one residue corticotropin releasing factor, vasopressin and high extracellular KC1 all stimulated the secretion of corticotropin. The hormonal response to corticotropin-releasing factor (10(-10) mol/l), vasopressin (10(-9) mol/l) as well as KC1 (48 mmol/l) was reduced by membrane permeant analogs of cGMP, such as 8-BrcGMP and dibutyryl-cGMP. The 8-BrcGMP analog (10(-5) mol/l) inhibited corticotropin release in response to corticotropin-releasing factor by 30%, that to vasopressin by 70%, and that to KCl by 50%. Atriopeptin1-28 (10(-8) and 10(-7) mol/l), a peptide known to activate membrane-bound guanylate cyclase in the anterior pituitary gland, decreased the release of corticotropin induced by vasopressin to about 30% of control. Similarly, activators of soluble guanylate cyclase, such as glyceryltrinitrate and sodium nitroprusside (10(-5) mol/l) inhibited vasopressin-stimulated corticotropin release by 60%. In conclusion, the data show that purported activators of particulate and soluble guanylate cyclase, as well as derivatives of cGMP itself are strong inhibitors of secretagogue-induced corticotropin release by corticotroph cells of the anterior pituitary gland.     

Effects of the kappa opioid receptor antagonist MR-2266-BS on the acquisition of ethanol preference

Using a paradigm by which rats forced to drink a weak ethanol solution (2.5% w/v) (conditioning session) develop ethanol preference in consecutive retention testing days, the effects of the administration of the kappa opioid antagonist MR-2266-BS, prior to or after the forced ethanol session, were studied. Pre-conditioning subcutaneous (s.c.) administration of 1 mg/kg of MR-2266-BS induced a decrease in subsequent ethanol consumption without significantly modifying the acquisition of ethanol preference. Post-conditioning administration of MR-2266-BS (0.1, 1, 5 or 10 mg/kg) induced both a dose-dependent reduction in ethanol consumption and in preference throughout the three following days. The results of the present study provide further support of the involvement of kappa-type opioids on drinking behavior, and suggest that kappa receptors may be involved in the consumption and development of preference to ethanol.     

Corticosteroidogenesis modulation by beta-endorphin and dynorphin1-17 in isolated rat adrenocortical cells

Two opioid peptides, beta-endorphin and dynorphin1-17 were bioassayed with isolated rat adrenocortical cells. beta-Endorphin increases basal production of corticosterone as well as the adrenal responsiveness to low doses of ACTH, these effects being partially reversed by naloxone. Dynorphin1-17, without affecting basal corticosterone synthesis, increases adrenocortical responsiveness to ACTH; naloxone does not influence this effect. It is suggested that peripheral opioid peptides may participate in the maintenance of the homeostatic balance by modulating adrenal corticosteroidogenesis.     

Exposure to T‐Cycles of 22 and 23 h during Lactation Modifies the Later Dissociation of Motor Activity and Temperature Circadian Rhythms in Rats

Early environmental conditions may affect the development and manifestation of circadian rhythms. This study sought to determine whether the maintenance of rats under different T‐cycles during lactation influences the subsequent degree of dissociation of the circadian rhythms of motor activity and core body temperature. Two groups of 22 day‐old Wistar rats were kept after weaning under T‐cycles of 22 h (T22) or 23 h (T23) for 70 days. Subsequently, they were kept in constant darkness (DD). Half of the animals in each group were born and reared under these experimental conditions, while the other half were reared until weaning under 24 h LD cycles (T24). Rats transferred from T24 to T22 or T23 showed two circadian components in motor activity and temperature, one entrained by light and the other free‐running. In T22, there was also desynchronization between temperature and motor activity. Rats submitted to T23 from birth showed higher stability of the 23 h component than rats transferred from T24 to T23 after weaning. However, in comparison to rats born under T24 and subsequently changed to T22, animals submitted to T22 from birth showed shorter values of the period of the non‐light‐dependent component during T22, more aftereffects when transferred to DD, and a lack of desynchronization between motor activity and temperature. The results suggest that T‐cycles in the early environment may modify overt rhythms by altering the internal coupling of the circadian pacemaker.     

Effects of Mediterranean climate annual variability on stream biological quality assessment using macroinvertebrate communities

Data on macroinvertebrates of selected reference sites were compiled from a long-term monitoring programme carried out in the Mediterranean Catalan Basins (NE Spain) that permitted analysis for nine years, from 1996 to 2004, using a homogeneous data collection procedure. This study aims to analyse the differences in composition and structure of macroinvertebrate communities at family level in five Mediterranean river types, and the values of biological quality metrics (IBMWP and IASPT indices, taxon richness and EPT) in reference conditions. Also differences between seasons (spring vs. summer) and between dry and wet periods were analysed. The dry and wet periods were determined using the Standardised Precipitation Index (SPI). A total of 29 reference sites were selected out of 184 sampling sites analysed, and 171 reference samples were available (from 1996 to 2004), of which 88 were sampled in dry periods, whereas 83 correspond to wet periods. Differences on community composition at family level were appreciated, clustering the rivers in three different groups: (1) rivers with a continuous flow regime located in siliceous zones; (2) rivers with a continuous flow regime located in calcareous zones; and (3) temporary rivers regardless of geology. Moreover, our results explain that the characteristics of hydrological periods (dry and wet) characterize the differences between communities better than just the season. The analysis of four biological quality metrics reveals clear differences between values obtained from dry and wet periods concerning taxon richness, EPT values and IBMWP biological indices, whereas the IASPT index does not show significant differences. The median taxonomic richness in wet periods is 32 macroinvertebrate families per sample while in dry periods this value falls to 22. Reference values of IBMWP index, the total number of taxa, and EPT metric are different between dry and wet periods in spring samples, while these differences are not relevant for IASPT index except for temporary streams. Hydrological specific conditions should additionally be considered in order to better calculate biological reference conditions, and to properly apply biological quality metrics used to establish the ecological status in Mediterranean rivers, especially in temporary ones. The use of the dry–wet period classification according to the climate characteristic results is a more accurate application of the Water Framework Directive in Mediterranean rivers. Implications of future climate change should be also considered from our results.     

Structural-Functional Analysis Reveals a Specific Domain Organization in Family GH20 Hexosaminidases

Hexosaminidases are involved in important biological processes catalyzing the hydrolysis of N-acetyl-hexosaminyl residues in glycosaminoglycans and glycoconjugates. The GH20 enzymes present diverse domain organizations for which we propose two minimal model architectures: Model A containing at least a non-catalytic GH20b domain and the catalytic one (GH20) always accompanied with an extra α-helix (GH20b-GH20-α), and Model B with only the catalytic GH20 domain. The large Bifidobacterium bifidum lacto-N-biosidase was used as a model protein to evaluate the minimal functional unit due to its interest and structural complexity. By expressing different truncated forms of this enzyme, we show that Model A architectures cannot be reduced to Model B. In particular, there are two structural requirements general to GH20 enzymes with Model A architecture. First, the non-catalytic domain GH20b at the N-terminus of the catalytic GH20 domain is required for expression and seems to stabilize it. Second, the substrate-binding cavity at the GH20 domain always involves a remote element provided by a long loop from the catalytic domain itself or, when this loop is short, by an element from another domain of the multidomain structure or from the dimeric partner. Particularly, the lacto-N-biosidase requires GH20b and the lectin-like domain at the N- and C-termini of the catalytic GH20 domain to be fully soluble and functional. The lectin domain provides this remote element to the active site. We demonstrate restoration of activity of the inactive GH20b-GH20-α construct (model A architecture) by a complementation assay with the lectin-like domain. The engineering of minimal functional units of multidomain GH20 enzymes must consider these structural requirements.     

Regulation of cytokines, cytokine inhibitors, and acute-phase proteins following anti-TNF-alpha therapy in rheumatoid arthritis.

Treatment with a chimeric mAb to TNF-alpha has been shown to suppress inflammation and improve patient well-being in rheumatoid arthritis (RA), but the mechanisms of action of such treatment have not been fully explored. Here we show that in vivo administration of anti-TNF-alpha Ab, using a longitudinal analysis, results in the rapid down-regulation of a spectrum of cytokines, cytokine inhibitors, and acute-phase proteins. Marked diurnal variation in the serum levels of some of these were detected. These results were consistent with the concept of a cytokine-dependent cytokine cascade, and the degree of clinical benefit noted after anti-TNF-alpha therapy is probably due to the reduction in many proinflammatory mediators apart from TNF-alpha, such as IL-6, which reached normal levels within 24 h. Serum levels of cytokine inhibitors such as soluble p75 and p55 TNFR were reduced as was IL-1 receptor antagonist. Reductions in acute-phase proteins occurred after serum IL-6 fell and included serum amyloid A, haptoglobin, and fibrinogen. The latter reduction could be of importance, as it is a risk factor for atherosclerosis, which is augmented in RA patients.     

On the study of evolution in species-level lineages in the fossil record: controlled methodological sloppiness

The focus of this discussion is on a research strategy to be employed in the paleontological study of evolution in species-level lineages. Such studies must begin with the identification of phena in a single sample, then trace their presumed ancestors and/or descendants through time along a single section, apply theoretical models to establish the most appropriate pattern in quantitative terms, and finally compare the pattern and its timing to patterns observed in presumably conspecific lineages over a substantial geographic area. Only thereafter can evolution be recognized as a plausbile explanation for paleontological observations. Decisions made at all these steps involve, however, more or less arbitrary judgements. Such studies, therefore, cannot be conducted with full methodological rigor. It is suggested that ‘controlled methodological sloppiness’ provides the best solution to the problem, and that it should be based on hermeneutic analysis.     

Structure of the O-polysaccharide from the lipopolysaccharide of Providencia alcalifaciens O28

An O-polysaccharide and oligosaccharides were isolated by GPC following mild acid degradation of the lipopolysaccharide of Providencia alcalifaciens O28. The O-polysaccharide was studied by sugar and methylation analyses, ¹H and ¹³C NMR spectroscopy, including 2D ROESY and H-detected ¹H,¹³C HSQC and HMBC experiments, and the following structure of the branched pentasaccharide repeating unit was established:This structure was confirmed by ESI MS of the isolated tridecasaccharide consisting of the lipopolysaccharide core and one O-polysaccharide repeat. The ESI mass spectrum also enabled inferring the composition of the core oligosaccharide.